Shop All Protein Quantitation Assay Kits, Reagents & Standards

Pierce™ Bovine Serum Albumin Standard, 2 mg/mL (Thermo Scientific™)

Thermo Scientific Pierce BSA Protein Assay Standards are high-quality reference samples for generating accurate standard curves and calibration controls in total protein assays.

Features of BSA Protein Assay Standards:

Convenient—50 mL bottle
Universal—recognized as the industry standard for protein quantitation in colorimetric protein assays
Pure and stable—supplied in ultrapure 0.9% saline solution with 0.05% sodium azide; room temperature stable
Accurate and consistent—precisely formulated at 2.00 +/-0.03 mg/mL compared to an NIST reference

These bovine serum albumin (BSA) solutions are protein concentration reference standards for use in BCA, Bradford and other protein assay protocols. BSA is the universally accepted reference protein for total protein quantitation. The albumin standard is precisely formulated at 2 mg/mL in an ultrapure 0.9% sodium chloride (saline) solution. The concentration of the stock solution is calibrated by direct comparison to purified BSA (Fraction V) from the National Institute of Standards and Technology (NIST).

Applications:
• Protein assay quantitation standard (BCA Protein Assay, Coomassie-Bradford Assay, etc.)
• Protein recovery control for desalting and other column procedures
• General calibration of spectrophotometer UV-lamp (absorbance at 280nm)

Selection of a protein standard is potentially the greatest source of error in any protein assay. The best choice for a standard is a purified, known concentration of the most abundant protein contained in the samples being tested. Often, a highly purified, known concentration of the protein of interest is not available or it is too expensive to use as the standard, or the sample itself is a mixture of many proteins (e.g., cell lysate). In such cases, the best standard is one that will produce a normal (i.e., average) color response curve with the selected protein assay method and is readily available to any researcher. BSA is such a protein, and the Pierce Albumin Standards are the most convenient source of ready-to-use BSA standard.

For greatest accuracy in estimating total protein concentration in unknown samples, it is essential to include a standard curve each time the assay is performed. This is particularly true for the protein assay methods that produce non-linear standard curves. Deciding on the number of standards and replicates used to define the standard curve depends upon the degree of non-linearity in the standard curve and the degree of accuracy required. In general, fewer points are needed to construct a standard curve if the color response is linear. Typically, standard curves are constructed using at least two replicates for each point on the curve.

Related Products
Pierce™ Bovine Serum Albumin Standard Pre-Diluted Set

Pierce™ Coomassie (Bradford) Protein Assay Kit (Thermo Scientific™)

The Thermo Scientific Pierce Coomassie Protein Assay Kit is a ready-to-use, stable formulation of the traditional Bradford assay reagent to measure (A595 nm) total protein concentration compared to a protein standard.

The Pierce Coomassie Protein Assay Kit is a ready-to-use formulation of the popular assay reagent originally described by Bradford in 1976. When mixed with a protein solution, the acidic coomassie-dye reagent changes color from brown to blue in proportion the amount of protein present in the sample. Protein determinations are made by comparison to the color response of protein assay standards, usually prepared as a series of known dilutions of bovine serum albumin (BSA) or bovine gamma globulin (BGG). The kit includes Coomassie Protein Assay Reagent and a package of Albumin Standard Ampules. The simple procedure is adaptable to nearly any volume scale, including test tubes, cuvettes and microplates.

Features of the Coomassie Protein Assay Kit:

Bradford reagent—stable, ready-to-use kit of the classical Bradford assay reagent
Colorimetric—measure with a standard spectrophotometer or plate reader (595 nm)
Easy to use—single reagent; no working reagent preparation required
Fast—almost immediate color development; add, mix and read results
Broad range—detects protein concentration in the range 1 to 1500 µg/mL
Flexible—microplate and cuvette protocols provided with the instructions and adaptable to several target working ranges

How the Coomassie (Bradford) Assay Detects Protein
Use of coomassie G-250 dye in a colorimetric reagent for the detection and quantitation of total protein was first described by Dr. Marion Bradford in 1976. In the acidic environment of the reagent, protein binds to the coomassie dye. This results in a spectral shift from the reddish/brown form of the dye (absorbance maximum at 465nm) to the blue form of the dye (absorbance maximum at 610 nm). The difference between the two forms of the dye is greatest at 595 nm, so that is the optimal wavelength to measure the blue color from the coomassie dye-protein complex. If desired, the blue color can be measured at any wavelength between 575 nm and 615 nm. At the two extremes (575 nm and 615 nm) there is a loss of about 10% in the measured amount of color (absorbance) compared to that obtained at 595 nm.

