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Protein purification is an essential component of protein research. The study of protein function, structure, and interactions heavily relies on the purity and quality of the isolated protein of interest. Here we present to you a five-step workflow that will help you in your quest. Learn about methods and technologies for protein expression, protein extraction and preservation, protein purification, protein clean up, and protein quantitation and detection.
Researchers have many methodology choices when it comes to producing recombinant proteins for early-stage discovery research through large-scale production of biotherapeutic drugs, vaccine development, and structural studies. It is imperative to use the right protein expression system for the target protein and application of interest. We offer a wide selection of superior mammalian, insect, bacterial, and yeast protein expression systems to suit your research needs.
Our Gibco Expi Transient Expression Systems, available in mammalian (CHO-S, 293F cells) and insect (Sf9 cells) expression formats, are completely optimized systems that enable rapid, high-yield production of proteins.
Recombinant protein titers in ExpiCHO and other transient CHO expression systems. Expression levels of human IgG, rabbit IgG and erythropoietin in ExpiCHO and other transient CHO expression systems are shown. ExpiCHO titers range from 25–160 times those of other transient CHO expression systems.
Discover more about Transient Protein Expression Platforms
Visit Protein Expression Learning Center
Visit Protein Expression Basics Technical Reference Library
Product highlight | Description |
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ExpiCHO Expression System | The Gibco ExpiCHO Expression System is a completely optimized system consisting of ExpiCHO-S(TM) cells that have been adapted to high-density, serum-free suspension culture in ExpiCHO(TM) Expression Medium, along with specially designed transfection reagents and enhancers, that provide the highest yields possible in a transient system (up to 3g/L). That means you can start your research work in CHO cells and stay in CHO cells through discovery. |
Expi293 Expression System | The Gibco Expi293 Expression System is a completely optimized system consisting of Expi293F(TM) cells that have been adapted to high-density, serum-free suspension culture in Expi293(TM) Expression Medium, along with specially designed transfection reagents and enhancers, that enable recombinant 293-derived recombinant protein expression in just five to seven days, with a 2-10-fold increase in protein yields compared to previous generation transient expression systems |
ExpiSf Expression System | The ExpiSf Expression System is the first-ever chemically defined baculovirus-insect cell protein expression system, delivering superior yields (3x more protein) and consistent performance run after run using a fast, streamlined workflow. |
Make sure to check for the following things when getting started with an Expi transient expression system:
Product selection guides | Products |
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Transient protein expression systems | Expi293 ExpiCHO ExpiSf Expression Systems |
Mammalian protein expression | Expi293 ExpiCHO Expression Systems |
Insect protein expression | ExpiSf Expression System |
Bacterial protein expression | Champion pET Expression System |
Yeast protein expression | PichiaPink Yeast Expression System |
Structural Biology and Membrane Protein Expression | Expi293 GnTI- Expi293 Inducible Expi293 Inducible GnTI- Expression Systems Expi293 Met(-) Protein Labeling Kit |
Protein expression selection guide | Protein Expression Systems Selection Guide |
Protein expression services | Gene-to-Protein Baculovirus Expression Mammalian Expression services |
Protein extraction techniques vary depending on the source of the starting material, the location of the protein of interest within the cell, and the downstream application. Other important considerations include the preservation of protein activity and function as well as the reduction of background effects.
Historically, mechanical disruption has been used to lyse cells and tissues; our gentle, detergent-based solutions have been developed to efficiently lyse cells and enable the separation of subcellular structures without requiring physical disruption, providing high yields of active proteins.
Protein extraction product features:
Protein extraction efficiency from major cellular compartments using M-PER Mammalian Protein Extraction Reagent. Lysates from established cell lines and primary cultures were prepared using M-PER reagent and extraction efficiency from the various cellular compartments evaluated. For each target protein, 10 µg of lysate was loaded for and electrophoresed by SDS PAGE, transferred to nitrocellose membrane and detected by western blot using SuperSignal West Pico PLUS Chemiluminescent Substrate.
Protein yield from various cell types using M-PER Mammalian Protein Extraction Reagent. Cells were harvested at 85% confluency, washed twice and collected in ice-cold PBS and counted. For each cell type, 1 x 106 cells were pelleted by centrifugation at 2,000 x g for 5 minutes and lysed in 1 mL M-PER Reagent for 5 minutes. The cell lysates were clarified by centrifugation at 14,000 x g for 10 minutes and the supernatant was collected and the protein concentration (µg/million cells) was determined using the Pierce BCA Protein Assay.
Improved protein yield using the Thermo Scientific Mem-PER Plus Membrane Protein Extraction Kit. Membrane proteins were isolated from mouse liver tissue and HeLa cells using Thermo Scientific Mem-PER Plus Membrane Protein Extraction Kit and three other commercial extraction kits. Protein yields (μg) for membrane, cytosolic, and total fractions were determined using the Thermo Scientific Pierce BCA Protein Assay Kit.
Tissue cell lysis protein yield with T-PER Tissue Protein Extraction Reagent. Duplicate tissue samples were weighed, resuspended in 1:10 to 1:20 w/v T-PER Reagent and disrupted in a chilled Dounce or benchtop tissue homogenizer. The resulting lysates were centrifuged at 10,000 x g for 5 minutes and the supernatant was collected. The protein concentration of each lysate was determined using the Pierce BCA Protein Assay to determine protein yield per milligram of starting tissue.
Protein yield comparison of two bacterial cell lysis reagents.E. coli ER2566/pLATE51-Klenow, ER2566/pGST-CC-StpB, and ER2566/pGS-Syk cell pellets (0.5 g), were resuspended in 2.5 mL aliquots of Thermo Scientific B-PER Complete Bacterial Protein Extraction Reagent or EMD Chemicals BugBuster Master Mix with gentle vortexing for 15 minutes at room temperature. Insoluble cell debris was removed by centrifugation at 16,000 x g for 20 minutes at 4°C. Protein yields (concentrations) for soluble fractions were determined using the Pierce BCA Protein Assay Kit.
Sample type | Goal | Product highlight | |
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Primary or cultured mammalian cells or tissues | Total protein extraction | M-PER reagent T-PER reagent N-PER reagent | RIPA Lysis and Extraction Buffer IP Lysis Buffer |
Cultured mammalian cells or tissues | Subcellular fractionation or organelle isolation | NE-PER reagent Subcellular Fractionation Kits Mitochondria Isolation Kits | GPCR Extraction and Stabilization Reagent Cell Surface Protein Isolation Kit Syn-PER Reagent Lysosome Enrichment Kit |
Bacterial cells | Total protein extraction | B-PER reagent | |
Yeast cells | Total protein extraction | Y-PER reagent | |
Insect cells (baculovirus) | Total protein extraction | I-PER reagent | |
Plant tissue (leaf, stem, root, flower) | Total protein extraction | Plan Total Protein Extraction Kit |
Detergents are frequently used in cell lysis reagent formulation and other protein research methods. Thermo Scientific Surfact-Amps Detergent Solutions are highly purified, precisely diluted (10%) formulations that are ideal for applications or assays that are sensitive to contaminants present in unpurified detergents.
Protein detergent product features:
Generic structure of a detergent molecule.
Cell lysis disrupts cell membranes and organelles, resulting in unregulated enzymatic activity that can reduce protein yield and function. To prevent these negative effects, protease and phosphatase inhibitors can be added to the lysis reagents. Numerous compounds have been identified that can inactivate or block the activities of proteases and phosphatases.
Protease and phosphatase inhibitor product features:
Performance comparison between three commercially available protease inhibitor tablets. Pancreatic extract (100 μL; 0.5 μg/μL) was incubated with quenched fluorescent protease-cleavable substrates for trypsin, cysteine, and metalloprotease and cathepsins, in the presence of the reformulated Thermo Scientific Pierce Protease Inhibitor Mini Tablets, Roche™ Complete™ Protease Inhibitor Tablets, and Sigma-Aldrich™ SIGMAFAST™ Protease Inhibitor Cocktail Tablets, with and without EDTA. Reactions were incubated for 1 hr at 37ºC, and fluorescence was determined at the appropriate emission wavelengths. The percent inhibition is shown for each protease inhibitor formulation.
Protein phosphorylation in cell extracts is broadly preserved by Thermo Scientific Phosphatase Inhibitor Mini Tablets.(A) HCT116 cells were serum-starved, then either treated with EGF for 15 min or left as control cells. Cell lysates were prepared in Thermo Scientific Pierce IP Lysis Buffer with Thermo Scientific Protease and Phosphatase Inhibitor Mini Tablets, EDTA-Free, or with no inhibitor. Lysate containing 500 μg of protein was then incubated with 5 μg of phospho-tyrosine antibody overnight at 4ºC. The complex was then incubated with Thermo Scientific Pierce Protein A/G Magnetic Beads for 1 hr at room temperature. Beads were washed, and low-pH elution was performed. The eluates were subjected to western blotting, and the membrane was then probed with EGFR antibody for chemiluminescence detection. (B) The degree of inhibition for protein, alkaline, and acid phosphatase activity was determined in kidney extract (25 μL; 0.5 μg/μL) by incubating extracts with a fluorogenic substrate (MFP or FDP) that measures phosphatase activity upon dephosphorylation in the presence of Pierce Phosphatase Inhibitor Mini Tablets, Roche™ PhosStop™ Phosphatase Inhibitor Tablets, and Sigma-Aldrich™ Phosphatase Inhibitor Cocktail 2 and 3 liquid formulations. Reactions were incubated for 1 hr at 37ºC, and fluorescence was determined at the appropriate emission wavelength. The percent inhibition is shown for each phosphatase inhibitor formulation.
