Purification Protocols

来自 : 发布时间：2024-09-20

GE-HEALTHCARE Handbook Protein Purification(pdf) GE-HEALTHCARE Strategies for Protein Purif ication Handbook (pdf) GE-HEALTHCAREProtein puri cation Applications (pdf) GE-HEALTHCARE Protein and Peptide Purification - Technique selection guide (pdf) GE-HEALTHCAREDOE: Design of Experiments in Protein Production and Purification Handbook (pdf) GE-HEALTHCARE Bio Process - Product guide(pdf)GE-HEALTHCARE Purifying Challenging Proteins: Membrane proteins, Multiprotein complexes and Inclusion bodies (pdf) GE-HEALTHCAREKTAdesign Purification Method handbook. This handbook is intended to help you make full use of your KTAexplorer or KTApurifier chromatography system.The handbook is a collection of useful step-by-step protocols to aid your everyday purification work. Detailed instructions and recommendations are presented in a straightforward format, which should be easy to follow without needing a high level of expertise in programming or in chromatography. (pdf)GE-HEALTHCARE Purification of tagged proteins(pdf)GE-HEALTHCARE Packing of Gel Filtration column (movie)GE-HEALTHCARE Packing of IEX, Affinity or HIC column (movie) GE-HEALTHCARE Column evaluation (movie) GE-HEALTHCARE PreDictor plates for High-throughput process development (HTPD). Disposable 96-well filter plates prefilled with GE Healthcare BioProcesschromatography media (Capto Q, S, DEAE, MMC, Adhere; Q and SP Sepharose MabSelect , SuRe and Xtra AIEX and CIEX screening plate. SupportHTPD by allowing parallel screening of chromatographic conditions. They can be used in automated workflows using robotic systems,or can be operatedmanually using multi-channel pipettes(pdf)(pdf Handbook) GE-HEALTHCARE Two-step purification with KTA pure, using loop collection (pdfI) (pdfII) Versatile modules enable automated multi-column puri cations on the KTA pure chromatography system Bastian Franke et. al. Journal of Chromatography A (2020), https://doi.org/10.1016/j.chroma.2019.460846 (pdf) Development of BioRad NGC and GE KTA pure systems for highly automated three column protein purification employing tandem affinity, buffer exchange and size exclusion chromatography Dwight Winters et. al. Protein Expression and Purification 165 (2020) 105497 https://doi.org/10.1016/j.pep.2019.105497 (pdf) BIA Separations CIM Monolithic Columns based on CIM Convective Interaction Media Technology; suitable for purification of large biomolecules such asviruses(viral vectors and vaccines), DNA (plasmid DNA) and larger proteins (Immunoglobulins G and M, pegylated proteins). CIM Monolithic Columns exhibitunrivaledcharacteristics in terms of operational flow rates, binding capacity and separation resolution for large biomolecules. Products are used in research,laboratory, pilot andindustrial production stages and are extremely simple to use, with no packing of columns needed(pdf-I) (pdf-II) (pdf-III) BIO RAD Sample Preparation Handbook(pdf) CLONTECH:Troubleshooting Guide (pdf) MERCKCleaning and Regeneraton of Fractogel EMD sorbents(pdf) MERCK Chromolith HighResolution columns allows you to enjoy greater flexibility of flow rates, faster analyses, and a longer column lifetime. Each Chromolith column consists of a single rod of high-purity polymeric silica gel with a bimodal pore structure of macro and mesopores. The macropores reduce column back pressure, allowing significantly faster flow rates. The mesopores form a fine porous structure, which creates a very large active surface area for high-efficiency separations. For even greater efficiency, multiple Chromolith columns can be coupled to achieve a higher theoretical plate count with still very low back pressure. (pdf) Chromolith HighResolution vs. Core-Shell: Approx. 5 times higher sample capacity. 2 to 3 times faster analysis. Substantially lower carry-over effect. Longer column lifetime. Same efficiency as sub-3 m core-shell columns. Similar selectivity NOVAGENBenzonase Nuclease - Removal of Nucleic Acids(pdf) NOVAGENBuffer Compositin for Fusion Tag Affinity Purification (pdf) PallChromatography Media Selection Guide(pdf)Protein Sample Preparation and Analysis Application Manual. Table of contents(pdf)1.0 Introduction: 1.1 Proteomics Process Flo -. 1.2 Product Selection (pdf1) 2.0 Pre-Analytical Applications: 2.1 Abundant Protein Removal - 2.2 Protein Fractionation - 2.3 Detergent Removal - 2.4 Concentration, Desalting, and Buffer Exchange - 2.5 Generic Clarification of Samples by Microporous Filtration (Particulate Removal) - 2.6 Endotoxin Removal (pdf2)3.0 Analysis: 3.1 Western Blotting - 3.2 Affinity Activated or Activatable Membranes (pdf3)4.0 Purification Applications: 4.1 Mustang Ion Exchange Membranes - 4.2 BioSepra Chromatography Resins - 4.3 Rapid Purification Development/Scouting (Process Proteomics) - 4.4 Buffer Exchange, Desalting, and Concentration - 4.5 Final Product Clarification (pdf4) 5.0 Supporting Technologies: 5.1 Water Supply - 5.2 Microbiology Testing(pdf5)6.0 Technical Appendices: 6.1 Evaluation Chromatography Column Packing Efficiency - 6.2 Optional Pre-Treatment to Improve Recovery of Samples from Ultrafiltration Spin Filters - 6.3 Vacuum Manifold for Use with Multi-Well Filter Plates - 6.4 General Filtration Procedures for AcroPrep and AcroWell Filter Plates - 6.5 Chromatography Products Selection Guide - 6.6 BioSepra Media - 6.7 Protein Sample Preparation and Analysis Media - 6.8 Chemical Compatibility Guide (pdf6)THERMO:Peptide solubility guidelines. Use amino acid characteristics to predict hydrophobicity. (pdf)TOSOH Chromatographic Media Catalog(pdf)TOSOHTOYOPEARL Orientation Sheet: Size Exclusion Chromatography, Ion Exchange Chromatography, Hydrophobic Interaction Chromatography and Affinity Chromatography (pdf) (pdf-II)(pdf-III)TOSOHGeneral Principles of Chromatography. Poster(pdf) Removal of Nucleic Acids - Various Protocols Test TubeRemoval of Chaperonins: Bacterial expression and purification of Interleukin-2 Tyrosine kinase: Single step separation of the chaperonin impurit. Raji E. Joseph, Amy H. Andreotti Protein Expression and Purification 60 (2008) 194 197(pdColumns DIBA industries Omnifit Labware Brochure Chromatography Columns and Bottle Caps (pdf)GE-HEALTHCARE columns are designed for standard liquid chromatography of macromolecules. XK columns (pdf) Tricorn (pdf) C-series columns (pdf)HR columns (pdf)HiScale columns (pdf) GE-HEALTHCARE Packing of Gel Filtration column (movie)GE-HEALTHCARE Packing of IEX, Affinity or HIC column (movie) GE-HEALTHCARE Column evaluation (movie) GE-HEALTHCARE Protein purification troubleshooting guide. This guide helps you quickly identify root causes for many of the common issues encountered during protein purification. It gives practical advice on small changes that you can make to fix these problems (pdf) Essential Life Solutions:SNAP Laboratory Glass Columns for high-performance preparative chromatography (pdf-literature) (pdf-tech manual) (video) PALLPall LRC Columns Chromatography Columns for Laboratory Applications (pdf) User Guide (pdf-II) SEPAX Generik FPLC Empty Column. Versatile glass empty column ideal for FPLC purification. High pressure tolerance - up to 6MPa (60bar) Wide range of ID 7 IDs (6.6 - 50mm) with various lengths (pdf) Purification of Recombinant ProteinsStructural Genomics Consortium Protein production and purification. Nature Methods 2008, 5: 135-146 (pdf-I) (pdf-II)GE-HEALTHCARE The Recombinant Protein Handbook (pdf-I) (pdf-II) (new pdf) GE-HEALTHCARE Purification of tagged proteins(pdf)PIERCE:Fusion Protein Fusion Protein Products(pdf) SIGMA:Detection and Purification Selection Guide (pdf-I) (pdf-II) GST-Fusion ProteinsSmall scale GST-fusion protein purification under nature conditions ABT :Glutathione Agarose resins for use in affinity purification of Glutathione-S-transferase (GST) and GST tagged fusion proteins. Bulk and Cartridges formats suitable for MPLC, FPLC , KTA design, batch, gravity, peristaltic pump syringe (pdf-I)(pdf-II) (pdf-III) Troubleshooting (pdf-IV) (pdf-brochure 2015)CLONTECH: Glutathione Resins (pdf) CLONTECH: GST products Handbook (pdf)CLONTECH: Protein Purification Products (pdf) G-BioscienceGlutathione Resin, for the single-step affinity purification of proteins with a glutathione S-transferase (GST) tag (pdf) GE-HEALTHCARE GST fusion protein Handbook(pdf) GE-HEALTHCARE Purification of tagged proteins(pdf)GE-HEALTHCARE Glutathione Sepharose High Performance is an affinity chromatography medium designed for easy, one-step purification of glutathione S-transferase (GST) tagged proteins (pdf)GE-HEALTHCARE Separation of DnaK from pGEX-GST by ion exchange chromatography (pdf) JENAGlutathione ChroMatrix , Fast Flow and GST Cleavage Capture Kit(pdf)NOVAGENGST.Bind Kits(pdf)NOVAGEN: His-Tag GST-Tag purification and Detection tools (pdf)NOVAGEN: Protein Purification and Detection. GST Tag Fusions (pg.30-36)(pdf) poly His-Tagged Fusion Proteins Small an large scale His-Tag fusion protein purification under nature conditions Small scale His-Tagfusion protein purification under denaturative conditions Protein Refolding on IMAC resin - Batch Screening Procedure - On-Column Scale-up ABT offers affinity tag chelating resins for purifications of his-tag proteins by Immobilized Metal Affinity Chromatography (IMAC).Selection of metal chelating beads ( 4 different metals). Two different degrees of loading ( high and low density). Three differentformats :bulk resins, pre-packed columns and cartridges. (pdf) (pdf-brochure 2015) Procedure for using chelating agarose beads test kits (pdf-I) . Troubleshooting (pdf-II)ACTIVE MOTIVE: Ni-Ted Manual(pdf) AFFILAND: Metal PurificationBioRad Profinity IMAC Resins (pdf)BioToolomics: IMAC SepFast BG Media (pdf-I) Super Spin columns instructions (pdf-II) CLONTECH: BD TalonTM Metal Affinity Resins User Manual(pdf) (pdf-II) CLONTECH: Talon Resin Protocol (pdf)CLONTECH: Talon Products (pdf) CLONTECH:HisTALON Superflow 1 ml 5 ml Cartridges User Manual (pdf) CLONTECH: TALON spin Columns Protocol (pdf) CLONTECH: Purification in the presence of Beta Mercaptoethanol (pdf) CLONTECH: Protein Purification Products (pdf)CLONTECH: BD HAT Protein Expression and Purification System User Manual (pdf)CLONTECH:Capturem His-Tagged Purification Miniprep Kit. The columns are suitable for use under native or denaturing conditions, in the presence of additives such as DTT (up to 10 mM), ME (up to 30 mM), TCEP (up to 5 mM), EDTA (up to 10 mM) (pdf) DYNAL Dynabeads TalonTM (pdf) G-BioscienceNickel Chelating Resin: A Ni-IDA IMAC resin for 6X-His Tagged Protein Purification (pdf)GE-HEALTHCARE Purification of tagged proteins(pdf) GE-HEALTHCAREHi Trap Chelating (pdf) GE-HEALTHCAREHi Trap Chelating Application Note(pdf) GE-HEALTHCAREChelating Sepharose Fast Flow Instructions(pdf) GE-HEALTHCAREHis GraviTrap prepacked, single-use column for purification of histidine-tagged proteins by immobilized metal affinity chromatography (IMAC) without any need for a purification system. Negligible nickel leakage and is compatible with denaturing andreducing agents as well as a wide range of additives. Short purification times. High protein binding capacity, 40 mg/column. Purifies unclarified samples (pdf) GE-HEALTHCAREHis SpinTrap is a prepacked, single-use spin column for purifying histidine-tagged proteins by immobilized metal ion affinity chromatography (IMAC). The resin has high protein binding capacity, low nickel ion (Ni2+) leakage, and excellent compatibility with denaturing agents plus a wide range of additives. His SpinTrap is used with a standard microcentrifuge and one purification run takes approx. 10 min. His SpinTrap columns allow direct purification of unclarified cell lysates.(pdf-I)(pdf-II) GE-HEALTHCARE HisTrap FF crude. This pre-prepacked column is intended for preparative puri cation of histidine-tagged recombinant proteins from unclari ed lysate without precentrifugation and ltration of the sample. Leakage of Ni2+ is very low. The medium is compatible with all commonly used aqueous buffers, reducing agents, denaturants such as 6 M Gua-HCland 8 M urea, and a range of other additives(pdf)GE-HEALTHCARENi Sepharose and IMAC Sepharose selection guide (pdf)GE-HEALTHCARE IMAC Sepharose High Performance. Possible to charge with various metal ions for optimized selectivity.High protein binding capacity. Compatible with commonly used reducing agents, such as DTT, DTE, TCEP, and -mercaptoethanol.Stable in a wide range of denaturants, detergents, and buffer systems. (pdf) GE-HEALTHCARENi Sepharose High Performance (pdf)(pdf-II) GE-HEALTHCARE Article: Addition of imidazole during binding improves purity of histidine-tagged proteins (pdf)GE-HEALTHCARE Article: Snags with tags: Some observations made with (His)6-tagged proteins. The need to remove or omit the affinity tag from the purification scheme.P.Ramage et.al (pdf)IBATools for protein expression purification. 6xHis-tag Ni-NTA technology: The optimal partner for Strep-tag indouble tag proteins (pdf)IBA: Expression and purification of proteins using Strep-tag and/or 6xHistidine-tag. A comprehensive manual. (pdf) IBA Mammalian expression and purification system using Strep-tag and/or 6xHistidine-tag (pdf)JENA6 % CL-Nickel ChroMatrix - Cobalt ChroMatrix , Fast Flow and Nickel, Zinc, Cobalt, Copper and Metal Free Agaroses,Gravity Flow (pdf) MERCK-NOVAGEN: Metal Chelate Affinity Chromatography. Ni-MAC , Co-MAC and u-MAC Metal Affinity Chromatography (MAC) Resins and Cartridges(pdf) (pdf-II)(pdf-III) NOVAGEN: pET purification manual(pdf)(pdf-2) NOVAGEN: Ni-NTA Hi-Bind Resins Protocols (pdf) NOVAGEN: Protein Purification and Detection. His Tag Fusions (pg.18-29)(pdf) NOVAGEN:His-Tag GST-Tag purification and Detection tools (pdf) NUNC: ProPur affinity spin columns contain a unique flow regulator, FlowGo, which maximizes the contact time between the target protein and the resin. ProPur columns offer the speed of a spin column with the yields and purities of a gravity column or a batch protocol. They are idel for rapid small-scale and pilot purifications of His-tagged proteins, method development, expression trials, solubility determination and separation of the proteolytically cleaved His tag from the purified protein. (pdf-I) THERMO: HisGrab Nickel Coated 96-Well Plates. Ideal for analyzing polyhistidine-tagged fusion proteins by ELISA-based methods. Can be used with colorimetric, chemiluminescent or fluorescent detection methods. (pdf) PROMEGA: MagneHisTM Protein Purification System(pdf) QIAGEN: Ni-NTA purification manual (pdf) QIAGEN: NiNTA cell lysis under nature conditions (pdf) QIAGEN:Ni-NTA Superflow Cartridges are pre-filled with 1 ml or 5 ml Ni-NTA Superflow and are ready to use for purification of 6xHis-tagged proteins using a syringe, peristaltic pump, or liquid chromatography system (pdf) ROCHEcOmplete His-Tag Purification Resin. Nickel-chelate chemistry that minimizes nickel ion leakage, and compatible with commonly usedreducing agents (such as DTT), chelating metalloprotease inhibitors(such as EDTA), and a wide range of buffer substances and salt conditions. (pdf-I) (pdf-II) SARTORIUS Sartobind IDA 75 - A Separation Technology Based on Metal Chelate Membrane Adsorbers - Operating Instructions(pdf)SIGMA: His Selected Nickel Affinity Gel(pdf) VIVASCIENCE: Vivapure Metal Chelate Mini SpinColumns(pdf-I)(pdf-II)(pdf-III) (pdf-IV)(pdf-V)(pdf-VI)(pdf-VII)(pdf-VIII) Maltose Binding Protein GE-HEALTHCAREMBPTrap HP is a ready to use HiTrap column for purifying recombinant proteins tagged with maltose binding protein (MBP). The column is packed with Dextrin Sepharose High Performance based on a 34 m beadsized matrix. Purification of MBP-tagged protein is done under physiological conditions, which together with mild elution by maltose, preserves the activity of the target protein.