[Skip to Content](https://www.sigmaaldrich.com#main-content) [](https://www.sigmaaldrich.com/US/en) Products Cart0 USEN Products ProductsApplicationsServicesResourcesSupport [Login / Register](https://www.sigmaaldrich.com/oidc-sign-in) [Order Lookup](https://www.sigmaaldrich.com/US/en/order-lookup) [Quick Order](https://www.sigmaaldrich.com/US/en/quick-order) Cart0 [Home](https://www.sigmaaldrich.com/US/en)[Applications](https://www.sigmaaldrich.com/US/en/applications)[Protein Biology](https://www.sigmaaldrich.com/US/en/applications/protein-biology)Protein Purification # Protein Purification  Numerous protein purification methods are widely used in both biological and biomedical research. Recombinant protein expression and purification workflows depend on many variables. These variables include but are not limited to the physical properties and biological function of the protein, and whether a bacterial or eukaryotic cell line should be used to express the protein of interest. Significant advancements have been made in the area of recombinant [protein expression](https://www.sigmaaldrich.com/US/en/applications/protein-biology/protein-expression)  and purification methodology along with a plethora of commercially available systems and kits. However, proteins are complex macromolecules, and optimal protein expression and purification strategies must be determined empirically.   * * * ## Related Products Slide 1 of 13 1 of 3 [ \ Sigma-Aldrich \ F4799 \ 3X FLAG® Peptide](https://www.sigmaaldrich.com/US/en/product/sigma/f4799) Quick View [ \ Millipore \ F3290 \ FLAG® Peptide](https://www.sigmaaldrich.com/US/en/product/sigma/f3290) Quick View [ \ Millipore \ A4596 \ ANTI-FLAG® M1 Agarose Affinity Gel](https://www.sigmaaldrich.com/US/en/product/sigma/a4596) Quick View [ \ Millipore \ A2220 \ ANTI-FLAG® M2 Affinity Gel](https://www.sigmaaldrich.com/US/en/product/sigma/a2220) Quick View [ \ Millipore \ M8823 \ Anti-FLAG® M2 Magnetic Beads](https://www.sigmaaldrich.com/US/en/product/sigma/m8823) Quick View [ \ Millipore \ F2426 \ EZview™ Red ANTI-FLAG® M2 Affinity Gel](https://www.sigmaaldrich.com/US/en/product/sigma/f2426) Quick View [ \ Millipore \ TP0010 \ FLAG® HA Tandem Affinity Purification Kit](https://www.sigmaaldrich.com/US/en/product/sigma/tp0010) Quick View [ \ Millipore \ CELLMM2 \ FLAG® M Purification Kit](https://www.sigmaaldrich.com/US/en/product/sigma/cellmm2) Quick View [ \ Sigma-Aldrich \ P7582 \ Amino-terminal FLAG-BAP™ Fusion Protein](https://www.sigmaaldrich.com/US/en/product/sigma/p7582) Quick View [ \ Millipore \ P5975 \ Amino-terminal Met-FLAG-BAP™ Fusion Protein](https://www.sigmaaldrich.com/US/en/product/sigma/p5975) Quick View [ \ Sigma-Aldrich \ P7457 \ Carboxy-terminal FLAG-BAP™ Fusion Protein](https://www.sigmaaldrich.com/US/en/product/sigma/p7457) Quick View [ \ Millipore \ H9914 \ HIS-Select® Nickel Magnetic Agarose Beads](https://www.sigmaaldrich.com/US/en/product/sigma/h9914) Quick View [ \ Millipore \ P6611 \ HIS-Select® Nickel Affinity Gel](https://www.sigmaaldrich.com/US/en/product/sigma/p6611) Quick View * * * ## Featured Categories [](https://www.sigmaaldrich.com/US/en/products/chemistry-and-biochemicals/biochemicals/biological-buffers) [Biological Buffers](https://www.sigmaaldrich.com/US/en/products/chemistry-and-biochemicals/biochemicals/biological-buffers) Our extensive portfolio of high-purity biological buffers in various formulation and packaging formats provide superior solution stability and pH control throughout your bioprocess workflow applications. [Shop Now](https://www.sigmaaldrich.com/US/en/products/chemistry-and-biochemicals/biochemicals/biological-buffers) [](https://www.sigmaaldrich.com/US/en/products/chemistry-and-biochemicals/biochemicals/detergents) [Detergents - Anionic, Cationic, Zwitterionic, Anti-foaming](https://www.sigmaaldrich.com/US/en/products/chemistry-and-biochemicals/biochemicals/detergents) Biological detergents and surfactants, including REACH-compliant biodegradable alternatives, are available for cell lysis, DNA/RNA electrophoresis, Western blotting, transfection, and many other life science research applications. [Shop Now](https://www.sigmaaldrich.com/US/en/products/chemistry-and-biochemicals/biochemicals/detergents) Overview Related Articles & Protocols Support ## Critical Factors for Protein Purification Protein structure and function are often critical factors to consider when selecting a protein purification strategy. Recombinant protein biochemical or biological activity is partially determined by discrete domains within the protein, of which are often reliant on the protein being folded into secondary, tertiary, and quaternary structures.  