Product selection guides | Products |
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Mammalian cell protein extraction | T-PER Tissue Protein Extraction Reagent N-PER Neuronal Protein Extraction Reagent M-PER Mammalian Protein Extraction Reagent RIPA Lysis Buffer IP Lysis Buffer |
Insect protein extraction | I-PER Insect Cell Protein Extraction Reagent |
Bacterial cell lysis | B-PER Complete Bacterial Protein Extraction Reagent B-PER Bacterial Protein Extraction Reagent B-PER (PBS) Bacterial Protein Extraction Reagent B-PER II (2X) Bacterial Protein Extraction Reagent |
Plant protein extraction | Plant Total Protein Extraction Kit |
Yeast protein extraction | Y-PER Yeast Protein Extraction Reagent |
Subcellular fractionation | NE-PER Nuclear and Cytoplasmic Extraction Reagents Subcellular protein fractionation kits (tissue or cultured cells) Syn-PER Synaptic Protein Isolation Kit |
Membrane protein extraction and isolation | GPCR Extraction and Stabilization Reagent Mem-PER Plus Membrane Protein Extraction Kit Cell Surface Protein Isolation Kit |
Organelle isolation | Lysosome Enrichment Kit for Tissues and Cultured Cells Organelle isolation using magnetic beads Mitochondrial isolation kits (tissue or cultured cells) |
Neuronal cell protein extraction | N-PER Neuronal Protein Extraction Reagent Syn-PER Synaptic Protein Isolation Kit |
Product selection guides | Products |
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Detergents for protein solubilization | Surfact-Amps detergents n-Dodecyl-beta-maltoside detergent CHAPS detergent (3-((3-cholamidopropyl) dimethylammonio)-1-propanesulfonate) Sodium dodecyl sulfate (SDS) Octylthioglucoside (OTG) detergent Octyl-beta-glucoside detergent |
Product selection guides | Products | |
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Protease and phosphatase inhibitors | Protease liquid cocktails, tablets, and capsules | Halt Protease Inhibitor Cocktail Halt Protease Inhibitor Cocktail, EDTA free Pierce Protease Inhibitor tablet Pierce Protease Inhibitor mini tablet Pierce Protease Inhibitor tablets, EDTA-free Pierce Protease Inhibitor mini tablets, EDTA-free Pierce Protease Inhibitor XL Capsules, EDTA-free |
Phosphatase liquid cocktail and tablets | Halt Phosphatase Inhibitor Cocktail Pierce Phosphatase Inhibitor Tablet | |
Combined protease and phosphatase liquid cocktails and tablets | Halt Protease and Phosphatase Inhibitor Cocktail Halt Protease and Phosphatase Inhibitor Cocktail, EDTA free Pierce Protease and Phosphatase Inhibitor Mini Tablet Pierce Protease and Phosphatase Inhibitor Mini Tablet, EDTA Free |
Various methods are used to enrich or purify a protein of interest from other proteins and components in a crude cell lysate or other sample. Ion exchange and affinity chromatography are two commonly used strategies for partial or 1-step purification.
Also known as affinity chromatography, this purification method is enabled by the specific binding properties of a protein to an immobilized ligand. Since the protein of interest is tightly bound, contaminants can be removed through wash steps, and the bound protein can be stripped (eluted) from the support in a highly purified form. Affinity purification is desirable because it often produces higher protein yields and requires less steps than other purification methods. It is the method of choice for purifying recombinant or biotinylated proteins and antibodies.
Discover more about protein purification
See protein isolation and purification learning resource
Also known as ion exchange chromatography, this purification method enables the separation of proteins based on the protein charge at a particular pH. Since multiple proteins may have similar charges, IEX chromatography generally enables only partial purification of a protein of interest when used early in a multistep purification process. However, IEX resins can also be used during a final polishing step to remove specific contaminants that persist after other purification steps. Typically, proteins bind to the IEX column at low ionic strength and elute differentially by increasing salt concentration or changing pH in a gradient. A cation exchange resin binds to positively charged proteins; an anion exchange resin binds to negatively charged proteins. Ion exchange resins are classified as “weak” or “strong”, which refers to the extent that the ionization state of the functional groups varies with pH.
Comparison of DYKDDDDK-tagged SUMO protein yield and background using Pierce anti-DYKDDDDK resin and other products C- and N-terminal DYKDDDDK-tagged SUMO proteins were expressed in E. coli and purified using Pierce Anti-DYKDDDDK Magnetic Agarose, Sigma-Aldrich® Anti-FLAG™ M2 Magnetic Beads, and MBL Anti-DDDDK-tag mAb-Magnetic Agarose. Tagged protein was competitively eluted with Pierce 3x DYKDDDDK Peptide, and the results were analyzed by SDS-PAGE(A) and densitometry using the Invitrogen iBright Imaging System(B). Comparison between the starting lysate and elution fractions shows effective immunoprecipitation and elution of DYKDDDDK-tagged protein, with minimal background from the Pierce magnetic agarose compared to the other suppliers’ products.
Types of affinity purification | Description |
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Protein immobilization (Activated supports for custom immobilization) | Uses activated supports and accessories for the immobilization of proteins, antibodies, and other molecules. These resins or magnetic beads are available separately or in convenient kits. Different reactive chemistries are available to optimize immobilization based on the ligand properties. See covalent immobilization of affinity ligands learning resource |
Antibody purification | Proteins A, G, A/G, and L have unique properties, which make each one suitable for different types of antibody targets (e.g., antibody subclass or animal species). These ligands enable purification of general immunoglobulins from a crude sample. Depending on the sample source, an antigen-specific antibody may account for only a small portion of the total immunoglobulin in the sample. For example, generally only 2–5% of total IgG in mouse serum is specific for the antigen used to immunize the animal. See antibody purification learning resource |
Recombinant protein purification (Fusion protein purification) | Uses resins for the purification of recombinant proteins from cultures such as E. coli or Pichia. These resins are available in multiple formats to accommodate a variety of needs, from high-throughput screening to batch and pilot-scale purification. Superflow resins have undergone extensive chemical characterization. We have ligands targeting a variety of fusion tags, including 6xHis, GST, anti-DYKDDDDK (anti-FLAG™), c-Myc, and HA. See fusion protein learning resource |
Purification using magnetic Agarose | Magnetic agarose beads consist of highly crosslinked agarose encapsulating a ferrimagnetic core. The beads are 10–40 uM in size and have higher binding capacity than traditional magnetic beads. We offer a variety of ligands for immunoprecipitation (IP), co-immunoprecipitation (co-IP), pull-down, and other high throughput affinity screening applications, utilizing immobilized Protein A/G, Ni-NTA, Glutathione, and Anti-DYKDDDDK. The beads are removed from the solution manually using a magnetic stand or by automation using an instrument such as the Thermo Scientific KingFisher Flex Magnetic Particle Processor. Automated instruments are especially useful for higher throughput purification and screening of purification conditions. See scientific poster: High capacity magnetic supports for automated antibody and epitope-tagged protein purifications |
Biotin affinity purification | Uses resins for the purification of biotinylated or desthiobiotinylated proteins, peptides, and other molecules. These resins are available in multiple pack sizes, as well as in spin columns, kits, FPLC cartridges, and coated plates. Different biotin-binding ligands are available based on elution conditions or level of purity. See avidin-biotin interaction learning resource |
Immunoprecipitation (IP) | Small-scale affinity purification of antigens using a specific antibody that is immobilized to a solid support such as magnetic beads or agarose resin. IP is one of the most widely used methods for isolation of proteins and other biomolecules from cell or tissue lysates for the purpose of subsequent detection by western blotting and other assay techniques. See immunoprecipitation learning resource See IP support |
Co-immunoprecipitation (co-IP) and pull-down | Similar to IP, except the target antigen precipitated by the antibody is used to co-precipitate its binding partner(s) or associated protein complex from the lysate and pull-downs. This method is used when antibodies to specific proteins are not available. These “bait” proteins are tagged with an epitope to which a high-affinity antibody is available and ectopically expressed in the cell of interest. See co-immunoprecipitation learning resource See pull-down assays learning resource See co-IP and pull-down support |
ChIP, RIP, and protein-nucleic acid pull-down | Chromatin immunoprecipitation (ChIP) assays are performed to identify regions of the genome with which DNA-binding proteins, such as transcription factors and histones, associate. In ChIP assays, proteins bound to DNA are temporarily crosslinked and the DNA is sheared prior to cell lysis. The target proteins are immunoprecipitated along with the crosslinked nucleotide sequences, and the DNA is then removed and identified by PCR, sequenced, applied to microarrays, or analyzed in some other way. RNA immunoprecipitation (RIP) uses an approach similar to ChIP, except that RNA-binding proteins are immunoprecipitated instead of DNA-binding proteins. Immunoprecipitated RNAs can then be identified by RT-PCR and cDNA sequencing. See ChIP learning resource See protein-nucleic acid interaction support |
Endotoxin detection and removal kits | Endotoxin is a type of pyrogen and is a component of the exterior cell wall of Gram-negative bacteria. It is an unwanted and toxic by-product of recombinant proteins purified from E coli. Therefore, its removal from protein samples is an important step for downstream applications. See app note: Eliminate endotoxins from protein and antibody samples |
Application | Purity level | Ligand and/or chemistry | Base bead type | Packaging options |
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Ion exchange purification | Medium to high (application-specific) | Strong anion exchange | POROS | Loose resin |
Strong cation exchange | ||||
Antibody purification | High | Protein A, protein G, protein A/G | Agarose, magnetic beads, magnetic agarose, POROS | Loose resins or beads, spin columns and kits, chromatography cartridges, 96-well spin plates |
Protein L | Agarose, magnetic beads | |||
Melon Gel | Agarose | |||
Fusion protein purification | High | Ni-NTA, Ni-IDA, high capacity EDTA compatible Ni-IMAC, cobalt, glutathione | Agarose, Superflow, magnetic beads, magnetic agarose | Loose resins or beads, spin columns and kits, chromatography cartridges, 96-well spin plates |
Anti-c-Myc, anti-HA, anti-FLAG | Agarose, UltraLink magnetic beads | Loose resins or beads, kits | ||
Biotin affinity purification | High | Avidin, streptavidin, NeutrAvidin, monomeric avidin | Agarose, magnetic beads | Loose resins, spin columns and kits, chromatography cartridges, 96-well spin plates |
Protein immobilization | High | Amine-reactive, sulfhydryl-reactive, carbonyl-reactive, carboxyl-reactive | Agarose | Loose resins or dry powder |
Epoxy, tosyl-activated, carboxylic acid, amine | Magnetic beads | Loose beads |
Scale | High-throughput screening | High-throughput batch | Batch | Pilot | Process |
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Description | Small scale, automation compatible | Lab or bench scale | Lab or bench scale | Scale-up desired | Production scale |
Yield | Microgram | Milligram | Milligram | Gram | Kilogram |
Format | Magnetic particle processor | Magnetic particle processor, 96-well spin plate (agarose) | Gravity flow, spin column (agarose), fast protein liquid chromatography (FPLC) at low flow rates | FPLC at medium flow rates | FPLC at high flow rates |
Application | High-throughput screening, interaction studies (IP, co-IP, pull-down), mutational analysis | High-throughput screening, interaction studies (IP, co-IP, pull-down), mutational analysis requiring mg scale | Functional assays, structural analysis | Structural analysis, intermediate-scale production | Bulk production |
Recommended resin type | Magnetic bead (1-2.8 µm) | ||||
Magnetic agarose (10-40 µm) | |||||
Agarose (45-165 µm) | |||||
Superflow (45-165 µm) | |||||
UltraLink resin (50-80 µm) | |||||
POROS resin (50 µm) |
The process of protein purification varies depending on the downstream analyses to be performed. Certain steps may be repeated or omitted to achieve the desired result.