(pdf-I) (pdf-II) GE-HEALTHCARE Purification of tagged proteins(pdf) NECTAGEN MBP purification by affinity chromatography with nanoCLAMP malE-A1resin (CLostridial Antibody Mimetic Proteins) affinity reagents are recombinant 15 kD antibody mimetic proteins selected for tight, selective and gently reversible binding. The nanoCLAMPs can be used to pull down proteins or protein complexes from whole cell extracts followed by elution with propylene glycol under conditions that preserve activity and protein-protein interactions. The nanoCLAMPtechnology is described in the Protein Expression and Purificationarticle \"Development of polyol-responsive antibody mimetics for single-step protein purification\" by Suderman et al. 2017. NEW ENGLAND BIOLAB: pMAL protein Fusion and Purification System (pdf-I)(pdf-II)PALLMixed-mode resins HEA HyperCel and PPA HyperCel can be used for MBP purification(pdf) Small and large scale MBP-fusion protein purificationD.R.Smyth et.al.: Crystal Structures of Fusion Proteins with Large-Affinity tags. Protein Science (2003), 12: 1313-1322(pdf) Calmodulin Binding Peptide G-BioscienceCalmodulin Resin: isolation of recombinant proteins that are fused to the calmodulin-binding peptide (CBP) (pdf) STRATAGENE: CBP Calmodulin-Binding Peptide Affinity Tag System(pdf-I)(pdf-II)(pdf-III) Intein Mediated Purification NEW ENGLAND BIOLAB:The IMPACT (Intein Mediated Purification with an Affinity Chitin-binding Tag) system is a novel protein purification system which utilizes the inducible self-cleavage activity of protein splicing elements (termed inteins) to separate the target proteinfrom the affinity tag. Each intein tag contains a chitin binding domain (CBD) for the affinity purification of the fusion protein on a chitin resin.Induction of on-column cleavage, using thiol reagents such as dithiothreitol (DTT), releases the target protein from the intein tag. Able to produce target protein without vector derived amino acids (pdf)NEW ENGLAND BIOLAB: INTEIN-TM System(pdf) NEW ENGLAND BIOLAB: INTEIN-TWIN System(pdf) T7.Tag NOVAGENT7.Tag Affinity Purification kit(pdf) (pdf-II pg 45-46)Cellulose Binding Domain NOVAGEN CBIND Kits (pdf) NusA Protein NOVAGENExpression of Soluble Heterologous Proteins via Fusion with NusA Protein - Article (pdf) NECTAGEN nanoCLAMP nusA-A1(Resin) (CLostridial Antibody Mimetic Proteins) affinity reagents are recombinant 15 kD antibody mimetic proteins selected for tight, selective and gently reversible binding. The nanoCLAMPs can be used to pull down proteins or protein complexes from whole cell extracts followed by elution with propylene glycol under conditions that preserve activity and protein-protein interactions. The nanoCLAMPtechnology is described in the Protein Expression and Purificationarticle \"Development of polyol-responsive antibody mimetics for single-step protein purification\" bySuderman et al. 2017. Biotinylated Fusion Protein - AviTag PROMEGAPinPointTM Xa Protein Purification System The System is designed for the production and purification of fusion proteins that are biotinylated in vivo. Biotinylated fusion proteins are produced in E. coli and are affinity-purified using the SoftLink Soft Release Avidin Resin. This proprietary resin allows elution of the fusion protein under nondenaturing conditions. The PinPoint Vectors feature the encoded endoproteinase Factor Xa proteolytic site that provides a way to separate the purification tag from the native protein. (pdf) GeneCopoeia TheAVI Tagis a commercially developed 15-amino acid peptidetagconsisting of GLNDIFEAQKIEWHE. This unique peptide is readily and specifically biotinylated by the E. coli biotin ligase, BirA, andAvi-tagged proteins can be detected or purified by avidin or streptavidin. Fused to your protein, theAviTag provides a multi-functional system useful for many applications including: Expression. Imaging. The AviTag technology is based on the biotinylation of AviTag by biotin ligase in vitro or in vivo and on the specific and reversible binding of avidin or streptavidin to biotin for immobilizing, purifying and visualizing proteins. FLAG and Anti HA GenScript DYKDDDDK or FLAG can be fused to the N-terminus or C-terminus of a target protein to facilitate detection and purification. Monoclonal Anti-DYKDDDDK G1 Affinity Resin (pdf) SIGMA:Detection and Purification - FLAG A Proven System for Detection and Purification of Proteins. (pdf) Anti-HA Agarose (pdf)THERMO: HA-epitope tag (YPYDVPDYA) derived from the human influenza hemagglutinin (HA) protein. (pdf) Strep-Tag GE-HEALTHCAREStrepTactin Sepharose High Performance is a chromatography medium for purifying Strep(II)-tagged proteins. Purification is done under physiological conditions and mild elution preserves the activity of the target protein(pdf) GE-HEALTHCARE Purification of tagged proteins(pdf)IBA: Expression and purification of proteins using Strep-tag and/or 6xHistidine-tag. A comprehensive manual. The Strep-tag II is a short peptide (8 amino acids, WSHPQFEK), which binds with high selectivity to Strep-Tactin, an engineered streptavidin. Elution of purified recombinant protein is performed by desthiobiotin. (pdf) IBATools for protein expression purification. Strep-tag technology and 6xHis-tag Ni-NTA technology: Double tag protein expression and purification (pdf) IBA Mammalian expression and purification systemusing Strep-tag and/or 6xHistidine-tag (pdf)IBA One-STrEP Kit. One-STrEP-tag Purification for the Isolation and Identification of Protein Complexes in Mammalian Cells. A comprehensive manual (pdf)IBA His-STREPPER: Adapter molecule for fast and easy conversion of 6xHistidine-tag fusion protein to Strep-tag II fusion proteins (pdf) NOVAGENStrep Tactin Purification Kits. Strep Tactin protein is a streptavidin derivative developed for optimal Strep Tag II binding. The binding affinity of Strep Tag II for Strep Tactin is approximately 100 times higher than for streptavidin. The purified target protein is competitively eluted with 2.5 mM desthiobiotin, an analog of biotin that reversibly binds Strep Tactin.(pdf-I) (pdf-II pg43-44)(pdf-III)QIAGEN:Two-Step Affinity Purification System Handbook - For expressing, purifying, and detecting proteins carrying a 6xHis and Strep-tag II. After elution from the Ni-NTA matrix using imidazole, recombinant proteins (which also carry the Strep-tag II epitope) are loaded directly onto a Strep-Tactin matrix and eluted using eitherbiotin or desthiobiotin.(pdf-I) (pdf-II)STRATAGENE:VariFlex bacterial protein expression system. Include three different solubility enhancement tags (SETs) which are designed to increase protein solubility, the streptavidin binding peptide (SBP) purification tag, and a tag that allows for rapid soluble protein quantification (Q-tag)(pdf) Fluorescent Protein BD BIOSCIENCES BD Living Colors AcGFP1 Fluorescent Protein. A novel monomeric green fluorescent protein for fusion tag applications. Ideal for multicolor applications in flow cytometry and fluorescence microscopy. (pdf) BD BIOSCIENCES BD Living Colors DsRed-Monomer Fluorescent Protein. A novel monomeric red fluorescent protein for fusion tag applications. Ideal for multicolor applications in flow cytometry and fluorescence microscopy. (pdf) Halo tag fusion PROMEGA:The HaloTag Interchangeable Labeling Technology is a novel tool for imaging live or fixed mammalian cells that express the HaloTag Protein or protein fusions, analyzing post-translational modification of labeled fusion proteins, and isolating proteins and protein complexes. The technology is based on efficient formation of a covalent bond between a specially designed reporter protein encoded by the HaloTag pHT2 Vector and a specific ligand in living cells, in solution or on a solid support. The HaloTag Ligand can carry a variety of functionalities, including fluorescent labels, affinity handles and attachments to a solid phase. The covalent bond forms rapidly under general physiological conditions, is highly specific and essentially irreversible, yielding a complex that is stable even under stringent conditions. The open architecture of the technology enables use of different ligands. Technical Manual. (pdf) PROMEGA: HaloLink Resin. The HaloTag technology comprises the HaloTag polypeptide, which can be fused to a protein of interest using the HaloTag Vectors, and a system of interchangeable synthetic ligands that covalently bind to the HaloTag polypeptide. HaloLink Resin, allows covalent and oriented surface immobilization of HaloTag fusion proteins. The HaloLink Resin can be used in a variety of applications including enzyme immobilization, detection of protein:protein interactions and purification of fusion proteins using protease cleavage. (pdf) Solubility enhancement tags (SETs)STRATAGENE:VariFlex bacterial protein expression system. Include three different solubility enhancement tags (SETs) which are designed to increase protein solubility, the streptavidin binding peptide (SBP) purification tag, and a tag that allows for rapid soluble protein quantification (Q-tag)(pdf)SUMO INVITROGEN:The Champion pET SUMO Protein Expression System utilizes a small ubiquitinlike modifier (SUMO) to allow expression, purification, and generation of native proteins in E. coli. SUMO fusions may increase the expression of recombinant proteins and enhance the solubilityof partially insoluble proteins. In addition, the tertiary structure of the SUMO protein is specifically recognized and cleaved by a ubiquitin-like protein-processing enzyme, SUMO Protease resulting in the production of native protein. (pdf) LIFESENSORS SUMO Gene Fusion Technology (pdf) LIFESENSORS SUMO Gene Fusion Technology - New Methods for Enhancing Protein Expression and Purification in Bacteria(pdf-I)(pdf-II) (pdf-III) LIFESENSORS SUMO Gene Fusion Technology - New Methods for Enhancing Protein Expression and Purification (intracellular and secretory expression) in Insect cells (pdf)LIFESENSORS SUMO Gene Fusion Technology - New Methods for Enhancing Protein Expression and Purification (intracellular and secretory expression) in Mammalian cells (pdf) LIFESENSORS SUMO Gene Fusion Technology - New Methods for Enhancing Protein Expression and Purification in Yeast cells: Saccharomyces (pdf) and Pichia (pdf)LIFESENSORS SUMO Gene Fusion Technology - New Methods for Rapid Recombinant Peptide Expression and Purification (pdf) NECTAGEN SUMO purification by affinity chromatography with nanoCLAMP SMT3-A1 (CLostridial Antibody Mimetic Proteins) affinity reagents are recombinant 15 kD antibody mimetic proteins selected for tight, selective and gently reversible binding. The nanoCLAMPs can be used to pull down proteins or protein complexes from whole cell extracts followed by elution with propylene glycol under conditions that preserve activity and protein-protein interactions. The nanoCLAMPtechnology is described in the Protein Expression and Purificationarticle \"Development of polyol-responsive antibody mimetics for single-step protein purification\" by Suderman et al. 2017.THIOREDOXIN NECTAGEN Thioredoxin purification by affinity chromatography with nanoCLAMP trxA-A1resin (CLostridial Antibody Mimetic Proteins) affinity reagents are recombinant 15 kD antibody mimetic proteins selected for tight, selective and gently reversible binding. The nanoCLAMPs can be used to pull down proteins or protein complexes from whole cell extracts followed by elution with propylene glycol under conditions that preserve activity and protein-protein interactions. The nanoCLAMPtechnology is described in the Protein Expression and Purificationarticle \"Development of polyol-responsive antibody mimetics for single-step protein purification\" by Suderman et al. 2017. Cold Expression CLONTECH: pCold TF DNA Vector is a fusion cold shock expression vector that expresses Trigger Factor (TF) chaperone as a soluble tag (48 kDa) which facilitates co-translational folding of newly expressed polypeptides. The vector has a His-Tag sequence, and a multicloning site (MCS). A lac operator is inserted downstream of the cspA promoter to ensure strict regulation of expression. Additionally, recognition sites for HRV 3C Protease, Thrombin, and Factor Xa are located between TF-Tag and the Multiple Cloning Site (MCS) and function to facilitate tag removal from the expressed fusion protein. Most E. coli strains can serve as expression hosts. The pCold TF DNA Vector provides cold shock technology for high yield protein expression combined with Trigger Factor (chaperone) expression to facilitate correct protein folding, thus enabling efficient soluble protein production for otherwise intractable target proteins.