Development of color in coomassie dye-based (Bradford) protein assays has been associated with the presence of certain basic amino acids (primarily arginine, lysine and histidine) in the protein. Van der Waals forces and hydrophobic interactions also participate in the binding of the dye by protein. The number of Coomassie dye ligands bound to each protein molecule is approximately proportional to the number of positive charges found on the protein. Free amino acids, peptides and low molecular weight proteins do not produce color with coomassie dye reagents. In general, the mass of a peptide or protein must be at least 3000 daltons to be assayed with this reagent. The assay is performed at room temperature and no special equipment is required. Simply add the sample to the tube containing reagent and the resultant blue color is measured at 595 nm following a short room-temperature incubation. The coomassie dye containing protein assay is compatible with most salts, solvents, buffers, thiols, reducing substances and metal chelating agents encountered in protein samples.

For more information, see the article "Chemistry of Protein Assays" in the Protein Methods Library.

Related Products
Pierce™ Coomassie Plus (Bradford) Assay Kit
Pierce™ Detergent Compatible Bradford Assay Kit
Ampule Breakers

Pierce™ Bovine Gamma Globulin Standard Ampules, 2 mg/mL (Thermo Scientific™)

Thermo Scientific Pierce BGG Protein Assay Standards are high-quality reference samples for generating accurate standard curves and calibration controls in total protein assays.

Features of BGG Protein Assay Standards:

Convenient—1 mL ampules
Antibody standard—best reference standard for immunoglobulin quantitation in colorimetric protein assays
Bradford standard—best general protein standard in coomassie-based (Bradford) protein assays
Pure and stable—supplied in ultrapure 0.9% saline solution with 0.05% sodium azide; room temperature stable
Accurate and consistent—precisely formulated at 2.00 +/-0.03 mg/mL, ensuring excellent lot-to-lot consistency

These bovine gamma globulin (BGG) solutions are protein concentration reference standards for use in BCA, Bradford and other protein assay protocols. BGG is an accepted reference protein for total protein quantitation of purified antibodies or immunoglobulin-rich samples. The IgG standard is precisely formulated at 2 mg/mL in an ultrapure 0.9% sodium chloride (saline) solution. The concentration of the stock solution of purified BGG (Fraction II) is calibrated by direct comparison to an internal standard to ensure lot-to-lot consistency and accuracy.

Applications:
• Protein assay immunoglobulin quantitation standard (Coomassie-Bradford Assay, etc.)
• Antibody recovery control for desalting and other column procedures
• Loading control for SDS-PAGE
• General calibration of spectrophotometer UV-lamp (absorbance at 280nm)

Selection of a protein standard is potentially the greatest source of error in any protein assay. The best choice for an antibody quantitation standard is a purified, known concentration of the specific immunoglobulin being tested. Often this is not available or it is too expensive to use as a standard. In such cases, the best standard is one that will produce a representative color response curve with the selected protein assay and is readily available to any researcher. BGG provides this function for all kinds of immunoglobulin samples, including IgG.

For greatest accuracy in estimating total protein concentration in unknown samples, it is essential to include a standard curve each time the assay is performed. This is particularly true for the protein assay methods that produce non-linear standard curves. Deciding on the number of standards and replicates used to define the standard curve depends upon the degree of non-linearity in the standard curve and the degree of accuracy required. In general, fewer points are needed to construct a standard curve if the color response is linear. Typically, standard curves are constructed using at least two replicates for each point on the curve.

For added safety when opening glass ampules, consider using our Ampule Breakers, which are disposable safety devices that protect the fingers when breaking open a glass ampule.