Product selection guides | Products |
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Protein immobilization | Pierce NHS-Activated Agarose AminoLink Coupling Resin AminoLink Plus Coupling Resin SulfoLink Coupling Resin GlycoLink Coupling Resin CarboxyLink Coupling |
Additional activated supports and accessories | Pierce NHS-Activated Agarose Spin Columns AminoLink Plus Immobilization Kit AminoLink Plus Micro Immobilization Kit AminoLink Immobilization Kit AminoLink Reductant Pierce CDI-activated Agarose Resin SulfoLink Immobilization Kit for Peptides UltraLink Iodoacetyl Micro Peptide Coupling Kit GlycoLink Immobilization Kit GlycoLink Micro Immobilization Kit CarboxyLink Immobilization Kit |
Product selection guides | Products | |||
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Antibody purification | Protein A | Magnetic beads | Pierce Protein A Magnetic Beads Dynabeads Protein A Magnetic Beads | |
Resins | Pierce Protein A Agarose Pierce Protein A Plus Agarose Pierce Recombinant Protein A Agarose POROS MabCapture A Select | |||
Spin columns and kits | NAb Protein A Plus Spin Columns NAb Protein A Plus Spin Kits | |||
Buffers | Pierce Protein A IgG Binding Buffer | |||
Protein G | Magnetic beads | Pierce Protein G Magnetic Beads Dynabeads Protein G Magnetic Beads | ||
Resins | Pierce Protein G Agarose Pierce Protein G Plus Agarose POROS MabCapture G Select | |||
Spin columns and kits | NAb Protein G Spin Columns NAb Protein G Spin Kits Pierce Recombinant Protein G | |||
Buffer | Pierce Protein G IgG Binding Buffer | |||
Protein A/G | Magnetic beads | Pierce Protein A/G Magnetic Agarose Beads Pierce Protein A/G Magnetic Beads | ||
Resins | Pierce Protein A/G Agarose Pierce Protein A/G Plus Agarose POROS MabCapture A/G Select | |||
Spin columns and kits | NAb Protein A/G Spin Columns NAb Protein A/G Spin Kit Pierce Recombinant Protein A/G | |||
Buffer | Pierce Protein A/G IgG Binding Buffer | |||
Protein L | Magnetic beads | Pierce Protein L Magnetic Beads | ||
Resins | Pierce Protein L Agarose Pierce Protein L Plus Agarose | |||
Spin columns and kits | NAb Protein L Spin Columns NAb Protein L Spin Kit Pierce Recombinant Protein L | |||
Melon Gel | Melon Gel IgG Spin Purification Kit Melon Gel IgG Purification Kit Melon Gel Monoclonal IgG Purification Kit Monoclonal IgG Purification Kit Melon Gel Spin Plate Kit for IgG Screening Melon Gel Chromatography Cartridges (1 mL or 5 mL) Melon Gel Regenerant Ascites Conditioning Reagent for Melon Gel Melon Gel Purification Buffer | |||
Buffers and related products | Pierce IgG Elution Buffer Pierce Gentle Ag/Ab Binding Buffer, pH 8.0 Pierce Gentle Ag/Ab Elution Buffer, pH 6.6 Pierce Gentle Ag/Ab Binding and Elution Buffer Kit Pierce Saturated Ammonium Sulfate Solution Pierce Thiophilic Adsorption Kit Pierce Mannan Binding Protein Agarose Pierce IgM Purification Kit Elution Buffer for Pierce IgM Purification Kit MBP Column Preparation Buffer for Pierce IgM Purification Kit Pierce Jacalin Agarose |
Product selection guides | Products |
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His-tagged (6xHis) protein purification | Pierce High-Capacity Ni-IMAC MagBeads, EDTA Compatible Pierce High-Capacity Ni-IMAC Resin, EDTA Compatible Pierce Ni-NTA Magnetic Beads Pierce Ni-NTA Magnetic Agarose Beads HisPur Ni-NTA Agarose Resin HisPur Ni-NTA Superflow Resin HisPur Cobalt Agarose Resin HisPur Cobalt Superflow Resin |
GST-tagged protein purification | Pierce Glutathione Magnetic Agarose Beads Pierce Glutathione Agarose Resin Pierce Glutathione Superflow Resin |
Other epitope-tagged protein purification | Pierce Anti-DYKD4K (FLAG) Magnetic Agarose Pierce Anti-DYKD4K (FLAG) Affinity Resin UltraLink Pierce Anti-c-Myc Magnetic Beads Pierce Anti-c-Myc Agarose (Superflow 6) Pierce Anti-HA Magnetic Beads Pierce Anti-HA Agarose |
Purification using magnetic agarose | Pierce Ni-NTA Magnetic Agarose Beads Pierce Glutathione Magnetic Agarose Beads Pierce Anti-DYKD4K (FLAG) Magnetic Agarose Beads |
Product selection guides | Products | |
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Biotin affinity purification | Magnetic beads | Pierce Streptavidin Magnetic Beads |
Resins | Pierce Avidin Agarose Pierce Streptavidin Agarose Pierce Streptavidin Agarose HC Pierce NeutrAvidin Agarose Pierce NeutrAvidin Agarose HC Pierce Monomeric Avidin Agarose | |
Related products for biotin-binding applications | Invitrogen CaptAvidin Agarose (Sedimented Bead Suspension) Pierce Streptavidin Agarose Columns Pierce High Capacity Streptavidin Chromatography Cartridge Pierce Monomeric Avidin Agarose Kit Pierce Biotin Pierce Iminobiotin Agarose |
Product selection guides | Products | ||||
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Immunoprecipitation (IP) | Antibody-binding IP products | Protein A, G, A/G | Magnetic beads |
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Kits |
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Secondary antibodies (anti-mouse, anti-rabbit) | Magnetic beads |
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Surface-activated beads (epoxy) | Kits |
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Biotin-binding IP products |
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Recombinant protein (fusion tag) IP products |
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Co-immunoprecipitation (co-IP) and pull-down | Spin column | ||||
Magnetic beads | |||||
ChIP, RIP, and Protein-Nucleic Acid Pull-Down | ChIP |
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RIP |
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Product selection guides | Products | |
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Ion exchange purification | Ion exchange spin columns | Pierce (SCX) Spin Columns Pierce (SAX) Spin Columns Ion Exchange Purification |
Ion exchange resins | POROS XS Resin POROS XQ Resin POROS 50 HQ Resin |
Product selection guides | Products |
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Endotoxin detection | Pierce Chromogenic Endotoxin Quant Kit Pierce LAL Chromogenic Endotoxin Quantitation Kit |
Endotoxin removal | Loose resin Spin columns (0.25 mL, 0.5 mL, or 1 mL) |
Many detergents and salts used in protein extraction formulations may have adverse effects on protein function or stability, or may interfere with downstream analysis. Therefore, it may be necessary to remove or reduce these contaminants following cell lysis or subsequent sample processing such as protein purification.