(pdf-I)Chaperone Plasmid Set / Chaperone Competent Cells (pdf-II) CL7/Im7 tag TRIALTUSThe ultra-high-affinity (CL7/Im7) purification system allows for one-step purification of a wide range of traditionally challenging biological molecules, including eukaryotic, membrane, toxic, and DNA/RNA-binding proteins and complexes. High activity, high purity. Excellent for low-abundance proteins. N or C terminal. Elution: specific proteases as TEVprotease , SUMO, others. Column could be re-use several times (pdf) Dmitry G. Vassylyev, Department of Biochemistry and Molecular Genetics, University of Alabama at Birmingham (pdf) (pdf Appendix) LYTAG and LYTAG Two Phase System BIOMEDALLYTAG system allows the single-step purification of proteins fusion to LYTAG and C-LYTAG using LYTRAP resin, a simple and efficient chromatographic support. Binding conditions are gentle and do not involve covalent modifications, in such a way that the fusion protein is highly stable once bound to the resin. LYTAG and C-LYTAG fusions protein are selectively eluted using choline-containing buffers. No unwanted amino acids after tag elimination by enterokinase digestion (pdf) BIOMEDALLYTAG Two-Phase is a protein purification system based on the use of two aqueouscomponents. The method relies on the affinity of the protein tag LYTAG for one of the two-phase components, allowing recombinant protein separation and purification from cellular extracts or culture media. In the procedure, the LYTAG-fused protein is retained in one of the aqueous phases while most of the undesired proteins can be removed by simply discarding the opposite phase. After replenishing the system with fresh phase, the protein of interest can be easily recovered in it, with high purity, by reversing its localization with the addition of choline, the specific LYTAG ligand.Purification can be easy and safely performed at low temperatures, requiring only a refrigerated centrifuge and an ice water bath. Alternative to conventional solid resins. (pdf-I) (pdf-II) Profinity eXact BIO-RADProfinity eXact fusion-tag system is an affinity tag-based protein purification system that utilizes a modified form of the subtilisin protease, which is immobilized onto a chromatographic support and used to generate pure, tag-free target protein in a single step. The tag in this system is the prodomain (prosignal sequence) of the subtilisin protease, a 75-amino acid sequence that is fused to the N-terminus of a target protein of interest. The mature and prodomain subtilisin protease sequences have been co-engineered to produce a highly specific, high-affinity interaction between the binding partners. Application of elution buffer triggers subtilisin s processing activity, which quickly and precisely cleaves the tag from the fusion protein and releases the purified target protein. At the end of the purification process, the tag remains tightly bound to the resin and contains only its native amino acid sequence. The structure and activity of the native protein is maintained, and the need for additional materials, such as cleavage enzymes and purification resin for postcleavage enzyme removal, is eliminated. (pdf-I) (pdf-II)(pdf-III) Profinity eXact purification resin (pdf-IV)(pdf-V)Bispecific Affinity Johan Nilvebrant, Tove Alm, Sophia Hober (2012)Orthogonal Protein Purification Facilitated by a Small Bispecific Affinity TagJournal of Visualized Experiments 59 e3370: 1-5Bispecific purification tag with two different binding sites on a 46 amino acid small protein domain: an albumin-binding domain (derived from Streptococcal protein G and with a strong affinity to human serum albumin (HSA), that can be recognized with HSA Sepharose column; and a second domain against a dimeric Z-domain derived from Staphylococcal protein A, that can bound HiTrap MabSelect SuRe from GE(pdf) The video component of this article can be found at http://www.jove.com/video/3370/ CaptureSelect C tag THERMO:CaptureSelect C tag. A small 4 amino acid peptide tag: E P E A (glutamic acid proline glutamic acid alanine) that binds toa 13 kDa Camelid antibody fragment affinity matrix.The CaptureSelect C tag affinity matrix purifies C terminal tagged proteins with high affinity and selectivity, even in the presence of Urea and Guanidine HCl, from complex mixtures like cytoplasm or periplasmatic fractions in a one step process. Mild elution conditions at neutral pH can be applied using 2M magnesium chloride or 50% propylene glycol, which ensures high activity recoveries of pH sensitive target proteins; or more specifically with 2mM S E P E A peptides.The affinity resin recognizes the E P E A tag sequence when fused either directly to the C terminus of a protein or through linkers between the C terminus and the E P E A tag(pdf) Cleavage of Recombinant Proteins Cleavage Sites Table for Fusion Proteins Proteases sensitivity to Detergents James M. Vergis, Michael C. Wiener (2011)The variable detergent sensitivity of proteases that are utilized for recombinant protein affinity tag removalProtein Expression and Purification 78: 139 142 (pdf)David S. Waugh (2011) An overview of enzymatic reagents for the removal of affinity tagsProtein Expression and Purification (pdf)Mohanty A. et.al. (2003) Inhibition of tobacco etch virus protease activity by detergents. Protein Expression and Purification 27: 109 114 (pdf) Factor XaFactor Xa Cleavage of a MBP fusion protein NOVAGEN: Factor-Xa kit (pdf)NOVAGEN: Restriction Grade Factor Xa.Factor Xa Cleavage Capture Kit .(pdf pg.54)QIAGEN: Factor Xa and Factor Xa Removal resin(pdf) Thrombin Thrombin Cleavage of a GST fusion protein NOVAGEN: Thrombin kit(pdf)NOVAGEN: Thrombin, Restriction Grade. Biotinylated Thrombin andThrombin Cleavage Capture Kit (pdf pg.53) SIGMA: rec Human Thrombin(pdf)Enterokinase NEW ENGLAND BIOLABS:Enterokinase (pdf) NOVAGEN: Enterokinase cleavage capture kit(pdf) NOVAGEN: Recombinant Enterokinase. Tag off High Activity rEK is a highly purified preparation of the catalytic subunit of human enterokinasethat recognizes the identical cleavage site as the native enzyme, AspAspAspAspLys . Enterokinase Cleavage Capture Kit. Tag off rEK CleavageCapture Kit(pdf pg.55-56) ROCHE:Enterokinase(pdf) TAGZyme UNIZYME The TAGZyme System is an efficient and specific method for complete removal of N-terminal his-tags by use of exopeptidases. The method is based on the use of dipeptidyl aminopeptidase I (DAPase) alone or in combination with glutamine cyclotransferase (Qcyclase) and pyroglutamyl aminopeptidase (pGAPase). These enzymes all have the ability to bind to IMAC matrices through an engineered his-tag in recombinant formsQIAGEN: His-tag Removal by exoproteolytic Digestion(pdf) Jos Arnau, et.al. (2006) Current strategies for the use of affinity tags and tag removal for the purification of recombinant proteins Protein Expression and Purification 48 1 13 (pdf) TEV-Protease INVITROGEN- LIFE TECHNOLOGIES TEV-Protease(pdf) SIGMA: TEV protease Biotin tagged ( pdf) Mohanty A. et.al. (2003) Inhibition of tobacco etch virus protease activity by detergents. Protein Expression and Purification 27: 109 114 (pdf) PROMEGA: ProTEV Protease is an improved 50kDa version of the NIa protease from tobacco etch virus (TEV) that has been engineered to be more stable than native TEV protease for prolonged enzymatic activity (pdf) CARDIFF UNIVERSITY - MOLECULAR CELL BIOLOGY - EHRMAN LABTEV NIa protease PreScission (HRV-3C protease) GE-HEALTHCAREApplication Note(pdf) GE-HEALTHCARELife Science News(pdf) NOVAGEN HRV 3C Protease(pdf) (pdf pg.57)SIGMA: HRV-3C protease from human Rhinovirus Type 14 is a protease that specifically cleaves within an eightresidue recognition sequence. This sequence is: Leu-Glu-Val-Leu-Phe-Gln- Gly-Pro (pdf)SIGMA: HRV 3C protease Biotin tagged (pdf) SUMO PROTEASE INVITROGENSUMO Protease (pdf) LIFESENSORS SUMO Protease 1(pdf) (pdf-II)(pdf-III) SIGMA: SUMO protease recognizes the tertiary structure of the Ubiquitin-like SUMO domain and hydrolyzes the peptide bond in the x Gly Gly x sequence after the Gly-Gly bond, at the C-terminus of the SUMO domain (pdf) SIGMA: SUMO protease Biotin tagged (pdf) FURIN NEW ENGLAND BIOLAB Furin is a ubiquitous subtilisin-like proprotein convertase. It is the major processing enzyme of the secretory pathway and islocalized in thetrans-golgi network. Substrates of Furin include blood clotting factors, serum proteins and growth factor receptors such as the insulinlikegrowth factor receptor. The minimal cleavage site is Arg-X-X-Arg\'. However, the enzyme prefers the site Arg-X-(Lys/Arg)-Arg\'. Anadditional arginine at the P6 position appears to enhance cleavage. Furin is inhibited by EGTA, 1- Antitrypsin Portland and polyarginine compounds.Isolated from Spodoptera frugiperda (Sf9) cells infected with recombinant baculovirus carrying truncated human furin (pdf) Affinity ChromatographyActivated Resins ABT Amino Ethyl and Glyoxal Agarose Beads (pdf-I)(pdf-II)Glyoxal Agarose Beads includes a broad selection of resins for the immobilization of biomolecules through amino groups (enzymes, antibodies, etc). There are resins, with either 4% or 6% agarose concentration, and with various degrees of activation (very high/ high/low). This permits the immobilization of biomolecules in different size ranges. (pdf-I)Aminoethyl Agarose Beads covalently bind the agarose to the acid groups of the amino acid of the target biomolecules: acidic amino acids like aspartic acid or glutamic acid.There are resins with agarose concentrationof 4% or 6% and with a range of degrees of activation. (pdf-I) BIO RADAffi-Gel 10 and Affi-Gel 15 activated immunoaffinity supports (N-hydroxysuccinimide esters of a derivatized crosslinked agarose), with a 10 and 15-atom spacer arm respectively, that offer rapid, high efficiency coupling for all ligands with a primary amino group aqueous or non-aqueous solution.(pdf)GE-HEALTHCARE Affinity Manual (pdf) Speci c Groups of Biomolecules (pdf) GE-HEALTHCAREAffinity Columns and Media (pdf) HiTrap NHS-activated HP, NHS-activated Sepharose 4 Fast Flow (10-atom spacer arms) is designed for the covalent coupling through the primary amine of a ligand and is the first choice for the preparation of immunospecific media. CNBr-activated Sepharose offers a well-established option for the attachment of larger ligands and as an alternative to NHS-activated Sepharose. Proteins, peptides, amino acids or nucleic acids can be coupled to CNBr-activated Sepharose, under mild conditions, via primary amino groups or similar nucleophilic groups. EAH Sepharose 4B for coupling small ligands containing free acarboxyl groups via a 10-atom spacer arm. Carbodiimide as the coupling reagent. Phenolic groups may be attached through others couple reagents. Thiol derivatives can couple carboxyl groups in the presence of carbodiimide and the thiol ester bond may be cleaved specifically using hydroxylamine, thus providing a simple and gentle method for eluting the intact ligand-protein complex. ECH Sepharose 4B for coupling small ligands containing free amino groups via a 9-atom spacer arm. Carbodiimide as the coupling reagent. Epoxy-activated Sepharose 6B for coupling through hydroxy, amino or thiol groups via a 12-carbon spacer arm. It is particularly useful for coupling small ligands such as choline, ethanolamine and sugars. The pre-activated matrix is formed by reacting Sepharose 6B with the bis oxirane, 1,4 bis-(2,3-epoxypropoxy-)butane. Free oxirane groups couple via stable ether bonds with hydroxyl-containing molecules such as sugars, via alkylamine linkages with ligands containing amino groups, and via thioether linkages with ligands containing thiol groups. Thiopropyl Sepharose 6B for coupling through a thiol group many types of small ligands Heavy metal ions and derivatives can be used as ligands to react with thiol groups forming mercaptides. Alkyl or aryl halide ligands give thioether derivatives. Ligands containing C=O, N=N and, under certain conditions, C=C bonds undergo addition reactions. The hydroxypropyl group acts as a small spacer arm. Ligands containing amino groups can be attached to Thiopropyl Sepharose 6B or Activated Thiol-Sepharose 4B by multi-point attachment or coupling through a small number of groups using the heterobifunctional thiolating reagent, SPDP. The coupled molecules may be recovered by eluting with a reducing agent. GE-HEALTHCARECNBr Activated Sepharose Data File - React with primary amines present on proteins, antibodies and other molecules. (pdf) GE-HEALTHCARENHS Activated Sepharose Data File -NHS (N-hydroxysuccinimide) coupling forms a chemically stable amide bond with ligands containing primary amino groups. NHS-activated Sepharose 4 Fast Flow provides a spacer arm and is therefore particularly suitable for immobilising small protein and peptide ligands. (pdf)GE-HEALTHCARERecommended carbodiimide coupling procedure for CH Sepharose 4B and AH Sepharose 4B (pdf) JENA BIOSCIENCEImmobilized Nucleotides for Affinity Chromatography (pdf)Immobilized nucleotides provide a convenient and rapid one-step purification procedure for a large number of proteins such as kinases, GTPases, chaperones, motor proteins, and others. Jena Bioscience provides NTPs and dNTPs that are linked to the matrix at various positions of sugar, base or phosphate moiety of the nucleotide; different types and lengths of linkers and different types of chromatography material ranging from bulk material to pre-made columns that fit any machineMERCK Fractogel EMD Amino(pdf) For immobilization of ligands with carboxy-functionional groups.MERCK Fractogel EMD Epoxy (M) (pdf) Suitable for coupling of low molecular weight ligands and pH stable proteins. The epoxyde reacts with primary amino groups, hydroxyl, and sulfhydryl groups. The resulting affinity matrix is very stable, due to the ether bonding of the ligand. NOVAGEN Preactivated Resins (pdf) PreACT Agarose ALD is an activated chromatography matrix for simple and efficient immobilization of small ligands to large proteins under mildphysiological conditions. Is supplied in an activated form containing bound reactive aldehyde groups. During ligand coupling, aldehydes react with primary amines on the ligand to form a Schiff-base. In a further reaction, the Schiff-base linkage is selectively converted into a stable, secondary amine linkage. PreACT Fractogel AZL is an activated resin with a high density of tentacle-bonded azlactone reactive groups suitable for the coupling of proteins under physiological conditions. The azlactone ring is opened by nucleophilic attack of appropriate groups present on the protein surface. Upon reaction with amines, a stable amide bond forms between the former carbonyl function