Protein folding is collectively referred to as the higher-order structure (HOS) and is essential for the correct three-dimensional shape and function of the protein. Additionally, protein solubility is a highly desirable attribute for successful protein purification and is influenced by numerous factors including, size and N- and C-terminal elements. Recombinant proteins commonly incorporate N- and C-terminal tags, which are small sequences that are used for immunohistochemical detection and purification, or protein affinity chromatography, depending on the specific N- and C-terminal tag and intended downstream application. ## Protein Purification Methods and Applications Whether researchers aim to study protein function or seek to scale up protein purification using strategies for downstream, industrial-scale biologics and pharmaceutical production, there are numerous protein purification methods, reagents, and tools available. The selected protein purification method will partially determine the sample preparation workflow. Affinity chromatography is a suitable initial purification step for purifying solubilized recombinant proteins that contain relevant tags; however, unwanted proteins are likely to also bind to the affinity resin column and will elute in the final wash along with the desired protein of interest. If additional purification is necessary, supplementary purification strategies are employed including, size-exclusion chromatography or ion exchange chromatography. Importantly, many affinity tags can be removed as researchers may want to remove any non-native sequences from the final purified protein. [Shop All Protein Purification Products](https://www.sigmaaldrich.com/search/facet-search?focus=products&page=1&perpage=30&sort=relevance&term=facet-search&type=product) [](https://www.sigmaaldrich.com/documents-search) [Looking for More Specific Information?](https://www.sigmaaldrich.com/documents-search) Visit our document search for data sheets, certificates and technical documentation. [Find Documents](https://www.sigmaaldrich.com/documents-search) ## Related Articles - [Immunoprecipitation of FLAG Fusion Proteins Using Monoclonal Antibody Affinity Gels](https://www.sigmaaldrich.com/US/en/technical-documents/protocol/protein-biology/protein-purification/flag-immuno-precipitation-protocol) Protocol for immunoprecipitation (IP) of FLAG fusion proteins using M2 monoclonal antibody 4% agarose affinity gels - [Desalting/Buffer Exchange and Concentration for Affinity Chromatography of Tagged Proteins](https://www.sigmaaldrich.com/US/en/technical-documents/protocol/protein-biology/protein-concentration-and-buffer-exchange/desalting-buffer-exchange-concentration) This page shows how to perform sample desalting, buffer exchange and concentration for affinity chromatography of tagged proteins. - [Solubilization of Membrane Proteins](https://www.sigmaaldrich.com/US/en/technical-documents/protocol/protein-biology/protein-purification/solubilization) This page shows how to solubilize membrane proteins with products from Cytiva. - [Column Packing and Preparation](https://www.sigmaaldrich.com/US/en/technical-documents/protocol/protein-biology/protein-purification/column-packing-and-preparation) This page describes efficient column packing and preparation for affinity chromatography of tagged proteins using Cytiva products. - [Sepharose Fast Flow: Purification with Good Resolution and Easy Scale-Up](https://www.sigmaaldrich.com/US/en/technical-documents/protocol/protein-biology/protein-purification/sepharose-fast-flow-good-resolution) This page covers the use of Sepharose Fast Flow for purification of proteins. - [See All (19)](https://www.sigmaaldrich.com/US/en/search/facet-search?focus=sitecontent&term=facet-search) ## Related Protocols - [GenElute™-E Single Spin DNA and RNA Purification Kits for Green Labs](https://www.sigmaaldrich.com/US/en/technical-documents/technical-article/protein-biology/protein-purification/genelute-e-sustainability) Reduce plastic waste and