Download Protein Clean-Up Technical Handbook
Discover more about protein dialysis, desalting, and concentration
Product category | Products |
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Protein dialysis | Slide-A-Lyzer products |
Protein desalting | Zeba products |
Protein concentration | Protein concentrators |
Detergent removal products | HiPPR Detergent Removal 96-well Spin Plates |
Abundant protein depletion | High Select Top 14 and Top 2 reagents |
Small molecule removal | Biotin, dye, crosslinkers, and reducing agent removal products |
Dialysis is a classic clean-up technique that removes small molecules and unwanted compounds by selective diffusion through a semi-permeable membrane. A sample and a buffer solution are placed on opposite sides of the membrane. Proteins that are larger than the membrane pores are retained on the sample side of the membrane, but smaller molecules (contaminants) diffuse freely through the membrane until an equilibrium concentration is achieved. Through this technique, the concentration of small contaminants in the sample can be decreased to acceptable levels.
Discover more about protein dialysis
See protein dialysis learning resource
MWCO* membrane | 10-100 µL Pierce 96-well Microanalysis Plate | 10-2,000 µL Slide-A-Lyzer MINI Dialysis Device | 0.1-70 mL Slide-A-Lyzer G2 Dialysis Cassette | 0.1-30mL Slide-A-Lyzer Dialysis Cassette | 150-250 mL Slide-A-Lyzer Dialysis Flask | 15-100 mL SnakeSkin Dialysis Tubing |
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2K | N/A | ✔ | ✔ | ✔ | ✔ See product | N/A |
3.5K | ✔ See product | ✔ | ✔ | ✔ | ✔ See product | ✔ |
7K | N/A | ✔ | ✔ | ✔ | N/A | ✔ |
10K | ✔ See product | ✔ | ✔ | ✔ | ✔ See product | ✔ |
20K | N/A | ✔ | ✔ | ✔ | ✔ See product | N/A |
See full comparison of these products › |
*MWCO: Molecular weight cut off.
Sample retention by the 2K, 3.5K, 7K, 10K, and 20K MWCO Thermo Scientific Slide-A-Lyzer cassette membrane. Individual proteins or vitamin B12 (1 mg/mL) in either saline or 0.2 M carbonate-bicarbonate buffer, pH 9.4 were dialyzed overnight (17 hours) at 4°C. The amount of retentate was estimated using either the Pierce BCA Protein Assay Kit or absorption at 360 nm (for vitamin B12).
The rate of removal of NaCl using various dialysis products. NaCl removal from samples was determined by measuring the conductivity of the retentate at the indicated times. (A)Slide-A-Lyzer MINI Dialysis Device (10K MWCO, 2 mL) versus conventional dialysis. Bovine serum albumin (BSA) samples (2 mL, 0.25 mg/mL in 1 M NaCl) were dialyzed against 45 mL of water in 50 mL disposable conical tubes on an orbital shaker (300 rpm) at room temperature. The water was changed once after 2 hours. Results are the average of two samples. For conventional dialysis, the samples were dialyzed against 2 L of water in a beaker with stirring. Greater than 95% of NaCl was removed within 4 hours. (B) Samples of 0.1 mL (0.4 mg/mL cytochrome C containing 1 M NaCl) were dialyzed in the Pierce 96-well Microdialysis Plate against 1.8 mL of water at RT with gentle shaking. The buffer was changed at 1-, 2-, and 3-hour intervals over a 4-hour period. Removal of NaCl was >83% after 2 hours and >99% after 4 hours. (C) Proteins in 200 mL samples containing 1 M NaCl were dialyzed at room temperature using Slide-A-Lyzer Dialysis Flasks with 2K, 3.5K, 10K, and 20K MWCOs. The dialysis buffer (4 L) was changed after 2 and 5 hours (triangles; also at 41 hours for the 2K condition). Greater than 95% of NaCl was removed within 8 to 18 hours (41 hours for the 2K condition).
Size exclusion chromatography, also described as gel filtration, can be used for removal of salts from samples. Using this technique, a resin is selected with pores large enough for salts to penetrate, but small for the protein of interest to enter. This causes contaminants to slow down their rate of migration. The larger and faster proteins separate from the slower and smaller molecules during gravity flow or centrifugation.
Discover more about protein desalting
See protein desalting learning resource
Type | Spin columns | Spin plates | Chromatography columns | ||||||
Format | Micro | 0.5 mL | 2 mL | 5 mL | 10 mL | 96-well | 1 mL | 5 mL | |
Resin bed | 75 µL | 0.5 mL | 2 mL | 5 mL | 10 mL | 550 µL | 1 mL | 5 mL | |
Sample volume (7K MWCO) | 2-12 µL | 30-130 µL | 200-700 µL | 500-2,000 µL | 700-4,000 µL | 20-100 µL | 50-250 µL | 100-1,500 µL | |
Sample volume (40K MWCO) | 5-14 µL | 70-200 µL | 200-900 µL | 300-2,000 µL | 1,000-4,000 µL | 20-100 µL | N/A | N/A | |
See full comparison of these products › |
Zeba Spin Desalting Columns result in a high protein recovery while providing minimal sample dilution over a wider range of sample concentrations and volumes compared to alternative products.Zeba Spin Desalting Columns, 10 mL (7K MWCO) and GE PD-10 Columns were used to desalt 1.5, 2.5, and 3.5 mL BSA samples at a concentration of 0.04, 0.2, and 1 mg/mL. Desalting was performed according to the manufacturers’ recommended protocols; both the spin and gravity protocols were used for the GE PD-10. Protein recovery was analyzed by SDS-PAGE. For each electrophoresis gel, an aliquot of starting sample equal to 1 μg of BSA was loaded in lane 1 as the loading control; all other desalted samples were loaded in the gel at the same volume as the loading control. Differences in intensity between lanes are a combination of protein recovery and sample dilution caused by desalting. The largest differences in recovery and concentration are noted in the highlighted area.
Protein concentration is similar to dialysis and uses a semi-permeable membrane to separate proteins from low molecular weight compounds. Unlike dialysis, which relies on passive diffusion, concentration is achieved by forcing solution through membrane using centrifugation. During centrifugation, both buffer and low molecular weight solutes are forced through the membrane where they are collected on the other side (filtrate). Macromolecules (proteins) remain on the sample side of the membrane, where they become concentrated to a smaller volume (retentate), as the reagent is forced across the membrane to the other side.
Discover more about protein concentrators
See protein dialysis, desalting, and buffer exchange learning resource
Volume range | 0.1–0.5 mL | 2–6 mL | 5–20 mL | 20–100 mL |
MWCOs available | 3K, 10K, 30K, 100K | 3K, 10K, 30K, 100K | 3K, 10K, 30K, 100K | 5K, 10K, 30K, 100K |
Processing time* | 3-15 min | 15-90 min | 15-60 min | 15-90 min |
Retentate volume range* | 9-67 µL | 51-174 µL | 121-777 µL | 1.9-3.5 mL |
Protein recovery range* | 95-100% | 94-100% | 94-100% | 92-98% |
See full comparison of these products › |
*Four different protein solutions were used for each MWCO
Comparison of protein recovery between Pierce Protein Concentrators (using 3K, 5K, 10K, 30K, or 100K MWCO) and other vendors for 0.5 mL, 6 mL, 20 mL, and 100 mL concentrators. Samples of different protein solutions were centrifuged in Pierce Protein Concentrators and other suppliers’ concentrators according to manufacturers’ instructions: 0.5 mL (15,000 x g), 5 mL (4,000 x g), 20 mL (4,700 x g), and 100 mL (1,200 x g). Samples were centrifuged until a greater than 15- to 30-fold decrease in sample volume was achieved; protein concentration was measured by either Pierce BCA Protein Assay Kit (0.5 mL concentrators only) or absorbance at A280.
Quantifying total protein concentration is an important step in workflows involving isolation, separation, and analysis of proteins by biochemical methods. Assay methods may use colorimetric or fluorescent detection with fluorometers, spectrophotometers, or plate readers. Every protein assay has limitations depending on the application and the specific protein sample analyzed. The most useful features to consider when choosing a protein assay are sensitivity (lower detection limit), compatibility with common substances in samples (e.g., detergents, reducing agents, chaotropic agents, inhibitors, salts, and buffers), standard curve linearity, and protein-to-protein variation.
Colorimetric signals can be detected using a microplate reader or spectrophotometer. The most popular colorimetric protein assays are:
Fluorescence-based protein quantitation is an alternative to colorimetric methods. Fluorescence detection methods offer excellent sensitivity, requiring less protein sample thereby leaving more sample available for your experiment. Additionally, read time is not a critical factor, so the assays can be readily adapted for automated high-throughput applications. The fluorescence signal can be detected using a fluorometer or microplate reader.
Discover more about protein assays and analysis
See protein quantitation and detection learning resource
Learn more about methods and technologies for identifying and measuring proteins
Product highlight | Category | Description |
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Pierce BCA Protein Assay Kit | Colorimetric | Two-component, high-precision, detergent-compatible protein assay. Compared to most dye-binding methods, the BCA assay is affected much less by protein compositional differences, providing greater concentration accuracy. |
Pierce Rapid Gold BCA Protein Assay Kit | Colorimetric | This kit maintains the key characteristics of the traditional BCA assay but allows a fast time and room temperature incubation equal to dye-binding methods. |
Quant-iT Protein Assay | Fluorescence | The assay is highly selective for protein and exhibits very little protein-to-protein variation. The assay is performed at room temperature, and the signal is stable for 3 hours. |
Qubit 4 Fluorometer | Fluorometer | The Qubit 4 Fluorometer is the latest version of the popular Qubit fluorometer designed to accurately measure protein quantity. |
Protein detection method attributes | ELISA | Western blotting | Mass spec |
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Advantages |
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Sensitivity | <5-10 pg/mL | low femtogram to high attogram* | attomolar range (1018) |
Lysis buffer compatibility |
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| Detergents and high salts must be removed prior to analysis |
Typical total protein required | 0.1 µg/mL - 1 µ g/mL | 1 µg- 50 µg | <1 µg |
Equipment required | Plate reader | X-ray film or CCD imaging equipment | Mass spectrometer |
*With high sensitivity HRP substrates, such as SuperSignal West Atto Ultimate Sensitivity Substrate
Discover more about western blotting, ELISA,mass spec
Researchers have many methodology choices when it comes to producing recombinant proteins for early-stage discovery research through large-scale production of biotherapeutic drugs, vaccine development, and structural studies. It is imperative to use the right protein expression system for the target protein and application of interest. We offer a wide selection of superior mammalian, insect, bacterial, and yeast protein expression systems to suit your research needs.