of the azlactone. PreACT Fractogel EPX is an activated resin for the immobilization of low molecular weight amine bearing ligands or alkaline-stable proteins. Bonded epoxide groups react with primary amino, hydroxyl and sulfhydryl groups to form stable ether linkages.PUROLITE Praesto CNBr: Pre-activated CNBr resin functionalized on a modern, high flow agarose base matrix for simplified ligand immobilization and fully customizable affinity chromatography purification solutions. Available in three particle sizes: 45 m, 65 m and 90 m (pdf pg27) (pdf)PUROLITE Praesto Epoxy: Pre-activated Epoxy resin functionalized on a modern, high flow agarose base matrix for simplified ligand immobilization and fully customizable affinity chromatography purification solutions. Available in three particle sizes: 45 m, 65 m and 90 m (pdf pg29) (pdf)PUROLITE Praesto NHS: Pre-activated NHS resin functionalized on a modern, high flow agarose base matrix for simplified ligand immobilization and fully customizable affinity chromatography purification solutions. Available in three particle sizes: 45 m, 65 m and 90 m (pdf pg31) (pdf)THERMO: Affinity Purification Handbook (pdf) THERMO: Coupling different Functional Groups: Tosyl, Tresyl and Epoxy Activated Agarose. Alkylamine Beads Tosyl Activated Agarose Hydroxyls on the surface of cross-linked beaded agarose are reacted with p-toluenesulfonyl chloride (tosyl chloride) to yield a sulfonated support. These sulfonates can couple to nucleophiles, such as primary amines or thiols, to yield a stable affinity support. The tosylated support also couples to imidazole, or tyrosine hydroxyl groups. Tosyl Activated Agarose couples more rapidly and more efficiently to sulfhydryl groups than to primary amines. Sulfhydryl coupling supports are convenient when accessible free sulfhydryls exist in the protein, or when one can be easily generated through reduction of a disulfide bond. Tresyl Activated Agarose Hydroxyls on the surface of agarose are reacted with 2,2,2-trifluoroethanesulfonyl chloride (tresyl chloride) to yield a sulfonated support. This sulfonated support is approximately 100-fold more reactive than a Tosyl agarose support. Tresyl Activated Agarose can couple to nucleophiles, such as primary amines or thiols, to yield a stable affinity matrix.. The tresylated support also couples to imidazole and tyrosine hydroxyl groups. Tresyl Activated Agarose couples more rapidly and more efficiently to sulfhydryl groups than to primary amines. ImmunoPure Epoxy-Activated Agarose Epoxide chemistry is a useful way to immobilize ligands like proteins, carbohydrates, peptides and amino acids, containing nucleophiles, such as amino,thiol and hydroxyl (including phenolic) functional groups. Epoxide-activated supports are produced by the immobilization of bifunctional oxiranes such as 1,4-butanediol diglycidyl ether onto agarose supports, introducing a hydrophilic spacer arm. These activated supports have limited stability in aqueous media, so it is necessary to use them quickly after they are generated or rehydrated.(pdf) THERMO: CarboLink Coupling Gel allows immobilization of glycoproteins through their oxidized carbohydrate moieties. Because carbohydrates are located on the Fc portion of antibody molecules, CarboLink Gel has the advantage of orienting the antibody binding sites to remain unobstructed, resulting in greater purification capability. (pdf) THERMO:Coupling Proteins trough Sulfhydril groups Pierce SulfoLink Coupling Gel is designed to efficiently react with thiol-containing molecules and immobilize them through a thioether linkage. The support contains an iodoacetyl group at the end of a long spacer arm, which reacts with sulfhydryls through displacement of the iodine. (pdf) THERMO: The AminoLink Coupling Gel support is 4% cross-linked beaded agarose that has been activated to form aldehyde functional groups. The aldehydes react spontaneously with primary amines found in lysine residues and at the amino terminus of a peptide chain. Reductive amination of the resulting Schiff base then forms a stable, secondary amine linkage with minimal leakage of the ligand. (pdf)THERMO: Batch and Spin Cup Methods for Affinity Purification of Proteins (pdf) THERMO: MicroLink Protein Coupling Kit allows immobilization of small amounts (25-100 g) of purified antibody and other proteins directly onto beaded agarose gel to create a permanent affinity support. The AminoLink Plus Coupling Gel included in this kit contains aldehyde functional groups that react with primary amines present on antibodies and other molecules. Reductive amination of the resulting Schiff bases forms a stable secondary amine linkage with minimal leakage.(pdf)THERMO:MicroLink Peptide Coupling Kit. For sulfhydryl-containing peptide or protein. This kit uses UltraLink Iodoacetyl Gel that reacts specifically with free sulfhydryls to form a stable thioether linkage. The support contains a 15-atom spacer that reduces steric hindrance, making binding interactions with the coupled molecule efficient.(pdf) THERMO: Ultra Link Immobilized Carboxy for Inmobilization of Peptides using EDC. Useful for coupling primary amine containing ligands like small peptides (pdf)THERMO:UltraLink Iodoacetyl Gel binds specifically to sulfhydryls when used under specific conditions with the iodoacetyl alkylating agent. The 15-atom spacer arm is especially ideal for conjugating small peptides to the support. The support will immobilize numerous types of molecules with a free sulfhydryl including antibodies, other proteins and peptides.(pdf)THERMO: Ultra Link Biosupport Medium with Azlactone Groups. Couples nucleophiles on ligands via a ring opening reaction to attach the ligand to the support through stable covalent linkages. Amino-functional ligands will form stable amide bonds at the end of a five-atom spacer. (pdf) SARTORIUS Sartobind Epoxy 75 - A Microporous Coupling Membrane for Affinity Chromatography - Operating Instructions - Any molecule containing amino-, hydroxyl- or thiol-groups may be immobilized by covalent coupling to the epoxy-activated membrane. The membrane is fitted into a filter holder with Luer Lock connectors for easy handling to quickly couple biomolecules like proteins or peptides covalently. (pdf) TOSOHAffinity Chromatography Activated Resins(pdf) TOYOPEARL AF-Tresyl-650M: For coupling of proteins through amine and thiol groups at slightly alkaline pH (7.0-8.0) TOYOPEARL AF-Epoxy-650M: Useful for attaching low MW ligands at high densities and for immobilization of carbohydrates or glycoproteins TOYOPEARL AF-Formyl-650M: (aldehide bearing) For coupling of proteins through amine groups at slightly alkaline pH (7.0-9.0) TOYOPEARL AF-Amino-650M: For coupling ligands through their carboxylate groups (peptide bond formation) or aldehyde groups (reductive amination) present in carbohydrates and glycoprotein ligands, or introduced into the ligand by mild periodate oxidation. Reaction at slightly acidic pH (4.5-6.0) TOYOPEARL AF-Amino-650M: Carbodiimide mediated coupling reaction to amino groups of proteins or low MW ligands. Reaction at slightly acidic pH (4.5-6.0) VIVASCIENCE: Vivapure Epoxy Coupling Kit - Mini Spin Columns - Vivapure Epoxy Mini spin columns provide a membrane to which any desired protein, containing amino-, sulfhydryl-, or hydroxyl-groups on its surface is coupled covalently. (pdf) Antibody Purification and antibody fragmentationRodrigo Vazquez-Lombardi et. al: Transient expression of human antibodies in mammalian cells Nature Protocols | VOL.13 NO.1 | 2018 | 99 Published online 14 December 2017; doi:10.1038/nprot.2017.126 (pdf) GE-HEALTHCAREAntibody Purification Manual(old pdf)(old pdf) (new pdf) GE-HEALTHCARE Columns and resinsfor antibody purification and immunoprecipitation. Selection guide (pdf) GE-HEALTHCAREApplication Note: Rapid optimisation and development of an automated two-step purification procedure for monoclonal IgG antibodies(pdf) GE-HEALTHCARECapto adhere: a strong anion exchanger with multimodal functionality ionic interaction, hydrogen bonding and hydrophobic interaction. Capto adhere is designed for post Protein A purification of monoclonal antibodies. Removal of leached Protein A, aggregates, host cell proteins, nucleic acids and viruses from monoclonal antibodies is performed in flowthrough mode at where the antibodies pass directly through the column while the contaminants are adsorbed. (pdf-I) (pdf-II) (pdf-III)GE-HEALTHCAREKappa Select is an affinity chromatography medium designed for the purification of Fab (kappa) with high binding capacity, purity and yield. The ligand binds to the constant region of the kappa light chain. Use for the production of Fabs for clinical applications. (pdf)GE-HEALTHCAREHiTrap MabSelect SuRe, HiTrap MabSelect and HiTrap MabSelect Xtra for purification of monoclonal antibodies (MAbs) (pdf)GE-HEALTHCARECapto L: strong affinity for the variable region of an antibody s kappa light chain. Suitable for capture of a wide range of antibody fragments such as Fabs, single-chain variable fragments (scFv), and domain antibodies (Dabs).(pdf) A platform approach for the purification of antibody fragments (Fabs) (pdf) BAC (BioAffinity Company) CaptureSelect Human IgA affinity matrix contains an affinity ligand that binds to a unique domain that is present on all subclasses of human IgA, with no cross reactivity with IgM or IgG. (pdf-I) (pdf-II) CaptureSelect IgM affinity matrix contains an affinity ligand that is directed towardsa unique domain that is present on both human and mouse IgM antibodies, and offers no cross-reactivity with human or mouse IgA or IgM.(pdf) CaptureSelect Fab kappa affinity matrix: for all human antibodies containing kappa light chains (pdf) CaptureSelect Fab lambda affinity matrix: for all human antibodies containing lambda light chains (pdf) CaptureSelect Human Fc affinity matrix: for all subclasses of human IgG (pdf) CaptureSelect Multi Species affinity matrix: for IgG from multiple species (pdf) CaptureSelect Multi Species affinity matrix: for human IgG4 (pdf) BIORAD Affi-Prep Protein A Matrix Instruction Manual(pdf)BIORAD Affi-Gel Protein A MAPS II Kit - Instruction Manual (pdf) BIORADEcono-Pac Protein A Kit Protein A Columns - Instruction Manual (pdf)BIO RADImmunoaffinity Kit for IgG coupling trough Fc region(pdf) BIO RAD Protein A removal from IgG on CHT Ceramic Hydroxyapatite Support(pdf)CALBIOCHEMHuman IgG subclasses(pdf) CLONTECH: Thiophilic - Purification of Immunoglobulins (pdf) CLONTECH:CapturemProteinAMiniprep,CapturemProteinAMaxiprep,andCapturemProteinA96. Spinnableaffinitycolumnsorplatescontaining novelhigh capacityProteinAnylonmembranes. Speed,ease of use,flexibility,andhighyieldtoantibodypurificationandscreening. (pdf) GENOVIS:FabRICATOR is the fastest and most accurate enzymatic tool to generate F(ab )2 and Fab fragments from antibodies. The FabRICATOR utilizes the unique specificity of a novel proteolytic enzyme that cleaves all human, rabbit, sheepand monkey IgG subclasses at the same place in thehingeregion creating pure F(ab )2 fragments without any further degradation. Cleave IgG within just a few minutes, at mild, physiological pH conditions. No sample degradation, high yields. (pdf)FragIT columns consist of the proteolytic antibody-specific enzyme FabRICATOR immobilized on agarose.FragIT is the fastest and easiest way tofragment antibodies into F(ab\')2 and Fc; cleave up to 100mg IgG in just half an hour. Reaction is carried out at pH 6.8. (pdf) FragIT microspin columns allow for easy and quick fragmentation of IgG into F(ab )2 and Fc fragment; fragmentation of up to 0.5 mg IgG in less than 30 minutes. FragIT microspin columns come prefilled with FabRICATOR enzyme immobilized on agarose (pdf) IgGZERO is a very efficient endoglucosidase with high specificity for IgG. The enzyme cleaves the chitobiose core of the glycan on IgG; it has the ability to hydrolyze human, Rhesus monkey, mouse, rat, rabbit, horse, goat, dog andporcine native and denaturated IgG.Removal of IgGZERO isextreamly easy by the included histidine tag in the amino terminal of the recombinant enzyme (pdf-I) Micrspin (pdf-II) Life Technologies Protein handbook: Protein purification using single-domain antibody fragments as affinity ligands. CaptureSelect affinity protocols (pdf-Chapter V) LigaTrapIgM Purification(pdf) LigaTrap Llama IgG Purification Kit and Resin. The LigaTrap Llama IgG Resin (LT-144) binds and captures allthree isotypes. LigaTrap Human IgA Purification Resinand column LigaTrap Chicken IgY PurificationMERCKCapture of Mouse Monoclonal Antibodies by Strong Cation Exchange Chromatography(pdf)Eshmuno S, a cation exchanger specifically designed for highly productive downstream purification of monoclonal antibodies. Hydrophilic polyvinyl ether base matrix that allows the use of much higher flow rates. (pdf)MILLIPOREAffinity Chromatography Media: ProSep -vA Ultra Media, ProSep-vA High Capacity Media, ProSep-A High Capacity Media ProSep-rA High Capacity Media. OPERATING INSTRUCTIONSProSep-A affinity media have been developed specifically for industrial scale purification of monoclonal antibodies where highly efficient purification can be achieved using clarified bioreactor feedstock at physiological pH and salt concentration. (pdf)MILLIPORE PROSEP-Thiosorb and PROSEP-Thiosorb M Chromatography Media - for the recovery and purification of Immunoglobulins (pdf)Pall BioSepra Protein A Ceramic HyperD F sorbent. High capacity affinity sorbent designed for process-scale purification of immunoglobulins G. (pdf)(pdf-II)PallPurification of mouse IgM from cell culture supernatant by Cation Exchange Chromatography on CM Ceramic HyperD F sorbent (pdf)PallPurification of mIgG1 on MEP HyperCel Mixed-Mode media. Optimization (pdf)PUROLITE Praesto AC: Modern agarose-based Protein A affinity resin. With capacity over 40 mg/mL at 4 minutes residence time or higher, PraestoA combines high capacity, excellent pressure/flow performance, and NaOH CIP stability for over 20 cycles, thus meeting the common requirements for production of materials for PI and PII clinical trials. It is an excellent choice for the capture step in a typical MAb platform process. (pdf pg13) THERMO: Antibody Technical Handbook. The handbook provides an overview of antibody structure and types, as well as technical information on the procedures, reagents and