eliminate hazardous liquid waste for more sustainable laboratories with GenElute™-E Single Spin DNA and RNA prep kits. - [Optimizing Purification of Histidine-Tagged Proteins](https://www.sigmaaldrich.com/US/en/technical-documents/technical-article/protein-biology/protein-purification/optimizing-purification-of-histidine-tagged-proteins) How to optimize purification of histidine-tagged proteins using Cytiva products. - [Immobilized Metal Chelate Affinity Chromatography (IMAC)](https://www.sigmaaldrich.com/US/en/technical-documents/technical-article/analytical-chemistry/purification/metal-chelate-afnity-chromatography) How to separate proteins and peptides with affinity for metal ions by immobilized metal chelate affinity chromatography. - [Manual Purification of Histidine-Tagged Proteins using His GraviTrap™ TALON® Columns](https://www.sigmaaldrich.com/US/en/technical-documents/protocol/protein-biology/protein-purification/manual-purification-using-his-gravitrap-talon) His GraviTrap™ TALON® columns are prepacked with 1 ml of TALON® Superflow medium. Each column provides simple manual purification of up to 15 mg of histidine-tagged proteins. - [Column Packing and Preparation for Hydrophobic Interaction and Reversed Phased Chromatography](https://www.sigmaaldrich.com/US/en/technical-documents/protocol/protein-biology/protein-purification/column-packing-and-preparation2) This page discusses column packing and preparation techniques for reverse phase chromatography. - [See All (4)](https://www.sigmaaldrich.com/US/en/search/facet-search?focus=sitecontent&term=facet-search) ### Find More Articles and Protocols Enter Keywords Search ## How Can We Help In case of any questions, please submit a [customer support request](https://www.sigmaaldrich.com/US/en/support/customer-support) or talk to our customer service team: Email [custserv@sial.com](mailto:custserv@sial.com) or call +1 (800) 244-1173 ## Additional Support - [Calculators & Apps](https://www.sigmaaldrich.com/US/en/support/calculators-and-apps) Web Toolbox - science research tools and resources for analytical chemistry, life science, chemical synthesis and materials science. - [Customer Support Request](https://www.sigmaaldrich.com/US/en/support/customer-support) Customer support including help with orders, products, accounts, and website technical issues. - [FAQ](https://maestro.my.site.com/knowledgeportal/s/) Explore our Frequently Asked Questions for answers to commonly asked questions about our products and services. * * * __Featured Articles__ [ANTI-FLAG® M2 Magnetic Beads](https://www.sigmaaldrich.com/US/en/technical-documents/technical-article/protein-biology/protein-purification/anti-flag-m2) The FLAG® Expression System is a proven method to express, purify and detect recombinant fusion proteins. Sigma®, the proven provider of FLAG®, now offers a magnetic bead for immunoprecipitation, protein purification, and the study of protein-protein interactions. The ANTI-FLAG® M2 Magnetic Bead is composed of murine derived, ANTI-FLAG® M2 monoclonal antibody attached to superparamagnetic iron impregated 4% agarose beads, with an average diameter of 50 µm. The M2 antibody is capable of binding to fusion proteins containing a FLAG peptide sequence at the N-terminus, Met-N-terminus, or C-terminus locations in mammalian, bacterial, and plant extracts. [Practical Considerations for IEX Separation](https://www.sigmaaldrich.com/US/en/technical-documents/technical-article/protein-biology/protein-purification/iex-separation) This page covers detailed aspects of each step in an IEX separation to improve resolution and overall performance. [GenElute™-E DNA and RNA Purification Kits](https://www.sigmaaldrich.com/US/en/technical-documents/technical-article/protein-biology/protein-purification/genelute-e-animation) Animation showing technology principle of GenElute™-E single spin negative chromatography nucleic acid purification kits [Optimizing Purification of Histidine-Tagged Proteins](https://www.sigmaaldrich.com/US/en/technical-documents/technical-article/protein-biology/protein-purification/optimizing-purification-of-histidine-tagged-proteins) How to optimize purification of histidine-tagged proteins using Cytiva products. Top __Sign In To Continue__ To continue reading please sign in or create an account. 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