Our Gibco Expi Transient Expression Systems, available in mammalian (CHO-S, 293F cells) and insect (Sf9 cells) expression formats, are completely optimized systems that enable rapid, high-yield production of proteins.
Recombinant protein titers in ExpiCHO and other transient CHO expression systems. Expression levels of human IgG, rabbit IgG and erythropoietin in ExpiCHO and other transient CHO expression systems are shown. ExpiCHO titers range from 25–160 times those of other transient CHO expression systems.
Discover more about Transient Protein Expression Platforms
Visit Protein Expression Learning Center
Visit Protein Expression Basics Technical Reference Library
Product highlight | Description |
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ExpiCHO Expression System | The Gibco ExpiCHO Expression System is a completely optimized system consisting of ExpiCHO-S(TM) cells that have been adapted to high-density, serum-free suspension culture in ExpiCHO(TM) Expression Medium, along with specially designed transfection reagents and enhancers, that provide the highest yields possible in a transient system (up to 3g/L). That means you can start your research work in CHO cells and stay in CHO cells through discovery. |
Expi293 Expression System | The Gibco Expi293 Expression System is a completely optimized system consisting of Expi293F(TM) cells that have been adapted to high-density, serum-free suspension culture in Expi293(TM) Expression Medium, along with specially designed transfection reagents and enhancers, that enable recombinant 293-derived recombinant protein expression in just five to seven days, with a 2-10-fold increase in protein yields compared to previous generation transient expression systems |
ExpiSf Expression System | The ExpiSf Expression System is the first-ever chemically defined baculovirus-insect cell protein expression system, delivering superior yields (3x more protein) and consistent performance run after run using a fast, streamlined workflow. |
Make sure to check for the following things when getting started with an Expi transient expression system:
Product selection guides | Products |
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Transient protein expression systems | Expi293 ExpiCHO ExpiSf Expression Systems |
Mammalian protein expression | Expi293 ExpiCHO Expression Systems |
Insect protein expression | ExpiSf Expression System |
Bacterial protein expression | Champion pET Expression System |
Yeast protein expression | PichiaPink Yeast Expression System |
Structural Biology and Membrane Protein Expression | Expi293 GnTI- Expi293 Inducible Expi293 Inducible GnTI- Expression Systems Expi293 Met(-) Protein Labeling Kit |
Protein expression selection guide | Protein Expression Systems Selection Guide |
Protein expression services | Gene-to-Protein Baculovirus Expression Mammalian Expression services |
Protein extraction techniques vary depending on the source of the starting material, the location of the protein of interest within the cell, and the downstream application. Other important considerations include the preservation of protein activity and function as well as the reduction of background effects.
Historically, mechanical disruption has been used to lyse cells and tissues; our gentle, detergent-based solutions have been developed to efficiently lyse cells and enable the separation of subcellular structures without requiring physical disruption, providing high yields of active proteins.
Protein extraction product features:
Protein extraction efficiency from major cellular compartments using M-PER Mammalian Protein Extraction Reagent. Lysates from established cell lines and primary cultures were prepared using M-PER reagent and extraction efficiency from the various cellular compartments evaluated. For each target protein, 10 µg of lysate was loaded for and electrophoresed by SDS PAGE, transferred to nitrocellose membrane and detected by western blot using SuperSignal West Pico PLUS Chemiluminescent Substrate.
Protein yield from various cell types using M-PER Mammalian Protein Extraction Reagent. Cells were harvested at 85% confluency, washed twice and collected in ice-cold PBS and counted. For each cell type, 1 x 106 cells were pelleted by centrifugation at 2,000 x g for 5 minutes and lysed in 1 mL M-PER Reagent for 5 minutes. The cell lysates were clarified by centrifugation at 14,000 x g for 10 minutes and the supernatant was collected and the protein concentration (µg/million cells) was determined using the Pierce BCA Protein Assay.
Improved protein yield using the Thermo Scientific Mem-PER Plus Membrane Protein Extraction Kit. Membrane proteins were isolated from mouse liver tissue and HeLa cells using Thermo Scientific Mem-PER Plus Membrane Protein Extraction Kit and three other commercial extraction kits. Protein yields (μg) for membrane, cytosolic, and total fractions were determined using the Thermo Scientific Pierce BCA Protein Assay Kit.
Tissue cell lysis protein yield with T-PER Tissue Protein Extraction Reagent. Duplicate tissue samples were weighed, resuspended in 1:10 to 1:20 w/v T-PER Reagent and disrupted in a chilled Dounce or benchtop tissue homogenizer. The resulting lysates were centrifuged at 10,000 x g for 5 minutes and the supernatant was collected. The protein concentration of each lysate was determined using the Pierce BCA Protein Assay to determine protein yield per milligram of starting tissue.
Protein yield comparison of two bacterial cell lysis reagents.E. coli ER2566/pLATE51-Klenow, ER2566/pGST-CC-StpB, and ER2566/pGS-Syk cell pellets (0.5 g), were resuspended in 2.5 mL aliquots of Thermo Scientific B-PER Complete Bacterial Protein Extraction Reagent or EMD Chemicals BugBuster Master Mix with gentle vortexing for 15 minutes at room temperature. Insoluble cell debris was removed by centrifugation at 16,000 x g for 20 minutes at 4°C. Protein yields (concentrations) for soluble fractions were determined using the Pierce BCA Protein Assay Kit.
Sample type | Goal | Product highlight | |
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Primary or cultured mammalian cells or tissues | Total protein extraction | M-PER reagent T-PER reagent N-PER reagent | RIPA Lysis and Extraction Buffer IP Lysis Buffer |
Cultured mammalian cells or tissues | Subcellular fractionation or organelle isolation | NE-PER reagent Subcellular Fractionation Kits Mitochondria Isolation Kits | GPCR Extraction and Stabilization Reagent Cell Surface Protein Isolation Kit Syn-PER Reagent Lysosome Enrichment Kit |
Bacterial cells | Total protein extraction | B-PER reagent | |
Yeast cells | Total protein extraction | Y-PER reagent | |
Insect cells (baculovirus) | Total protein extraction | I-PER reagent | |
Plant tissue (leaf, stem, root, flower) | Total protein extraction | Plan Total Protein Extraction Kit |
Detergents are frequently used in cell lysis reagent formulation and other protein research methods. Thermo Scientific Surfact-Amps Detergent Solutions are highly purified, precisely diluted (10%) formulations that are ideal for applications or assays that are sensitive to contaminants present in unpurified detergents.
Protein detergent product features:
Generic structure of a detergent molecule.
Cell lysis disrupts cell membranes and organelles, resulting in unregulated enzymatic activity that can reduce protein yield and function. To prevent these negative effects, protease and phosphatase inhibitors can be added to the lysis reagents. Numerous compounds have been identified that can inactivate or block the activities of proteases and phosphatases.
Protease and phosphatase inhibitor product features:
Performance comparison between three commercially available protease inhibitor tablets. Pancreatic extract (100 μL; 0.5 μg/μL) was incubated with quenched fluorescent protease-cleavable substrates for trypsin, cysteine, and metalloprotease and cathepsins, in the presence of the reformulated Thermo Scientific Pierce Protease Inhibitor Mini Tablets, Roche™ Complete™ Protease Inhibitor Tablets, and Sigma-Aldrich™ SIGMAFAST™ Protease Inhibitor Cocktail Tablets, with and without EDTA. Reactions were incubated for 1 hr at 37ºC, and fluorescence was determined at the appropriate emission wavelengths. The percent inhibition is shown for each protease inhibitor formulation.
Protein phosphorylation in cell extracts is broadly preserved by Thermo Scientific Phosphatase Inhibitor Mini Tablets.(A) HCT116 cells were serum-starved, then either treated with EGF for 15 min or left as control cells. Cell lysates were prepared in Thermo Scientific Pierce IP Lysis Buffer with Thermo Scientific Protease and Phosphatase Inhibitor Mini Tablets, EDTA-Free, or with no inhibitor. Lysate containing 500 μg of protein was then incubated with 5 μg of phospho-tyrosine antibody overnight at 4ºC. The complex was then incubated with Thermo Scientific Pierce Protein A/G Magnetic Beads for 1 hr at room temperature. Beads were washed, and low-pH elution was performed. The eluates were subjected to western blotting, and the membrane was then probed with EGFR antibody for chemiluminescence detection. (B) The degree of inhibition for protein, alkaline, and acid phosphatase activity was determined in kidney extract (25 μL; 0.5 μg/μL) by incubating extracts with a fluorogenic substrate (MFP or FDP) that measures phosphatase activity upon dephosphorylation in the presence of Pierce Phosphatase Inhibitor Mini Tablets, Roche™ PhosStop™ Phosphatase Inhibitor Tablets, and Sigma-Aldrich™ Phosphatase Inhibitor Cocktail 2 and 3 liquid formulations. Reactions were incubated for 1 hr at 37ºC, and fluorescence was determined at the appropriate emission wavelength. The percent inhibition is shown for each phosphatase inhibitor formulation.