tools used to produce, purify, fragment and label antibodies. (pdf-1) (pdf-2) (pdf-3)(pdf-THERMO)THERMO:Affinity Purification Handbook (pdf)THERMO:Binding Characteristics of Immunoglobulin Binding Proteins and Thiophilic Gel (PowerPoint) THERMO:Fab Preparation Kit enables efficient Fab generation from IgG. This kit uses papain, a nonspecific thiol-endopeptidase, immobilized on agarose resin.Immobilized enzyme is advantageous because digestion can be immediately stopped by simply removing the IgG solution from the resin, resulting in adigest that is enzyme-free. Digestion by papain produces 50 kDa Fab and Fc fragments (pdf)THERMO: Immobilized E. coli Lysate. Partially purified bacterial proteins from a suspension of E. coli cells (strain BMH 71-18) immobilized onto agarose, provides a simple and efficient method of removing E. coli-reactive proteins from antibody preparations (pdf_I) (pdf-II) THERMO: Antibody Immobilization: Choosing the Best Support(PowerPoint) THERMO: Immobilized Jacalin (human IgA and IgD binding lectin).Jacalin immobilized on supports such as agarose has been useful for the purification of human serum or secratory IgA1(pdf) (pdf-II)THERMO: T-Gel Purification Kit (pdf) THERMO: Pierce Protein L Agarose. Protein L binds to immunoglobulin kappa light chains without interfering with the antigen-binding site and binds awider range of Ig classes and subclasses than other antibody-binding proteins such as Protein A or Protein G. Protein L binds to all classes of Ig(i.e., IgG, IgM, IgA, IgE and IgD). Protein L also binds single chain variable fragments (Scfv) and Fab fragments. Protein L only binds to immunoglobulinscontaining light chains of type kappa I, III, and IV in human and kappa I in mouse. Protein L also may be specific for certain kappa subgroups in other species.Protein L binds scfv without interfering with antigen binding. Protein L binds weakly to rabbit immunoglobulins and does not bind immunoglobulins from bovine,goat or sheep; nor does it bind to lambda light chains. (pdf)SARTORIUS Sartobind Protein A 75 Membrane Adsorbers - Operating Instructions A Separation Technology Based on Microporous Membranes(pdf) TOSOHTOYOPEARL AF-rProtein L-650F Resins. Methacrylic polymer. Protein L based affinity chromatography is used for the capture of antibodies and antibody fragments that do not bind to Protein A. Unlike Protein A and G, which bind to the Fc region of immunoglobulins (IgGs), Protein L binds through interactions with the variable region of an antibody s kappa light chain (pdf) (pdf-flyer) VIVASCIENCE:Vivapure Protein A - Mini Spin Columns (pdf)General BIO RADPolymixin matrix - For removal of Endotoxin molecules (pdf)GE-HEALTHCAREVIII Select. Highly selective to recombinant b-domain depleted Factor VIIIa key recombinant blood factor used for treatment of Hemophilia A. Efficient purification of recombinant b-domain.(pdf) GE-HEALTHCAREKappa Select is an affinity chromatography medium designed for the purification of Fab (kappa) with high binding capacity, purity and yield. The ligand binds to the constant region of the kappa light chain. Use for the production of Fabs for clinical applications. (pdf) MILLIPORE Matrex Cellufine Sulfate - For concentration, purification and depyrogenation of Virus, Viral/Microbial Antigen, Heparin binding proteins (pdf) Pall BioSepra Blue Trisacryl M is an affinity chromatographic sorbent used for the purification of a wide variety of enzymes and proteins such as kinases, albumin, interferons, and some coagulation factors. (pdf)Pall BioSepra Heparin HyperD M composite chromatography sorbent for the purification of biological molecules that bind to heparin, such as coagulation factors, growth factors, lipoproteins. (pdf) THERMO: CaptureSelect Protein Affinity Resins: Human alpha-1 anti-trypsin, Human Antithrombin, Human C1 esterase inhibitor, C-terminal amino acid tag E-P-E-A, Human fibrinogen from human plasma, Follicle stimulating hormone, Human growth hormone (pdf) Immunoprecipitation ABCAM: Immunoprecipitation protocol. 1. Lysis buffers and other reagents 2. Preparation of lysates 3. Pre-clearing the lysates 4. Immunoprecipitation5. Choosing the correct beads(pdf-1)Troubleshhoting(pdf-II) Procedure to cross link the antibody to the beads to enable elution of protein withlittle antibody contamination (pdf-III) INVITROGEN: Dynabeads for protein complex isolation. Add your specific antibody or interacting protein with tags to Dynabeads , then immunoprecipitateyour protein of interest. Once the beads are exposed to a magnet, they are efficiently drawn to the tube wall, taking only your protein complex with them.As the process is gentle, yet very quick, complexes remain intact and functional. The complexes can be resuspended in a small volume ready for downstreamanalysis with mass spectrometry, gels, etc. (pdf) INVITROGEN: Immunoprecipitation with Dynabeads Protein A or Protein G. The captured immune complexes are easily removed from the supernatant bymagnetic separation. (pdf)THERMO: Seize X Immunoprecipitation Kits. Recover protein without antibody protein band interference. The Kit combine cross-linking and affinity chromatography to offer a new and improved immunoprecipitation method. First, the primary antibody is bound and immobilized to a Protein A or Protein G support using cross-linking agent (DSS). This properly orients the antibody to \"seize\" protein from crude cell lysate applied to the immobilized antibody support. Unbound proteins are then centrifuged away and the protein is recovered by using an elution buffer. (pdf -I) (pdf-II) (pdf-III) (pdf-IV)THERMO:The Thermo Scientific Pierce Direct IP Kit enables highly effective and efficient antigen immunoprecipitations by directly immobilizing purified antibodies onto an agarose support. Immobilizing the antibody provides faster and easier immunoprecipitations, enables reuse of the immobilized antibody and results in purified antigen free from antibody contamination. Immunoprecipitation is achieved using less than 10 g of antibody and a short coupling protocol to AminoLink Resin. The antigen sample is incubated with the immobilized antibody to form immune complex. The complex is washed to remove non-bound material, and a low pH elution buffer is used to dissociate the bound antigen from the antibody. (pdf) Lectins and Glycoprotein Purification CLONTECH:Glycoprotein Enrichment Resin is a phenylboronic acid-based resin which provides quick, efficient, and specific enrichment of glycoproteinsfrom complex samples such as human serum. The resin consists of an m-aminophenylboronic acid ligand coupled to agarose beads. The ligand bindsto cis-diol groups on sugar residues such as mannose, galactose, or glucose that are present within the saccharide moiety of glycoprotein molecules. This complex can be dissociated by lowering the pH, or by using an elution buffer containing either Tris or sorbitol. (pdf)GE-HEALTHCARE Con A Sepharose 4B (Con A binds molcules containing Alpha-D-mannopyranosyl, Alpha-D-glucopyranosyl and sterically related residues)(pdf-I) (pdf-II) GE-HEALTHCARE HiTrap Lectin Test Kit consists of four glycoprotein binding columns, HiTrap Con A, HiTrap Lentil Lectin, HiTrap Wheat Germ Lectin and HiTrap Peanut Lectin.(pdf) GE-HEALTHCARE HiTrap Wheat Germ Lectin (high affinity to N-acetylglucosamine and reacts strongly with the chitobiose core of N-linked oligosaccharides and N-acetylneuraminic acid) (pdf) GE-HEALTHCARE Lentil Lectin Sepharose 4B (binds to polysaccharides and glycoconjugates containing glucose or mannose type sugars) (pdf) CALBIOCHEMLectins (pdf) EY Laboratories Lectin Catalog. Lectin carbohydrate binding chart. Immobilized lectins and carbohydrates. Lectins kits. (pdf)THERMO: Immobilized Jacalin (human IgA and IgD binding lectin). Jacalin immobilized on supports such as agarose has been useful for the purification of human serum or secratory IgA1 (pdf) (pdf-II) THERMO:Glycoprotein Isolation Kit, WGA isolates glycoproteins from complex protein mixtures using the lectin wheat germ agglutinin (WGA) immobilized on agarose. The WGA lectin preferentially binds N-acetyl glucosamine (GlcNAC) and terminal GlcNAC structures that are commonly present in many serum and membrane glycoproteins. WGA also has affinity for sialic acid. (pdf)THERMO:Glycoprotein Isolation Kit, ConA isolates glycoproteins from complex protein mixtures using the lectin concanavalin A (ConA) immobilizedon agarose. The ConA lectin preferentially recognizes -linked mannose and, to a lesser extent, terminal glucose residues (pdf) PROMETICAminophenyl Boronate Affinity Chromatography for capture of glycoproteins. The aminophenyl boronate-glycoprotein covalent bond can be subsequently disrupted to elute the protein of interest with a pH change or by competitive elution using a sugar such as sorbitol.(pdf-I)(pdf-II)QIAGEN: Qproteome Glycoprotein Fractionation Handbook. For the fractionation of glycoproteins in proteomic samples (pdf) Qproteome Total Glycoprotein Kit. The Total Glycoprotein Spin Columns in the Qproteome Total Glycoprotein Kit contain ConA and WGA lectins. They are used for a general enrichment of the total glycoprotein population from a cell or serum sample. Qproteome Mannose Glycoprotein Kit. The ConA, GNA, and LCH lectin spin columns in the Qproteome Mannose Glycoprotein Kit are used for specific enrichment of glycoproteins with mannose-rich glycan moieties. The three lectins each bind different subclasses of these moieties. Qproteome Sialic Glycoprotein Kit. The WGA, SNA, and MAL lectin spin columns in the Qproteome Sialic Glycoprotein Kit are used for specific enrichment of glycoproteins with sialicacid-rich glycan moieties. The three lectins each bind different subclasses of these moieties. Qproteome O-Glycan Glycoprotein Kit. The AIL, and PNA lectin spin columns in the Qproteome O-Glycan Glycoprotein Kit are used for specific enrichment of glycoproteins with a glycan structure of the type that are found on T-antigens. The two lectins each bind different subclasses of these glycoproteins.QIAGEN: Qproteome GlycoArrays is a simple, rapid, kit-based method for determining the pattern and relative abundance of specific mammalian glycosylation epitopes in a glycosylated protein. The analysis can be performed on crude samples in growth media, eliminating the need for time-consuming purification and sample preparation steps. The technology consists of arrays of selected lectins, which are used to determine the glycosylation features of the analyzed glycoproteins. (pdf) Mimetics Ligands, Antibody fragments and Similar TechnologyTHERMO:These CaptureSelect products arecreated by a proprietary technology based on Camelid derived single domain antibody fragments. These antibody fragments are produced efficiently by our host-vector system based on the yeast Saccharomyces cerevisiae, enabling high level production with minimal purification effort as well as easy scale-up. CaptureSelect products possess a combination of unique properties such as stability, affinity, and selectivity that provides competitive benefits in terms of reduced cost of purification, higher quality product, and increased flexibility in the purification process.Custom ligands. CaptureSelect technology enables new opportunities in the field of custom designed ligands. It is an unique ligand discovery program that is applicable to a wide variety of target molecules, ranging from bacteria and viruses to proteins, carbohydrates and even smallmolecules like haptens and peptide tags.(pdf-1)(pdf-2) (pdf-3)NECTAGEN nanoCLAMP Single DomainAntibody Mimetics.nanoCLAMP(CLostridial Antibody Mimetic Proteins) affinity reagents are recombinant 15 kD antibody mimetic proteins selected for tight, selective and gently reversible binding. The nanoCLAMP scaffold is based on an IgG-like, thermostable carbohydrate binding module from the Clostridium perfringens hyaluronidase (Mu toxin).nanoCLAMPs are 4 nm X 2.5 nm in size, about the same size as a nanobody. Nectagen have generated nanoCLAMP libraries with variable loops that are analogous to the complement determining regions of immunoglobulins and used these libraries to isolate nanoCLAMPs recognizing specific immunogens. The nanoCLAMPs can be used to pull down proteins or protein complexes from whole cell extracts followed by elution with propylene glycol under conditions that preserve activity and protein-protein interactions. The nanoCLAMPtechnology is described in theProtein Expression and Purificationarticle \"Development of polyol-responsive antibody mimetics for single-step protein purification\" bySuderman et al. 2017. PROMETICMimetic Ligand Affinity Chromatography. The Mimetic Ligand adsorbent range has been designed to enable purification of many proteins, providing a general technique that can replace ion-exchange, gel permeation and hydrophobic interaction chromatography, but with higher recovery and purity. Mimetic Ligand Screening Column Kit contains 1ml pre-packed columns of ten different Mimetic LigandTM adsorbents, suitable for attachment to automated chromatography work stations. (pdf) SCIL Proteins Affilin molecules are small non-immunoglobulin proteins which are designed for specific binding of proteins and small molecules.New Affilin molecules can be quickly selected from libraries which are based on two different human derived scaffold proteins.Affilin moleculescan be further modified for various applications. Additional functionalities can be generated through genetic fusions with enzymes or effector domains. Specific labeling, chemically or with isotopes, extends the field of applications for Scil Proteins individualised binding proteins.Affilin proteins are easily produced in the cytoplasm of E. coli and are fully functional without further posttranslational modifications such asglycosylation. Affilin molecules will findtheir applications in therapy, chromatography, in vivo and in vitro diagnostics and as research tools.Albumin and Other Abundant Serum Proteins Removal AGILENT The Agilent Multiple Affinity Removal System for Proteomics Sample Preparation. Designed to remove greater than 98-99% of six interfering high-abundant proteins using Affinity-purified polyclonal antibodies to: albumin, IgG, IgA, transferrin, haptoglobin, and antitrypsin from human serum or three proteins (albumin, IgG, and transferrin) from mouse serum samples (pdf-I) (pdf-II) (pdf-III) (pdf-IV) (pdf-V)AGILENTThe Agilent Multiple Affinity Removal System. Designed to remove six interfering high-abundant proteins from Monkey Plasma for Proteomics Sample Preparation (pdf)AGILENTAgilent Human 14 Multiple Affinity Removal System Columns for the Fractionation of High-Abundant Proteins from Human Proteomic Samples (pdf) CALBIOCHEMProteoExtract Removal Kits (for Enhancing Resolution of Low Abundance Proteins) (pdf) GE-HEALTHCARE Capto Blue a new affinity chromatography prototype resin for purification or removal of HSA.Made by immobilizing Cibacron Blue to a new High Flow Agarose matrix (pdf) GE-HEALTHCARE HiTrap Albumin IgG Depletion and Albumin IgG Depletion SpinTrap: prepacked columns for the depletion of albumin and IgG from human serum and plasma. For small-scale preparation of protein samples prior to downstream analyses such as 1-D or 2-D gel electrophoresis and mass spectrometry. (pdf)NORGENProteoSpin Abundant Serum Protein Depletion Kit. The kit depletes 70% of albumin, 90% of -antitrypsin, and 50% of transferin and haptoglobin, enabling the visualization of low abundance proteins. The kit is unique in that it is based on an ion-exchange mechanism,silicon carbide (SiC), and not the use of specific antibodies. As a result, the kit can be used to deplete serum proteins from a wide variety of samples, including human and various animals. (pdf) PALL HyperCel STAR AX. Salt Tolerant Advanced Recovery Anion Exchange. Composed of a rigid cellulose matrix. (pdf) Capture of Human Serum Albumin from Plasma (pdf) THERMO:SwellGel Blue Albumin Removal Kit (pdf)QIAGEN:Qproteome Albumin/IgG Depletion Handbook. Fast and specific removal of albumin and IgG from human serum and plasma samples. Based on monoclonal antibodies that bind HSA and human IgG with high affinity and specificity.(pdf)SIGMA:ProteoPrep 20 Plasma Immunodepletion Kit is a complete kit with all necessary reagents and consumable equipment to deplete 20 high abundance proteins from human plasma or serum. This kit is designed to specifically remove the 20 proteins from human plasma. Specifically, 8 mL of plasma may be depleted in preparation for proteomic analysis, two-dimensional electrophoresis (2DE), or liquid chromatography (LC). (pdf). VIVASCIENCE: Vivapure Anti-HSA Kit for Human Albumin DepletionHighly specific human albumin depletion with unique antibody fragments (pdf-I) (pdf-II)(pdf-IV)(pdf-V)Vivapure Anti-HSA Affinity Resin (pdf-III)VIVASCIENCE:Vivapure Anti-HSA/IgG Kits for Human Albumin and Human Albumin/IgG Depletion (pdf)ChromatofusingGE-HEALTHCAREChromatofusing Handbook(pdf) GE-HEALTHCARE Ion Exchange Cromatography and Chromatofusing Handbook(pdf) Crystallography and Recombinant Methods HAMPTON: Solubility Stability Screen is designed to assist in the identification of solution conditions which promote protein solubility and stability, and minimize protein precipitation. Solubility Stability Screen is a solubility screen, a stability screen, and may also be used as an additive screen in the presence of a crystallization reagent.(pdf-I)(pdf-II) JENAMacromolecular crystallography products. Crystallization Screens: JBScreen Basic and JBScreen Membrane. Crystallization Optimization: JBS Solubility Kit, JBS Methylation Kit, JBScreen Plus, JBScreen Detergents, JBScreen Buffer Kits and JBScreen Cryo (pdf-I) (pdf-II) JENA Crystallization Freshman Kit - Junior andCrystallization Freshman Kit - Scholar are designed for screening of initial crystallization conditions ofproteins, peptides, nucleic acids and macromolecular complexes in order to grow single crystals suitable for X-ray diffraction analysis. They includeeverything you need to start your crystallization experiment: pregreased sitting drop plates, cover slides, our unique JBScreen reagents as well adetailed user guide. (pdf-I) (pdf-II) (pdf-III) (pdf-IV) (pdf-V)JENAProtein Crystallization Starter Kit. Introduction and Theory ofCrystallization. Which Materials are Useful as Precipitants(pdf)Derewenda Z., The use of recombinant methods and molecular engineering in protein crystallization.Methods (2004),34: 354 363(pdf) Smyth D., REVIEW Crystal structures of fusion proteins with large-affinity tags. Protein Science (2003), 12:1313 1322(pdf)Geerlof A. et.al. The impact of protein characterization in structural proteomics. Acta Cryst.(2006) D62: 1125-1136(pdf)Newby Z. et.al. A general protocol for the crystallization of membrane proteins for X-ray structural investigation. Nature Protocols (2009) 4: 619-637 (pdf) Lee J. et.al. An efficient platform for screening expression and crystallization of glycoproteins produced in human cells. Nature Protocols (2009) 4: 592-604 (pdf)Deglycosylation NEW ENGLAND BIOLAB Rapid PNGase F is the most effective enzymatic method for removing almost all N-linked oligosaccharidesfrom glycoproteins. PNGase F digestion deaminates the aspargine residue to aspartic acid, and leaves the oligosaccharide intact, keeping it suitable for further analysis(pdf) NORTHSTAR Enzyme Deglycosylation Kit. Contains all enzymes needed to completely remove all N- simple O-linked carbohydrates from glycoproteins (pdf) QAbioEnzymatic CarboRelease Kit will remove all N-linked oligosaccharides and many O-linked oligosaccharides from glycoproteins. N-links (Asn-linked) are removed using the enzyme PNGase F. In addition, all Ser/Thr-linked (O-linked) Gal-( 1-3)-GalNAc-( 1) and all sialic acid substituted Gal-( 1-3)-GalNAc-( 1) will be removed using the combination of Sialidase and O-Glycosidase. The addition of -Galactosidaseand Hexosaminidase will assist in the deglycosylation of larger O-link structures. (pdf) PROZYME-GLYKO Enzymatic Deglycosylation Kit. Contains all enzymes reagents needed to completely remove all N-linked simple O-linked carbohydrates from glycoproteins (pdf) ROCHEThe N-Glycosidase F Deglycosylation Kit can be used to test for the existence of asparagine-linked glycan chains on glycoproteins. The new Endoglycosidase H Deglycosylation Kit is useful for testing the existence of high mannose type and hybrid type asparagine-linked glycan chains on purified glycoproteins. (pdf-I) (pdf-II)SIGMA Deglycosylation Selection Guide(pdf) Expanded Bed Adsorption GE-HEALTHCAREIntroduction to Expanded Bed Adsorption (pdf) GE-HEALTHCAREStreamline Application Note (pdf)Streamline Data File(pdf) BIORAD CHT Ceramic Hydroxyapatite: Use in Expanded Bed Adsorption Mode (pdf) GelFiltration Protein concentration by ultrafiltrationBioWorks WorkBeads 40/100 SEC, WorkBeads 40/1000 SEC and WorkBeads 40/10 000 SEC resins are used for preparative size exclusion chromatography (SEC) resins in laboratory and process scale purification of proteins, virus and other biomolecules by utilizing the differences in their size. The resins are based on agarose, which is biopolymer suitable for separation of biomolecules (pdf) GE-HEALTHCAREGel Filtration Handbook(pdf)(pdf new) Fundamentals. New-generation agarose resins. Applications. (pdf) GE-HEALTHCAREGel Filtration of peptides - buffer conditions (pdf) GE-HEALTHCARE Packing of Gel Filtration column (movie) GE-HEALTHCARE Column evaluation (movie) GE-HEALTHCARE Maintenance and cleaning of size exclusion chromatography columns (pdf) GE-HEALTHCARE Superdex 200 Increase column (pdf) Superdex Peptide PE 7.5x300 (pdf) MERCKFractogel EMD BioSEC (S)(pdf) SEPAX Zenix SEC; silica 3 m particle size analytical column (pdf) SEPAX SEC-C line columns: for samples that shows a tendency to stick to traditional size exclusion resins with delayed elution time, low recovery, varying HMWS, or excessive tailing (pdf-user manual)(pdf-comparision) (pdf-cataloge 2016) SEPAX Preparative Size Exclusion Chromatography - SRT-10 SEC-300 Prep Column - 21.2 x 400 mm (141 mL) - Pressure: 17 bar at 7 mL/min, 25 bar at 10 mL/min (pdf) SHODEX SEC columns(pdf)TOSOH TSKgel SW series SEC columns. TSKgel SW, SWXL and SuperSW columns are stable from pH 2.0 to 7.5 and have excellent solvent stability up to 100% organic solvent. (pdf) TOSOH Chromatographic Media Catalog(pdf) TOSOHSize Exclusion Chromatography (pdf-I) (pdf-II) (pdf-III) (pdf-IV)Hydrophobic Interaction Chromatography Test Tube for HIC Resins BioToolomics: SepFast Butyl, SepFast Pentyl, SepFast Hexyl, SepFast Phenyl, SepFast Heptyl and SepFast Octyl are hydrophobic interaction chromatography (HIC) adsorbents having a carbon chain of C4, C5, C6, C6 ring, C7 and C8, respectively. There is a choice of 3 different base matrices according to the pore accessibility of target molecules:400 serial) to purify peptides or small proteins; 500 serial) to purify most medium to large proteins, and 600 serial) to purify large to very large proteins. (pdf) G-Bioscience G-Sep Butyl Agarose Fast Flow (FF) (pdf) GE-HEALTHCARE Butyl-S Sepharose 6 Fast Flo (pdf)GE-HEALTHCARE Hydrophobic Interaction and Reversed Phase Chromatography Principles and Methods (pdf) GE-HEALTHCARE Hydrophobic Interaction Chromatography Manual(pdf) GE-HEALTHCARE HiTrap HIC Selection Kit consists of six Hydrophobic Interaction Chromatography media (HIC) with different hydrophobic characteristics. The kit provides the possibility to screen for the most appropriate HIC medium for specific applications. (pdf-I) (pdf-II) GE-HEALTHCARERESOURCE ETH (ether), ISO (isopropyl) and PHE (phenyl) are pre-packed high performance columns for separating biomolecules by hydrophobic interaction chromatography (HIC)(pdf)GE-HEALTHCARE Packing of IEX, Affinity or HIC column (movie) GE-HEALTHCARE Column evaluation (movie) BIORAD Macro-Prep Hydrophobic Interaction Chromatography Support - Instruction Manual (pdf-I)(pdf-II) MERCKCleaning and Regeneraton of Fractogel EMD sorbents(pdf)MERCK Fractogel EMDPhenyl (S) Hydrophobic Interaction Chromatography (HIC)(pdf) MERCK Fractogel EMD Propyl (S) Hydrophobic Interaction Chromatography(HIC) (pdf) SEPAX Polar MC chromatographic media is composed of hydrophilic polymethacrylate beads with high physical and chemical stability (pdf) SEPAX Generik MC chromatographic media are composed of polymethacrylate beads with high physical and chemical stability (pdf)TOSOHUse of Hydrophobic Interaction Chromatography With a Non-Salt Buffer System for Improving Process Economics in Purification of Monoclonal Antibodies(pdf) TOSOH Chromatographic Media Catalog(pdf)TOSOH HIC Effects of Mixed Electrolites on Protein Separation (pdf) TOSOHHydrophobic Interaction Chromatography(pdf_I) (pdf-II) (pdf-III) HydroxyapatiteBIORAD Bio-Gel HT - Bio-Gel HTP DNA Grade Bio - Gel HTP Hydroxyapatite - Instruction Manual (pdf) BIORAD CHT Ceramic Hydroxyapatite (pdf)BIORAD CHT Ceramic Hydroxyapatite: Use in Expanded Bed Adsorption Mode (pdf) Application guide for process development and Scale-Up (pdf) How CHT Ceramic Hydroxyapatite works (pdf) Instruction Manual (pdf) BIORAD CFT Ceramic Fluoroapatite - A Chromatographic Support for Protein Purifications Requiring Acidic Conditions (pdf) BIO RADProtein A removal from IgG on CHT Ceramic Hydroxyapatite Support(pdf) BIORADMacro Prep Chromatography Supports(pdf) PallHA Ultrogel Hydroxyapatite Chromatography Sorbents (pdf)TOSOHCaPure-HA Resin for Aggregate Removal from Monoclonal Antibodies (pdf) DoE Optimization of Elution Conditions (pdf)Ion Exchange Chromatography Test Tube for IEX Resins BioToolomics: SepFast HighRes Q, SepFast HighRes DEAE, SepFast HighRes S and SepFast HighRes CM are, respectively, strong anion, weak anion, strong cation and weak cation exchange adsorbents. They are specially designed for high resolution purification of biological molecules. For each type of ion exchange medium, there is a choice of three different base matrices according to pore accessibility of target molecules (pdf) BioToolomics:SepFast Purifier Q, SepFast Purifier DEAE, SepFast Purifier S and SepFast Purifier CM are, respectively, strong anion, weak anion, strong cation and weak cation exchange adsorbents. They are specially designed for large-scale purification of biological molecules.For each type of ion exchange medium, there is a choice of three different base matrices accordingto pore accessibility of target molecules (pdf)BioToolomics:SepFast Capture Q, SepFast Capture DEAE, SepFast Capture S and SepFast Capture CM are,respectively, strong anion, weak anion, strong cation and weak cation exchange adsorbents. They are specially designed for cost-effective large-scalecapturing of biological molecules in the initial downstream recovery step.For each type of ion exchange medium, there is a choice of three different base matrices accordingto pore accessibility oftarget molecules (pdf)BioToolomics: SepFast Supor Q a strong anion exchange chromatography product. The working medium possesses a combination of small pore (50-100nm) and large pore (micro level). It shows fast accessibility to both small and large molecules (such as endotoxin, DNA, virus and virus like particles)(pdf) CryoBioPhysicapISep buffers for separation of proteins by anion or cation exchange chromatography, applying usercontrol external pH gradients. Can be apply to all kind of ion exchange columns, together with NaCl and Urea gradients. pISep software to calculate pH gradients in the presence of salt, urea and somenon-ionic detergents. Introduction (pdf) and overview of technology (pdf)G-BioscienceG-Sep Ion Exchange Agarose Fast Flow: CM, DEAE, Q SP Agarose Fast Flow(pdf)GE-HEALTHCAREIon Exchange Chromatography Manual(pdf) (pdf new) GE-HEALTHCARE Ion Exchange Cromatography and Chromatofusing Handbook (pdf) GE-HEALTHCAREIon Exchange Media - Selection Guide(pdf)(pdf-II) (pdf-III)GE-HEALTHCARE Recommended buffers for Anion Exchange Chromatography GE-HEALTHCARERecommended buffers for Cation Exchange Chromatography GE-HEALTHCARE Recommended Volatile buffers systems for Ion Exchange Chromatography GE-HEALTHCARECapto adhere: a strong anion exchanger with multimodal functionality ionic interaction, hydrogen bonding and hydrophobic interaction. Capto adhere is designed for post Protein A purification of monoclonal antibodies. Removal of leached Protein A, aggregates, host cell proteins, nucleic acids and viruses from monoclonal antibodies is performed in flowthrough mode at where the antibodies pass directly through the column while the contaminants are adsorbed. (pdf-I) (pdf-II) (pdf-III) GE-HEALTHCARE Capto MMC: a multimodal cation exchanger. Has a multimodal ligand that may interact with target molecules in several different ways. It contains a carboxylic group and thus its features partly resemble those of a weak cation exchanger. In addition to the ionic interactions several other types of interactions involved, including hydrogen bonding and hydrophobic interaction. The design of the ligand enables binding of proteins at high conductivity. (pdf-I) (pdf-II) (pdf-III)GE-HEALTHCARE MacroCap SP is a highly porous cation exchangerhigh available surface area for adsorption of of large biomolecules such as IgM and polyethylene glycol(PEG)-modified proteins (i.e., PEGylated proteins) that are intended for use as biopharmaceuticals.