Product selection guides | Products |
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Mammalian cell protein extraction | T-PER Tissue Protein Extraction Reagent N-PER Neuronal Protein Extraction Reagent M-PER Mammalian Protein Extraction Reagent RIPA Lysis Buffer IP Lysis Buffer |
Insect protein extraction | I-PER Insect Cell Protein Extraction Reagent |
Bacterial cell lysis | B-PER Complete Bacterial Protein Extraction Reagent B-PER Bacterial Protein Extraction Reagent B-PER (PBS) Bacterial Protein Extraction Reagent B-PER II (2X) Bacterial Protein Extraction Reagent |
Plant protein extraction | Plant Total Protein Extraction Kit |
Yeast protein extraction | Y-PER Yeast Protein Extraction Reagent |
Subcellular fractionation | NE-PER Nuclear and Cytoplasmic Extraction Reagents Subcellular protein fractionation kits (tissue or cultured cells) Syn-PER Synaptic Protein Isolation Kit |
Membrane protein extraction and isolation | GPCR Extraction and Stabilization Reagent Mem-PER Plus Membrane Protein Extraction Kit Cell Surface Protein Isolation Kit |
Organelle isolation | Lysosome Enrichment Kit for Tissues and Cultured Cells Organelle isolation using magnetic beads Mitochondrial isolation kits (tissue or cultured cells) |
Neuronal cell protein extraction | N-PER Neuronal Protein Extraction Reagent Syn-PER Synaptic Protein Isolation Kit |
Product selection guides | Products |
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Detergents for protein solubilization | Surfact-Amps detergents n-Dodecyl-beta-maltoside detergent CHAPS detergent (3-((3-cholamidopropyl) dimethylammonio)-1-propanesulfonate) Sodium dodecyl sulfate (SDS) Octylthioglucoside (OTG) detergent Octyl-beta-glucoside detergent |
Product selection guides | Products | |
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Protease and phosphatase inhibitors | Protease liquid cocktails, tablets, and capsules | Halt Protease Inhibitor Cocktail Halt Protease Inhibitor Cocktail, EDTA free Pierce Protease Inhibitor tablet Pierce Protease Inhibitor mini tablet Pierce Protease Inhibitor tablets, EDTA-free Pierce Protease Inhibitor mini tablets, EDTA-free Pierce Protease Inhibitor XL Capsules, EDTA-free |
Phosphatase liquid cocktail and tablets | Halt Phosphatase Inhibitor Cocktail Pierce Phosphatase Inhibitor Tablet | |
Combined protease and phosphatase liquid cocktails and tablets | Halt Protease and Phosphatase Inhibitor Cocktail Halt Protease and Phosphatase Inhibitor Cocktail, EDTA free Pierce Protease and Phosphatase Inhibitor Mini Tablet Pierce Protease and Phosphatase Inhibitor Mini Tablet, EDTA Free |
Various methods are used to enrich or purify a protein of interest from other proteins and components in a crude cell lysate or other sample. Ion exchange and affinity chromatography are two commonly used strategies for partial or 1-step purification.
Also known as affinity chromatography, this purification method is enabled by the specific binding properties of a protein to an immobilized ligand. Since the protein of interest is tightly bound, contaminants can be removed through wash steps, and the bound protein can be stripped (eluted) from the support in a highly purified form. Affinity purification is desirable because it often produces higher protein yields and requires less steps than other purification methods. It is the method of choice for purifying recombinant or biotinylated proteins and antibodies.
Discover more about protein purification
See protein isolation and purification learning resource
Also known as ion exchange chromatography, this purification method enables the separation of proteins based on the protein charge at a particular pH. Since multiple proteins may have similar charges, IEX chromatography generally enables only partial purification of a protein of interest when used early in a multistep purification process. However, IEX resins can also be used during a final polishing step to remove specific contaminants that persist after other purification steps. Typically, proteins bind to the IEX column at low ionic strength and elute differentially by increasing salt concentration or changing pH in a gradient. A cation exchange resin binds to positively charged proteins; an anion exchange resin binds to negatively charged proteins. Ion exchange resins are classified as “weak” or “strong”, which refers to the extent that the ionization state of the functional groups varies with pH.
Comparison of DYKDDDDK-tagged SUMO protein yield and background using Pierce anti-DYKDDDDK resin and other products C- and N-terminal DYKDDDDK-tagged SUMO proteins were expressed in E. coli and purified using Pierce Anti-DYKDDDDK Magnetic Agarose, Sigma-Aldrich® Anti-FLAG™ M2 Magnetic Beads, and MBL Anti-DDDDK-tag mAb-Magnetic Agarose. Tagged protein was competitively eluted with Pierce 3x DYKDDDDK Peptide, and the results were analyzed by SDS-PAGE(A) and densitometry using the Invitrogen iBright Imaging System(B). Comparison between the starting lysate and elution fractions shows effective immunoprecipitation and elution of DYKDDDDK-tagged protein, with minimal background from the Pierce magnetic agarose compared to the other suppliers’ products.
Types of affinity purification | Description |
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Protein immobilization (Activated supports for custom immobilization) | Uses activated supports and accessories for the immobilization of proteins, antibodies, and other molecules. These resins or magnetic beads are available separately or in convenient kits. Different reactive chemistries are available to optimize immobilization based on the ligand properties. See covalent immobilization of affinity ligands learning resource |
Antibody purification | Proteins A, G, A/G, and L have unique properties, which make each one suitable for different types of antibody targets (e.g., antibody subclass or animal species). These ligands enable purification of general immunoglobulins from a crude sample. Depending on the sample source, an antigen-specific antibody may account for only a small portion of the total immunoglobulin in the sample. For example, generally only 2–5% of total IgG in mouse serum is specific for the antigen used to immunize the animal. See antibody purification learning resource |
Recombinant protein purification (Fusion protein purification) | Uses resins for the purification of recombinant proteins from cultures such as E. coli or Pichia. These resins are available in multiple formats to accommodate a variety of needs, from high-throughput screening to batch and pilot-scale purification. Superflow resins have undergone extensive chemical characterization. We have ligands targeting a variety of fusion tags, including 6xHis, GST, anti-DYKDDDDK (anti-FLAG™), c-Myc, and HA. See fusion protein learning resource |
Purification using magnetic Agarose | Magnetic agarose beads consist of highly crosslinked agarose encapsulating a ferrimagnetic core. The beads are 10–40 uM in size and have higher binding capacity than traditional magnetic beads. We offer a variety of ligands for immunoprecipitation (IP), co-immunoprecipitation (co-IP), pull-down, and other high throughput affinity screening applications, utilizing immobilized Protein A/G, Ni-NTA, Glutathione, and Anti-DYKDDDDK. The beads are removed from the solution manually using a magnetic stand or by automation using an instrument such as the Thermo Scientific KingFisher Flex Magnetic Particle Processor. Automated instruments are especially useful for higher throughput purification and screening of purification conditions. See scientific poster: High capacity magnetic supports for automated antibody and epitope-tagged protein purifications |
Biotin affinity purification | Uses resins for the purification of biotinylated or desthiobiotinylated proteins, peptides, and other molecules. These resins are available in multiple pack sizes, as well as in spin columns, kits, FPLC cartridges, and coated plates. Different biotin-binding ligands are available based on elution conditions or level of purity. See avidin-biotin interaction learning resource |
Immunoprecipitation (IP) | Small-scale affinity purification of antigens using a specific antibody that is immobilized to a solid support such as magnetic beads or agarose resin. IP is one of the most widely used methods for isolation of proteins and other biomolecules from cell or tissue lysates for the purpose of subsequent detection by western blotting and other assay techniques. See immunoprecipitation learning resource See IP support |
Co-immunoprecipitation (co-IP) and pull-down | Similar to IP, except the target antigen precipitated by the antibody is used to co-precipitate its binding partner(s) or associated protein complex from the lysate and pull-downs. This method is used when antibodies to specific proteins are not available. These “bait” proteins are tagged with an epitope to which a high-affinity antibody is available and ectopically expressed in the cell of interest. See co-immunoprecipitation learning resource See pull-down assays learning resource See co-IP and pull-down support |
ChIP, RIP, and protein-nucleic acid pull-down | Chromatin immunoprecipitation (ChIP) assays are performed to identify regions of the genome with which DNA-binding proteins, such as transcription factors and histones, associate. In ChIP assays, proteins bound to DNA are temporarily crosslinked and the DNA is sheared prior to cell lysis. The target proteins are immunoprecipitated along with the crosslinked nucleotide sequences, and the DNA is then removed and identified by PCR, sequenced, applied to microarrays, or analyzed in some other way. RNA immunoprecipitation (RIP) uses an approach similar to ChIP, except that RNA-binding proteins are immunoprecipitated instead of DNA-binding proteins. Immunoprecipitated RNAs can then be identified by RT-PCR and cDNA sequencing. See ChIP learning resource See protein-nucleic acid interaction support |
Endotoxin detection and removal kits | Endotoxin is a type of pyrogen and is a component of the exterior cell wall of Gram-negative bacteria. It is an unwanted and toxic by-product of recombinant proteins purified from E coli. Therefore, its removal from protein samples is an important step for downstream applications. See app note: Eliminate endotoxins from protein and antibody samples |
Application | Purity level | Ligand and/or chemistry | Base bead type | Packaging options |
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Ion exchange purification | Medium to high (application-specific) | Strong anion exchange | POROS | Loose resin |
Strong cation exchange | ||||
Antibody purification | High | Protein A, protein G, protein A/G | Agarose, magnetic beads, magnetic agarose, POROS | Loose resins or beads, spin columns and kits, chromatography cartridges, 96-well spin plates |
Protein L | Agarose, magnetic beads | |||
Melon Gel | Agarose | |||
Fusion protein purification | High | Ni-NTA, Ni-IDA, high capacity EDTA compatible Ni-IMAC, cobalt, glutathione | Agarose, Superflow, magnetic beads, magnetic agarose | Loose resins or beads, spin columns and kits, chromatography cartridges, 96-well spin plates |
Anti-c-Myc, anti-HA, anti-FLAG | Agarose, UltraLink magnetic beads | Loose resins or beads, kits | ||
Biotin affinity purification | High | Avidin, streptavidin, NeutrAvidin, monomeric avidin | Agarose, magnetic beads | Loose resins, spin columns and kits, chromatography cartridges, 96-well spin plates |
Protein immobilization | High | Amine-reactive, sulfhydryl-reactive, carbonyl-reactive, carboxyl-reactive | Agarose | Loose resins or dry powder |
Epoxy, tosyl-activated, carboxylic acid, amine | Magnetic beads | Loose beads |
Scale | High-throughput screening | High-throughput batch | Batch | Pilot | Process |
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Description | Small scale, automation compatible | Lab or bench scale | Lab or bench scale | Scale-up desired | Production scale |
Yield | Microgram | Milligram | Milligram | Gram | Kilogram |
Format | Magnetic particle processor | Magnetic particle processor, 96-well spin plate (agarose) | Gravity flow, spin column (agarose), fast protein liquid chromatography (FPLC) at low flow rates | FPLC at medium flow rates | FPLC at high flow rates |
Application | High-throughput screening, interaction studies (IP, co-IP, pull-down), mutational analysis | High-throughput screening, interaction studies (IP, co-IP, pull-down), mutational analysis requiring mg scale | Functional assays, structural analysis | Structural analysis, intermediate-scale production | Bulk production |
Recommended resin type | Magnetic bead (1-2.8 µm) | ||||
Magnetic agarose (10-40 µm) | |||||
Agarose (45-165 µm) | |||||
Superflow (45-165 µm) | |||||
UltraLink resin (50-80 µm) | |||||
POROS resin (50 µm) |
The process of protein purification varies depending on the downstream analyses to be performed. Certain steps may be repeated or omitted to achieve the desired result.