(pdf) GE-HEALTHCARE Q-Sepharose XL (anion exchange). Q-Sepharose XL virus licensed. SP-Sepharose XL (cation exchange)(pdf) GE-HEALTHCARE Capto Q a strong anion exchange medium for packed bed chromatography that allows increased speed and throughput in capture and intermediate purification. It combines high capacity with high flow velocity and low backpressure(pdf)GE-HEALTHCAREANX Sepharose 4 Fast Flow (high sub) is a weak anion exchanger with different selectivity. Larger pores than DEAE Sepharose Fast Flow which improves the dynamic binding capacity when separating larger molecules (pdf)GE-HEALTHCARECapto S ImpAct: Polishing of monoclonal antibodies. Good pressure/flow properties. Bead size of 50 m an optimized porosity. Gives high resolution. Mix of two different building blocks: a negatively charged sulfonate group and a neutral pyrrolidone: high binding capacity. (pdf)GE-HEALTHCARE Packing of IEX, Affinity or HIC column (movie) GE-HEALTHCARE Column evaluation (movie)BIORAD Macro-Prep Ion Exchange Support - Instruction Manual (pdf)BIORADUNOTMQ S Continuous (pdf) (pdf II)MERCK Cleaning and Regeneraton ofFractogel EMD sorbents (pdf)MERCKIon Exchange Chromatography Using Fractogel EMD Tentacle Supports (pdf) MERCKFractoprep DEAE Weak Anion Exchange chromatography(pdf)MERCKFractoprep SO3 - Strong Cation Exchange chromatography (pdf)MERCKFractoprep TMAE Strong Anion Exchange (pdf)MERCKFractogel EMD COO- (S) (M) Weak Cation Exchange (pdf) MERCK Fractogel EMD DEAE (S) (M) Weak Anion Exchange (pdf)MERCKFractogel EMD DMAE (S) (M) Weak Anion Exchange (pdf) MERCK Fractogel EMD SE High Capacity (M)Strong Cation Exchange (pdf)MERCKFractogel EMD SO3- (S) (M) Strong Cation Exchange (pdf) MERCKFractogel EMD TMAE (S) (M) Strong Anion Exchange (pdf) MERCK Fractogel EMD TMAEHigh Capacity (M) Strong Anion Exchange (pdf)MERCKEshmuno S is a cation exchanger specifically designed for highly productive downstream purification of monoclonal antibodies.Hydrophilicpolyvinyl ether base matrix that allows the use of much higher flow rates. (pdf) NATRIX NatriFlo HD-Q Membrane is an advanced material with athree-dimensional macroporous hydrogel structurethat provides a High Density of binding sites and rapid mass transfer. NatriFlo HD-Q Membranes deliver binding capacity that exceeds resin-based columns with fast flow-rates typical of membrane adsorbers.(pdf products) (pdf Instruction Guide)(pdf Method development) (pdf quick start guide)Adsept Cross Flow Technology (pdf).Adsept Process Technology High Performance Disposable Capture Chromatography (pdf)PallQ, S, DEAE, CM Ceramic HyperD ion exchangers. Of particular value in process scale application, these sorbents are designed to maintainhigh dynamic binding capacity (DBC) under conditions where conventional sorbents display significant capacity or productivity limitations. (pdf) Prepacked RPC columns (pdf)Pall Pall PRC Columns Prepacked Columns for Ion Exchange and Mixed-Mode Chromatography (pdf)PallHyperCel STAR AX Salt Tolerant Advanced Recovery Anion Exchange Chromatography Sorbent. Designed for high productivity protein capture at moderate or higher salt conductivity (2 to 15 mS/cm), typical of undiluted biological feedstocks (mammalian cell culture supernatants, E. coli feedstock, plasma, others) (pdf) Pall Purification of mouse IgM from cell culture supernatant by Cation Exchange Chromatography on CM Ceramic HyperD F sorbent (pdf) PallQ, S HyperCel ion exchangers. Different selectivities with improved productivity, available in prepacked PRC 1mL and 5mL column and in bulk. (pdf) Article describing comparative characteristics and advantages. (pdf-II) Pall Q Mustang XT capsules (5ml, 140ml and 5L volume): High throughput, scalable, and reusable ion exchange membrane chromatography. High capacities and high flow at low pressure. Contaminant removal for DNA, virus, host cell protein, and endotoxin. Plasmid, Virus, protein capture, and oligonucleotide purification (pdf I) (pdf II) (pdf III) Application Note: Purification of Influenza Virus by Ion Exchange Chromatography on Mustang Q XT Membranes (pdf IV)Pall Q, S Mustang Membrane Adsorbers. These single-use or reusable capsules can be used for small scale quick purification at lab scale withXT Acrodisc , or even scaled-up. Capacities are several times higher than traditional resins, especially for large molecules (DNA or viruses).No packing is needed and it is compatible with existing pumping systems, or can be used with a syringe. (pdf) PUROLITE Praesto SP and Praesto Q are available in 90 m, 65 m and 45 m particle sizes, covering the use of ion exchange in high-productivity capture steps as well as high-resolution polishing applications. Highly cross-linked, agarose-based ion exchange chromatography resins (pdf pg16) SARTORIUS SartobindTM - Membrane Adsorbers Discs and Cassettes. Reusable Sartobind Membrane Ion Exchangers for Adjustable Filter Holders (pdf) SARTORIUSSartobind Membrane Adsorbers - A Separation Technology Based on Microporous Membrane Ion Exchangers (pdf) SARTORIUSSartobind SingleSep Disposable Capsules - A Separation Technology Based on Microporous Membranes - Operating Instructions(pdf)THERMO: Strong Ion Exchange Spin Columns; use membrane-adsorbent technology as a chromatographic matrix to fractionate proteins.The spin column capacities are 4 mg proteins/peptides for the mini and 60-80 mg protein for the maxi. Actual capacity depends on the specific protein sample, selected pH and salt condition. (pdf) THERMO:ProPac line of ion-exchange columns from Dionex specifically to provide high-resolution, high-efficiency separations of proteins and glycoproteins(pI= 3 10; MW 10,000). Available with weak or strong anion-exchange or cation-exchange resins packed in 2-, 4-, 9-, and 22-mm i.d. formats. (pdf) TOSOH TSK-GEL BioAssyst Series Ion Exchange Column (pdf) TOSOH Ion Exchange Chromatography (pdf) TOSOH Chromatographic Media Catalog(pdf)TOSOHTOYOPEARL NH2-750F SALT TOLERANT ANION EXCHANGE RESIN (dpf) TOSOHTOYOPEARL SULFATE-650F SALT TOLERANT CATION EXCHANGE RESIN (pdf) VIVASCIENCE: Vivapure IEX kits include everything required for rapid purification of protein samples or contaminant removal from proteinsamples: clarification spin columns for initial sample clearing, Vivapure spin columns and ready-to-use buffers in different concentrations forthe protein bind-wash-elute steps, and Vivaspin ultrafiltration devices for final sample concentration and desalting. Basic and acidic proteinpurification kits. (pdf Brochure 1) (pdf Brochure 2) (pdf spin columns) Multimodal and Hydrophobic Charge-Induction Chromatography Test Tube for Multi Mode ResinsBioToolomics: SepFast MM AH-1 a mixed mode chromatography medium. The ligand contains a combination of anionic and hydrophobic groups.The product shows good binding capacity to molecules rich of hydrophobic moieties. It shows little binding to DNA or albumins under moderateionic strength (e.g. 0.15 M salt) (pdf) GE-HEALTHCARE Multimodal Chromatography Handbook (pdf)GE-HEALTHCARECapto adhere: a strong anion exchanger with multimodal functionality ionic interaction, hydrogen bonding and hydrophobic interaction. Capto adhere is designed for post Protein A purification of monoclonal antibodies. Removal of leached Protein A,aggregates, host cell proteins, nucleic acids and viruses from monoclonal antibodies is performed in flowthrough mode at where the antibodies pass directly through the column while the contaminants are adsorbed. (pdf-I) (pdf-II) (pdf-III) ImpRes (pdf-IV) GE-HEALTHCARE Capto MMC: a multimodal cation exchanger. Has a multimodal ligand that may interact with target molecules in several different ways. It contains a carboxylic group and thus its features partly resemble those of a weak cation exchanger. In addition tothe ionic interactions several other types of interactions are involved, including hydrogen bonding and hydrophobic interaction. The design ofthe ligand enables binding of proteins at high conductivity. (pdf-I) (pdf-II) (pdf-III) GE-HEALTHCARECapto core 700, a media for flow through applications in intermediate purification and polishing of viruses and other large biomolecules. By using Capto Core 700 it ispossible to remove contaminants Mr ~700kDa and allow for larger targets to be collected in the flowthrough. Capto Core 700 is targeted for flow through applications in intermediate purification of viruses and other large biomolecules. The novel core bead technology allows each bead in this medium to be designed with a ligand-activated core and an inactivate shell (a shell without ligands). inactive shell excludes large biomolecules from entering the corethrough the pores in the shell. These larger biomolecules are therefore collected in the flowthrough while smaller contaminants enter through the pores in the shell and bind to the internalized ligands. The use of Gel filtration chromatography is a bottleneck in vaccine processes (polishing steps) because of the low column loading and slow flow rates. Capto Core 700 offer a high productivity solution to size exclusion chromatography in this field. (pdf-data file) (pdf-instructions) (pdf-Influenza purific) MERCK Eshmuno S resin and Eshmuno Q resin. Tentacle structure with the new hydrophilic polyvinyl ether base matrix. (pdf) PallMEP HyperCel was the first modern Mixed-Mode media on the market, combining hydrophobic interaction for binding and ionic repulsion for eluting.It can help purify antibodies and fragments as well as recombinant proteins (enzymes and others) with less or even no salt compared to HIC, whileallowing an anionic repulsion pH drop at low conductivity. Together with HEA and PPA chemistries, these 3 ligands can be screened at the same timesince providing different selectivities.(pdf) Pall Pall HEA and PPA HyperCel operate on a \"mixed-mode\" mechanism, based on a combination of electrostatic and hydrophobic propertiesof the protein and ligands. Ligands: aliphatic (HEA hexylamine) and aromatic (PPA phenylpropylamine) amines, which offer differentselectivity and hydrophobicity options (pdf)Poster on different selectivities (pdf) PallPall HEA and PPA HyperCel operate on a \"mixed-mode\" mechanism, based on a combination of electrostatic and hydrophobic properties of the protein and ligands. Ligands: aliphatic (HEA hexylamine) and aromatic (PPA phenylpropylamine) amines, which offer differentselectivity and hydrophobicity options.(pdf) Pall HyperCel STAR AX. Salt Tolerant Advanced Recovery Anion Exchange. Composed of a rigid cellulose matrix. (pdf-1) (pdf-2) Capture of Human Serum Albumin from Plasma (pdf) Pall Performance of mixed-mode cation exchange (CEX) CMM HyperCel sorbent, to both monoclonal antibody (mAb) polishing and recombinant protein purification. Behavior of the CMM HyperCel sorbent versus conventional cation exchange (sulfopropylgroups) and a weak cation exchange multi-modal sorbent (cross-linked agarose) in terms of dynamic binding capacity (DBC) and selectivity in aggregate removal (pdf)PallPurification of mIgG1 on MEP HyperCel Mixed-Mode media. Optimization (pdf)Pall Pall PRC Columns Prepacked Columns for Ion Exchange and Mixed-Mode Chromatography (pdf) TOSOH TOYOPEARL MX-Trp-650M: a new multimodal cation exchange resin for protein purification. It combines the selectivity options of mixed-mode chromatography with the binding capacity of modern ion exchange resins. Excellent choice for intermediate and polishing steps, such as aggregate removal in antibody purification. (pdf-1) (pdf-2)TOSOH Chromatographic Media Catalog(pdf) Phospho-Protein Purification CLONTECH: BD Phosphoprotein Kit User Manual(pdf)(pdf-II)(pdf-III) QIAGEN Phosphoprotein Purification Kit. For purification and analysis of phosphorylated proteins from eukaryotic cells(pdf-I)(pdf-II)THERMO Fe-NTA Phosphopeptide Enrichment Kit for efficient enrichment of phosphorylated peptides by spincolumns. Useful for Sample Prep for Mass Spec Analysis. (pdf) Polyubiquitin-modified proteins THERMO:Ubiquitin Enrichment Kit. For the isolation and study of intracellular polyubiquitin-modified proteins. Through the use of a high-binding affinity resin, polyubiquitinated proteins are isolated from cell or tissue lysates. The bound proteins can then be eluted from the affinity resin and analyzed using the anti-ubiquitin antibody. (pdf)Protein AggregationAdditives Used to Stabilize Folding and Prevent AggregationBuffer solubility screen to avoid aggregation during protein concentration and ultrafiltrationProtein Expression Facility of The Hebrew University of Jerusalem: Heat shock growth procedure Lebendiker M., and Danieli T. (2014) Production of prone to aggregate proteins.FEBS Lett. (2014) 588:236-246 , http://dx.doi.org/10.1016/j.febslet.2013.10.044 (pdf)Lebendiker M., Maes M. and Friedler A (2015)A screening methodology for purifying proteins with aggregation problems. Methods in Molecular Biology: Insoluble Proteins book (Springer) 1258: 261-281(pdf)AVACTAOptim 1000 developed to reduce the time and cost of therapeutic protein pre-formulation studies, stability testing and formulation.Thermal unfolding and aggregation curves are simultaneously acquired for 48 samples run in a single experiment, enabling 96 samples to be analysed in one working day. Sample volumes as low as 1 l. Simultaneous optical measurements: 1) Intrinsic fluorescence to monitor tertiary structure 2) Static light scattering to detect aggregates. Sample heating and cooling to determine: 1) Protein unfolding temperature (Tm) 2) Protein aggregation onset temperature (Tagg) 3) Time dependent unfolding and aggregation at a fixed temperature(pdf) BioTek and ENZO ProteoStat Protein aggregation assay provides a simple, homogenous assay format for monitoring protein aggregation in a microplate assay format. The assay can be employed to streamline protein processing and optimize formulation procedures; in a wide pH and ionic strength range. The Synergy Mx Multi-Mode Microplate Reader is used for all assays. Its quadruple monochromator system in top-reading mode is used with slit widths of 9 nm. The excitation monochromator is set to 500 nm and emission to 600 nm. (pdf-manual) (pdf-I) (pdf-II)DILYX Biotechnologies OptiSol Protein Solubility Screening Kit Application Manual. Array-based filtration technology that enables to either: 1) identify formulations that protect a target protein from aggregation, or 2) gently solubilize an aggregated protein sample. This screening kit contains a systematically varied array of buffers (from pH 3 to pH 10) and a series of solubility enhancers (salts, amino acids, sugars, polyols, reducing reagents) that enable the determination of conditions under which a particular protein sample is protected from aggregation or can be de-aggregated. Filtration principle: soluble proteins pass the filter due to their smaller size, while aggregated proteins do not pass filter. (pdf-1) (pdf-2)OptiResc Reagent Listing (pdf-3) HAMPTON: Solubility Stability Screen is designed to assist in the identification of solution conditions which promote protein solubility and stability, and minimize protein precipitation. Solubility Stability Screen is a solubility screen, a stability screen, and may also be used as an additive screen in the presence of a crystallization reagent.