Product selection guides | Products |
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Protein immobilization | Pierce NHS-Activated Agarose AminoLink Coupling Resin AminoLink Plus Coupling Resin SulfoLink Coupling Resin GlycoLink Coupling Resin CarboxyLink Coupling |
Additional activated supports and accessories | Pierce NHS-Activated Agarose Spin Columns AminoLink Plus Immobilization Kit AminoLink Plus Micro Immobilization Kit AminoLink Immobilization Kit AminoLink Reductant Pierce CDI-activated Agarose Resin SulfoLink Immobilization Kit for Peptides UltraLink Iodoacetyl Micro Peptide Coupling Kit GlycoLink Immobilization Kit GlycoLink Micro Immobilization Kit CarboxyLink Immobilization Kit |
Product selection guides | Products | |||
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Antibody purification | Protein A | Magnetic beads | Pierce Protein A Magnetic Beads Dynabeads Protein A Magnetic Beads | |
Resins | Pierce Protein A Agarose Pierce Protein A Plus Agarose Pierce Recombinant Protein A Agarose POROS MabCapture A Select | |||
Spin columns and kits | NAb Protein A Plus Spin Columns NAb Protein A Plus Spin Kits | |||
Buffers | Pierce Protein A IgG Binding Buffer | |||
Protein G | Magnetic beads | Pierce Protein G Magnetic Beads Dynabeads Protein G Magnetic Beads | ||
Resins | Pierce Protein G Agarose Pierce Protein G Plus Agarose POROS MabCapture G Select | |||
Spin columns and kits | NAb Protein G Spin Columns NAb Protein G Spin Kits Pierce Recombinant Protein G | |||
Buffer | Pierce Protein G IgG Binding Buffer | |||
Protein A/G | Magnetic beads | Pierce Protein A/G Magnetic Agarose Beads Pierce Protein A/G Magnetic Beads | ||
Resins | Pierce Protein A/G Agarose Pierce Protein A/G Plus Agarose POROS MabCapture A/G Select | |||
Spin columns and kits | NAb Protein A/G Spin Columns NAb Protein A/G Spin Kit Pierce Recombinant Protein A/G | |||
Buffer | Pierce Protein A/G IgG Binding Buffer | |||
Protein L | Magnetic beads | Pierce Protein L Magnetic Beads | ||
Resins | Pierce Protein L Agarose Pierce Protein L Plus Agarose | |||
Spin columns and kits | NAb Protein L Spin Columns NAb Protein L Spin Kit Pierce Recombinant Protein L | |||
Melon Gel | Melon Gel IgG Spin Purification Kit Melon Gel IgG Purification Kit Melon Gel Monoclonal IgG Purification Kit Monoclonal IgG Purification Kit Melon Gel Spin Plate Kit for IgG Screening Melon Gel Chromatography Cartridges (1 mL or 5 mL) Melon Gel Regenerant Ascites Conditioning Reagent for Melon Gel Melon Gel Purification Buffer | |||
Buffers and related products | Pierce IgG Elution Buffer Pierce Gentle Ag/Ab Binding Buffer, pH 8.0 Pierce Gentle Ag/Ab Elution Buffer, pH 6.6 Pierce Gentle Ag/Ab Binding and Elution Buffer Kit Pierce Saturated Ammonium Sulfate Solution Pierce Thiophilic Adsorption Kit Pierce Mannan Binding Protein Agarose Pierce IgM Purification Kit Elution Buffer for Pierce IgM Purification Kit MBP Column Preparation Buffer for Pierce IgM Purification Kit Pierce Jacalin Agarose |
Product selection guides | Products |
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His-tagged (6xHis) protein purification | Pierce High-Capacity Ni-IMAC MagBeads, EDTA Compatible Pierce High-Capacity Ni-IMAC Resin, EDTA Compatible Pierce Ni-NTA Magnetic Beads Pierce Ni-NTA Magnetic Agarose Beads HisPur Ni-NTA Agarose Resin HisPur Ni-NTA Superflow Resin HisPur Cobalt Agarose Resin HisPur Cobalt Superflow Resin |
GST-tagged protein purification | Pierce Glutathione Magnetic Agarose Beads Pierce Glutathione Agarose Resin Pierce Glutathione Superflow Resin |
Other epitope-tagged protein purification | Pierce Anti-DYKD4K (FLAG) Magnetic Agarose Pierce Anti-DYKD4K (FLAG) Affinity Resin UltraLink Pierce Anti-c-Myc Magnetic Beads Pierce Anti-c-Myc Agarose (Superflow 6) Pierce Anti-HA Magnetic Beads Pierce Anti-HA Agarose |
Purification using magnetic agarose | Pierce Ni-NTA Magnetic Agarose Beads Pierce Glutathione Magnetic Agarose Beads Pierce Anti-DYKD4K (FLAG) Magnetic Agarose Beads |
Product selection guides | Products | |
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Biotin affinity purification | Magnetic beads | Pierce Streptavidin Magnetic Beads |
Resins | Pierce Avidin Agarose Pierce Streptavidin Agarose Pierce Streptavidin Agarose HC Pierce NeutrAvidin Agarose Pierce NeutrAvidin Agarose HC Pierce Monomeric Avidin Agarose | |
Related products for biotin-binding applications | Invitrogen CaptAvidin Agarose (Sedimented Bead Suspension) Pierce Streptavidin Agarose Columns Pierce High Capacity Streptavidin Chromatography Cartridge Pierce Monomeric Avidin Agarose Kit Pierce Biotin Pierce Iminobiotin Agarose |
Product selection guides | Products | ||||
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Immunoprecipitation (IP) | Antibody-binding IP products | Protein A, G, A/G | Magnetic beads |
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Kits |
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Secondary antibodies (anti-mouse, anti-rabbit) | Magnetic beads |
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Surface-activated beads (epoxy) | Kits |
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Biotin-binding IP products |
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Recombinant protein (fusion tag) IP products |
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Co-immunoprecipitation (co-IP) and pull-down | Spin column | ||||
Magnetic beads | |||||
ChIP, RIP, and Protein-Nucleic Acid Pull-Down | ChIP |
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RIP |
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Product selection guides | Products | |
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Ion exchange purification | Ion exchange spin columns | Pierce (SCX) Spin Columns Pierce (SAX) Spin Columns Ion Exchange Purification |
Ion exchange resins | POROS XS Resin POROS XQ Resin POROS 50 HQ Resin |
Product selection guides | Products |
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Endotoxin detection | Pierce Chromogenic Endotoxin Quant Kit Pierce LAL Chromogenic Endotoxin Quantitation Kit |
Endotoxin removal | Loose resin Spin columns (0.25 mL, 0.5 mL, or 1 mL) |
Many detergents and salts used in protein extraction formulations may have adverse effects on protein function or stability, or may interfere with downstream analysis. Therefore, it may be necessary to remove or reduce these contaminants following cell lysis or subsequent sample processing such as protein purification.