(pdf-I)(pdf-II) INTEGRITY BIOSOLUTION Protein Aggregation (page) NOVAGEN Enhancing solubility during protein expression in E. Coli (site) (site-II)Bondos Detection and prevention of protein aggregation before, during, and afterpurification. Analytical Biochemistry (2003), 316 (2 ), 223-231 (pdf)Cromwell M., Protein Aggregation and Bioprocessing The AAPS Journal 2006; 8 (3) Article 66 pg.E-572 (pdf)Hamada H., Effect of additives on Protein Aggregation Current Pharmac. Biotech. (2009), 10, 400-407 (pdf)Lebendiker M., and Danieli T. (2014) Production of prone to aggregate proteins.FEBS Lett. (2014) 588:236-246 , http://dx.doi.org/10.1016/j.febslet.2013.10.044 (pdf)Lebendiker M., Maes M. and Friedler A (2015)A screening methodology for purifying proteins with aggregation problems. Methods in Molecular Biology: Insoluble Proteins book (Springer) 1258: 261-281(pdf)Philo J.,Mechanisms of Protein Aggregation Current Pharmac. Biotech. (2009), 10, 348-351 (pdf)Pullara F.et al, A general path for large-scale solubilization of cellular proteins: From membrane receptors to multiprotein complexesProtein Expression and Purification 87 (2013) 111 119 (pdf )(pdf Supplement) Protein Labeling with Biotin MERCK INNOLINK Biotin 354S: A modified biotin conjugate in which biotin is linked to a bis-aryl hydrazone chromophore an aromatic succinimidyl ester via a long-chain PEG4 linker. The bis-aryl hydrazone chromophore allows for direct spectrophotometric quantitation of total incorporated biotin. The long-chain PEG4 linker preserves biotin/avidin affinity as well as increases solubility. The aromatic succinimidyl ester allows for higher efficiency modification of amines in aqueous buffers. MOLECULAR PROBESBiotin-XX Microscale Protein Labeling Kit. This kit has been optimized for labeling small amounts (20 100 g) of purified proteins with molecular weights between 12 and 150 kDa, and contains everything needed to perform three labeling reactions and to separate the resulting conjugates from excess reactive biotin.(pdf) MOLECULAR PROBES FluoReporter Biotin Quantitation Assay Kit for biotinylated proteins. Sensitive fluorometric assay for accurately determining the number of biotin labels on a protein. The assay can detect from 4 to 80 pmol of biotin in a sample. (pdf) THERMO EZ-Link NHS-Biotin Reagents. N-Hydroxysuccinimide (NHS) esters of biotin are the most popular type of biotinylation reagent. NHS-activated biotins react efficiently with primary amino groups (-NH2) in pH 7-9 buffers to form stable amide bonds (pdf)Protein Refolding - Inclusion Bodies GE-HEALTHCAREPurifying Challenging Proteins: Membrane proteins, Multiprotein complexes and Inclusion bodies (pdf) AVIDIS-TECHNOLOGY Refolding Chromatography Refolding Chromatography with Mini-Chaperones(pdf) AthenaESQuickFold Protein Refolding Kit with a Detergent and a Cyclodextrin Screening Kit(pdf)BIO-VECTRAVectrase AT - Folding Proteins with Disulfide Bands BIO-VECTRAVectrase CD - Quick Screening of Refolding Conditions(pdf) BIO-VECTRA Vectrase DK - Refolding at High Protein Concentration BIO-VECTRAVectrase P - Protein Disulfide Isomerase (PDI) Mimic (pdf-I)(pdf-II)GENO-Protein Foldase-Protein Folding Optimization Kit NORGEN The ProteoSpin Inclusion Body Isolation Kit. Facilitates the isolation of recombinant proteins in the form of inclusion bodies from E. coli. The kit includes reagents specially formulated to achieve rapid and high-quality purification of inclusion body proteins using three processes: 1. Lysis of bacterial cells to release inclusion bodies in solid form 2. Solubilization of purified inclusion bodies 3. Purification of the recombinant protein using spin column chromatography with Norgen s proprietary resin as an ion-exchanger Each spin column is able to purify up to 12 mg of recombinant proteins from 100 mL of culture. The kit is designed to purify both acidic and basic proteins. (pdf Maxi Kit) (pdf Micro Kit) NOVAGEN Enhancing solubility during protein expression in E. Coli (site) (site-II) NOVAGEN-Protein Refolding kit (pdf) NOVAGENInformation on Protein Refolding (pdf)NOVAGEN iFOLD Protein Refolding System . The system includes inclusion body purification reagents combined with a dispensed 96-well plate-based protein refolding buffer matrix. (pdf-I)(pdf-II) NOVEXINThe technology and reagents protect the protein during vulnerable steps in the refolding process, providing an opportunity for the protein to refold corrctly, reducing losses due to aggregation.Employs linear carbohydrate polymers of ~5kDa that enhance protein solubility and stability through the formation of reversible complexes with proteins without altering their structure and preventing aggregation.Protein Refolding Starter Kit (pdf) Stability P.A.C. (pdf)Acidic Protein Refolding Kit (pdf)Basic Protein Refolding Kit (pdf) Refolding Kit for Histidine-tagged proteins (pdf-I) (pdf-II)Bulk Protection and Release Reagents (pdf)THERMOPro-MatrixTM Protein Refolding Kit(pdf) PROTEIN EXPRESSION FACILITY OF THE HEBREW UNIVERSITY OF JERUSALEM Heat shock growth procedure PROTEIN EXPRESSION AND PURIFICATION FACILITY OF THE EUROPEAN MOLECULAR BIOLOGY LABORATORY In vitro denaturation and refoldingAdditives Used to Stabilize Folding and Prevent Aggregation Protein Refolding on IMAC resin - Batch Screening Procedure - On-Column Scale-upContaminant Removal from Inclusion Bodies Before Solubilization UNIVERSITY OF OKLAHOMA School of Chemical Engineering and Materials ScienceRecombinant Protein Solubility Prediction Recommended Reading Altamirano M., Refolding Chromatography with Immobilized Mini-Chaperones. PNAS 1997, 94: 3576-3578(pdf)Armstrong N., A New Protein Folding Screen...etc. Protein Science 1999, 8: 1475-1483 (pdf) Chen G., Overexpression of a Glutamate Receptor (GluR2) ligand binding domain in E.Coli: Application of a novel protein folding screen (pdf)De Bernardez Clark E.,Refolding of RecombinantProteins . Current Opinion in Biotechnology 1998,9:157 163 (pdf)De Bernardez Clark E., Protein refolding for industrial processes. Current Opinion in Biotechnology 2001, 12:202 207 (pdf).Eiler S., Overexpression, Purification, and Crystal Structure of Native ER alphaLBD Protein Expression and Purification (2001) 22, 165 173 (pdf) Machida S., Cycloamylose as an efficient artificial chaperone for protein refolding. FEBS Letters 486 (2000) 131-135 (pdf) Middelberg A., Preparative Protein Folding. TRENDS in Biotechnology 2002, 20 (10): 437-443 (pdf) Ming Li et al., In vitro protein refolding by chromatographic procedures. Protein Expr. and Purif. 2004,33: 1-10 (pdf) Tsumoto K., Practical Considerations in Refolding Proteins from Inclusion Bodies. Protein Expr. and Purif. 2003,28: 1-8 (pdf) Reverse Phase Chromatography GE-HEALTHCAREHydrophobic Interaction and Reversed Phase Chromatography Principles and Methods (pdf)GE-HEALTHCAREReverse Phase Chromatography Manual (pdf)PIERCE:Peptide solubility guidelines. Use amino acid characteristics to predict hydrophobicity. (pdf)TOSOH BIOSCIENCEReverse Phase Chromatography Brochure (pdf) (pdf - new)VYDACThe Handbook of Analysis and Purification of Peptides and Proteins by Reverse-Phase HPLC(pdf) Simulated Moving Bed Chromatography SEMBA Biosciences: Poster: Continuous highly efficient protein purification using simulated moving bed chromatography (pdf)SEMBA Biosciences:Application Note: The Semba Octave Chromatography System. Simulated moving bed chromatography. Continuous AffinityPurication of Recombinant Proteins (pdf) SEMBA Biosciences:The Semba Octave Chromatography System. A simulated moving bed chromatography system. Continuous unattended purification, milligrams to grams. Compatible with chemical and biological samples. SMBC emulates countercurrent separation where the mobile opposite direction of the stationary phase. The stationary phase is represented by individual columns connected in series, and the mobile phase by inlet streamsof Feed and Desorbent and outlet streams of Raffinate and Extract. (pdf) Storage of Purified Proteins PROTEIN EXPRESSION AND PURIFICATION FACILITY OF THE EUROPEAN MOLECULAR BIOLOGY LABORATORY Storage of Purified Proteins THERMO Protein stability and storage (pdf) Thiophilic Chromatography CLONTECH: Thiophilic - Purification of Immunoglobulins (pdf)CLONTECH: Protein Purification Products(pdf) G-BioscienceThiophilic Adsorption: For the Purification of Immunoglobulins with Thiophilic Resin(pdf) GE-HEALTHCARE Activated Thiol Sepharose 4B reacts with solutes containing thiol groups under mild conditions to form mixed disulphides. This reaction forms the basis of covalent chromatography and a procedure for immobilizing thiol containing biomolecules. (pdf) GE-HEALTHCAREThiopropyl Sepharose (pdf)MERCK Fractogel Thiophilic(pdf)THERMO T-Gel Purification Kit (pdf) MILLIPOREPROSEP-Thiosorb and PROSEP-Thiosorb M Chromatography Media - for the recovery and purification of Immunoglobulins (pdf) Viral PurificationADEMTECHViro-Adembeads are specifically developed magnetic particles to capture viruses from biological samples. Following incubationin a virus containing sample, Viro-Adembeads associated to viruses are recovered and can be directly used to infect target cells increasing infection efficiency. The mechanism of virus capture is based on electrosteric interactions(pdf-I) (pdf-II) BIA Separations CIM Monolithic Columns based on CIM Convective Interaction Media Technology; suitable for purification of large biomolecules such as viruses(viral vectors and vaccines), DNA (plasmid DNA) and larger proteins (Immunoglobulins G and M, pegylated proteins). CIM Monolithic Columns exhibit unrivaled characteristics in terms of operational flow rates, binding capacity and separation resolution for large biomolecules. Products are used in research, laboratory, pilot andindustrial production stages and are extremely simple to use, with no packing of columns needed(pdf-I) (pdf-II) (pdf-III) GE-HEALTHCARE Q-Spharose XL (anion exchange). Q-Spharose XL virus licensed. SP-Spharose XL (cation exchange) (pdf)GE-HEALTHCARE Rapid Adenovirus Purification Using Q-Spharose XL(pdf-I) or Source 15Q (pdf-II)GE-HEALTHCARE AVB Sepharose High Performance is an affinity medium designed for the purification of adeno associatedvirus (AAV). The ligand, a 14 kD recombinant protein, is attached to the base matrix via a long, hydrophilic spacer arm to make it easily available for binding of the virus (pdf)GE-HEALTHCARECapto core 700, a media for flow through applications in intermediate purification and polishing of viruses and other large biomolecules. By using Capto Core 700 it ispossible to remove contaminants Mr ~700kDa and allow for larger targets to be collected in the flowthrough. Capto Core 700 is targeted for flow through applications in intermediate purification of viruses and other large biomolecules. The novel core bead technology allows each bead in this medium to be designed with a ligand-activated core and an inactivate shell (a shell without ligands). inactive shell excludes large biomolecules from entering the corethrough the pores in the shell. These larger biomolecules are therefore collected in the flowthrough while smaller contaminants enter through the pores in the shell and bind to the internalized ligands. The use of Gel filtration chromatography is a bottleneck in vaccine processes (polishing steps) because of the low column loading and slow flow rates. Capto Core 700 offer a high productivity solution to ize exclusion chromatography in this field. (pdf-data file) (pdf-instructions) (pdf-Influenza purific) GE-HEALTHCAREDownstream process development for efficient purification of adenovirus (pdf) CLONTECH: Adeno-X Virus Purification Kits User Manual (pdf-I) Protocol (pdf-II). A complete filtration-based system for purifying and concentrating recombinant adenovirus. It provides a superior alternative to cesium chloride (CsCl) density gradient centrifugation. Use an adsorbent membrane that selectively binds adenoviral particles based on their distinctive surface-associated properties.CHISSO CorpMILLIPORE Matrex Cellufine Sulfate - For concentration, purification and depyrogenation of Virus, Viral/Microbial Antigen, Heparin binding proteins (pdf-I)(pdf_II) References: (pdf-III) PALLDynamic High Capacity Mustang Q Membrane Units for Scaleable Anion Exchange Chromatography Purification of Adenoviral Vectors (pdf) PURESYNPuresyn s Adenopure Kit couples membrane adsorber technology attaches ion exchange functional groups to the inner surfaceof synthetic microporous membranes in a syringe-filter format(pdf) THERMOPOROS CaptureSelect AAV Affinity Resins AAV8 and AAV9 (pdf) VIVASCIENCE: Vivapure AdenoPACK 100. Adenovirus (Ad5) purification and concentration kit for up to 100 ml cell culture volume. AdenoPACK syringe filters containing an ion exchange membrane adsorber that binds adenoviral particles. (pdf-I) (pdf-II) (pdf-III)