Download Protein Clean-Up Technical Handbook
Discover more about protein dialysis, desalting, and concentration
Product category | Products |
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Protein dialysis | Slide-A-Lyzer products |
Protein desalting | Zeba products |
Protein concentration | Protein concentrators |
Detergent removal products | HiPPR Detergent Removal 96-well Spin Plates |
Abundant protein depletion | High Select Top 14 and Top 2 reagents |
Small molecule removal | Biotin, dye, crosslinkers, and reducing agent removal products |
Dialysis is a classic clean-up technique that removes small molecules and unwanted compounds by selective diffusion through a semi-permeable membrane. A sample and a buffer solution are placed on opposite sides of the membrane. Proteins that are larger than the membrane pores are retained on the sample side of the membrane, but smaller molecules (contaminants) diffuse freely through the membrane until an equilibrium concentration is achieved. Through this technique, the concentration of small contaminants in the sample can be decreased to acceptable levels.
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MWCO* membrane | 10-100 µL Pierce 96-well Microanalysis Plate | 10-2,000 µL Slide-A-Lyzer MINI Dialysis Device | 0.1-70 mL Slide-A-Lyzer G2 Dialysis Cassette | 0.1-30mL Slide-A-Lyzer Dialysis Cassette | 150-250 mL Slide-A-Lyzer Dialysis Flask | 15-100 mL SnakeSkin Dialysis Tubing |
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2K | N/A | ✔ | ✔ | ✔ | ✔ See product | N/A |
3.5K | ✔ See product | ✔ | ✔ | ✔ | ✔ See product | ✔ |
7K | N/A | ✔ | ✔ | ✔ | N/A | ✔ |
10K | ✔ See product | ✔ | ✔ | ✔ | ✔ See product | ✔ |
20K | N/A | ✔ | ✔ | ✔ | ✔ See product | N/A |
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*MWCO: Molecular weight cut off.
Sample retention by the 2K, 3.5K, 7K, 10K, and 20K MWCO Thermo Scientific Slide-A-Lyzer cassette membrane. Individual proteins or vitamin B12 (1 mg/mL) in either saline or 0.2 M carbonate-bicarbonate buffer, pH 9.4 were dialyzed overnight (17 hours) at 4°C. The amount of retentate was estimated using either the Pierce BCA Protein Assay Kit or absorption at 360 nm (for vitamin B12).
The rate of removal of NaCl using various dialysis products. NaCl removal from samples was determined by measuring the conductivity of the retentate at the indicated times. (A)Slide-A-Lyzer MINI Dialysis Device (10K MWCO, 2 mL) versus conventional dialysis. Bovine serum albumin (BSA) samples (2 mL, 0.25 mg/mL in 1 M NaCl) were dialyzed against 45 mL of water in 50 mL disposable conical tubes on an orbital shaker (300 rpm) at room temperature. The water was changed once after 2 hours. Results are the average of two samples. For conventional dialysis, the samples were dialyzed against 2 L of water in a beaker with stirring. Greater than 95% of NaCl was removed within 4 hours. (B) Samples of 0.1 mL (0.4 mg/mL cytochrome C containing 1 M NaCl) were dialyzed in the Pierce 96-well Microdialysis Plate against 1.8 mL of water at RT with gentle shaking. The buffer was changed at 1-, 2-, and 3-hour intervals over a 4-hour period. Removal of NaCl was >83% after 2 hours and >99% after 4 hours. (C) Proteins in 200 mL samples containing 1 M NaCl were dialyzed at room temperature using Slide-A-Lyzer Dialysis Flasks with 2K, 3.5K, 10K, and 20K MWCOs. The dialysis buffer (4 L) was changed after 2 and 5 hours (triangles; also at 41 hours for the 2K condition). Greater than 95% of NaCl was removed within 8 to 18 hours (41 hours for the 2K condition).
Size exclusion chromatography, also described as gel filtration, can be used for removal of salts from samples. Using this technique, a resin is selected with pores large enough for salts to penetrate, but small for the protein of interest to enter. This causes contaminants to slow down their rate of migration. The larger and faster proteins separate from the slower and smaller molecules during gravity flow or centrifugation.
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Type | Spin columns | Spin plates | Chromatography columns | ||||||
Format | Micro | 0.5 mL | 2 mL | 5 mL | 10 mL | 96-well | 1 mL | 5 mL | |
Resin bed | 75 µL | 0.5 mL | 2 mL | 5 mL | 10 mL | 550 µL | 1 mL | 5 mL | |
Sample volume (7K MWCO) | 2-12 µL | 30-130 µL | 200-700 µL | 500-2,000 µL | 700-4,000 µL | 20-100 µL | 50-250 µL | 100-1,500 µL | |
Sample volume (40K MWCO) | 5-14 µL | 70-200 µL | 200-900 µL | 300-2,000 µL | 1,000-4,000 µL | 20-100 µL | N/A | N/A | |
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Zeba Spin Desalting Columns result in a high protein recovery while providing minimal sample dilution over a wider range of sample concentrations and volumes compared to alternative products.Zeba Spin Desalting Columns, 10 mL (7K MWCO) and GE PD-10 Columns were used to desalt 1.5, 2.5, and 3.5 mL BSA samples at a concentration of 0.04, 0.2, and 1 mg/mL. Desalting was performed according to the manufacturers’ recommended protocols; both the spin and gravity protocols were used for the GE PD-10. Protein recovery was analyzed by SDS-PAGE. For each electrophoresis gel, an aliquot of starting sample equal to 1 μg of BSA was loaded in lane 1 as the loading control; all other desalted samples were loaded in the gel at the same volume as the loading control. Differences in intensity between lanes are a combination of protein recovery and sample dilution caused by desalting. The largest differences in recovery and concentration are noted in the highlighted area.
Protein concentration is similar to dialysis and uses a semi-permeable membrane to separate proteins from low molecular weight compounds. Unlike dialysis, which relies on passive diffusion, concentration is achieved by forcing solution through membrane using centrifugation. During centrifugation, both buffer and low molecular weight solutes are forced through the membrane where they are collected on the other side (filtrate). Macromolecules (proteins) remain on the sample side of the membrane, where they become concentrated to a smaller volume (retentate), as the reagent is forced across the membrane to the other side.
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Volume range | 0.1–0.5 mL | 2–6 mL | 5–20 mL | 20–100 mL |
MWCOs available | 3K, 10K, 30K, 100K | 3K, 10K, 30K, 100K | 3K, 10K, 30K, 100K | 5K, 10K, 30K, 100K |
Processing time* | 3-15 min | 15-90 min | 15-60 min | 15-90 min |
Retentate volume range* | 9-67 µL | 51-174 µL | 121-777 µL | 1.9-3.5 mL |
Protein recovery range* | 95-100% | 94-100% | 94-100% | 92-98% |
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*Four different protein solutions were used for each MWCO
Comparison of protein recovery between Pierce Protein Concentrators (using 3K, 5K, 10K, 30K, or 100K MWCO) and other vendors for 0.5 mL, 6 mL, 20 mL, and 100 mL concentrators. Samples of different protein solutions were centrifuged in Pierce Protein Concentrators and other suppliers’ concentrators according to manufacturers’ instructions: 0.5 mL (15,000 x g), 5 mL (4,000 x g), 20 mL (4,700 x g), and 100 mL (1,200 x g). Samples were centrifuged until a greater than 15- to 30-fold decrease in sample volume was achieved; protein concentration was measured by either Pierce BCA Protein Assay Kit (0.5 mL concentrators only) or absorbance at A280.
Quantifying total protein concentration is an important step in workflows involving isolation, separation, and analysis of proteins by biochemical methods. Assay methods may use colorimetric or fluorescent detection with fluorometers, spectrophotometers, or plate readers. Every protein assay has limitations depending on the application and the specific protein sample analyzed. The most useful features to consider when choosing a protein assay are sensitivity (lower detection limit), compatibility with common substances in samples (e.g., detergents, reducing agents, chaotropic agents, inhibitors, salts, and buffers), standard curve linearity, and protein-to-protein variation.
Colorimetric signals can be detected using a microplate reader or spectrophotometer. The most popular colorimetric protein assays are:
Fluorescence-based protein quantitation is an alternative to colorimetric methods. Fluorescence detection methods offer excellent sensitivity, requiring less protein sample thereby leaving more sample available for your experiment. Additionally, read time is not a critical factor, so the assays can be readily adapted for automated high-throughput applications. The fluorescence signal can be detected using a fluorometer or microplate reader.
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Learn more about methods and technologies for identifying and measuring proteins
Product highlight | Category | Description |
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Pierce BCA Protein Assay Kit | Colorimetric | Two-component, high-precision, detergent-compatible protein assay. Compared to most dye-binding methods, the BCA assay is affected much less by protein compositional differences, providing greater concentration accuracy. |
Pierce Rapid Gold BCA Protein Assay Kit | Colorimetric | This kit maintains the key characteristics of the traditional BCA assay but allows a fast time and room temperature incubation equal to dye-binding methods. |
Quant-iT Protein Assay | Fluorescence | The assay is highly selective for protein and exhibits very little protein-to-protein variation. The assay is performed at room temperature, and the signal is stable for 3 hours. |
Qubit 4 Fluorometer | Fluorometer | The Qubit 4 Fluorometer is the latest version of the popular Qubit fluorometer designed to accurately measure protein quantity. |
Protein detection method attributes | ELISA | Western blotting | Mass spec |
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Advantages |
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Sensitivity | <5-10 pg/mL | low femtogram to high attogram* | attomolar range (1018) |
Lysis buffer compatibility |
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| Detergents and high salts must be removed prior to analysis |
Typical total protein required | 0.1 µg/mL - 1 µ g/mL | 1 µg- 50 µg | <1 µg |
Equipment required | Plate reader | X-ray film or CCD imaging equipment | Mass spectrometer |
*With high sensitivity HRP substrates, such as SuperSignal West Atto Ultimate Sensitivity Substrate
Discover more about western blotting, ELISA,mass spec
For Research Use Only. Not for use in diagnostic procedures.