Related Products
Pierce™ Bovine Gamma Globulin Standard Pre-Diluted Set

Pierce™ Bovine Gamma Globulin Standard Pre-Diluted Set (Thermo Scientific™)

Thermo Scientific Pierce BGG Protein Assay Standards are high-quality reference samples for generating accurate standard curves and calibration controls in total protein assays.

Features of BGG Protein Assay Standards:

Convenient—complete set containing seven ready-to-use dilutions
Antibody standard—best reference standard for immunoglobulin quantitation in colorimetric protein assays
Bradford standard—best general protein standard in coomassie-based (Bradford) protein assays
Pure and stable—supplied in ultrapure 0.9% saline solution with 0.05% sodium azide; room temperature stable
Accurate and consistent—precisely formulated at 2.00 +/-0.03 mg/mL, ensuring excellent lot-to-lot consistency

These bovine gamma globulin (BGG) solutions are protein concentration reference standards for use in BCA, Bradford and other protein assay protocols. BGG is an accepted reference protein for total protein quantitation of purified antibodies or immunoglobulin-rich samples. The IgG standard is precisely formulated at 2 mg/mL in an ultrapure 0.9% sodium chloride (saline) solution. The concentration of the stock solution of purified BGG (Fraction II) is calibrated by direct comparison to an internal standard to ensure lot-to-lot consistency and accuracy.

Applications:
• Protein assay immunoglobulin quantitation standard (Coomassie-Bradford Assay, etc.)
• Antibody recovery control for desalting and other column procedures
• Loading control for SDS-PAGE
• General calibration of spectrophotometer UV-lamp (absorbance at 280nm)

Selection of a protein standard is potentially the greatest source of error in any protein assay. The best choice for an antibody quantitation standard is a purified, known concentration of the specific immunoglobulin being tested. Often this is not available or it is too expensive to use as a standard. In such cases, the best standard is one that will produce a representative color response curve with the selected protein assay and is readily available to any researcher. BGG provides this function for all kinds of immunoglobulin samples, including IgG.

For greatest accuracy in estimating total protein concentration in unknown samples, it is essential to include a standard curve each time the assay is performed. This is particularly true for the protein assay methods that produce non-linear standard curves. Deciding on the number of standards and replicates used to define the standard curve depends upon the degree of non-linearity in the standard curve and the degree of accuracy required. In general, fewer points are needed to construct a standard curve if the color response is linear. Typically, standard curves are constructed using at least two replicates for each point on the curve.

Related Products
Pierce™ Bovine Gamma Globulin Standard Ampules, 2 mg/mL

Pierce™ Modified Lowry Protein Assay Kit (Thermo Scientific™)

The Thermo Scientific Pierce Modified Lowry Protein Assay is a stable form of a traditional, 2-component, folin phenol- and copper-based reagent system to measure (A750nm) total protein concentration compared to a protein standard.

Features of Modified Lowry Protein Assay:

Popular method—widely cited in protein research literature
Colorimetric—measure with a standard spectrophotometer or plate reader (750nm)
Stable—uses a modified cupric sulfate-tartrate reagent that is stable at room temperature
Large range—exhibits good linearity in the range 1 to 1500 µg/mL (tested with BSA protein)
Convenient—microplate and cuvette protocols provided with the instructions

The Modified Lowry Protein Assay Kit combines a stabilized formulation of the original Lowry Reagents and the essential Folin-Ciocalteu Phenol reagent in a complete kit for accurately determining protein concentration in a variety of samples types. Although newer protein assay methods provide greater speed and convenience, the Lowry method remains a popular, accurate and useful option for many applications.

Related Products
Pierce™ Modified Lowry Protein Assay Kit
Ampule Breakers

ProteinSEQ™ Protein A Core Kit (Applied Biosystems™)

The ProteinSEQ™ Protein A Core Kit is a partial kit that enables users to customize their ProteinSEQ protein measurements for leeched Protein A, or any other ligand used in their purification columns. It contains the buffers and reagents, but not the antibody-loaded capture beads, needed to enable PCR-based quantitation of Protein A or any other purification ligand. The kit contains sufficient reagents for 200 reactions. For a kit that includes Protein A capture beads and standards, for use in the quantitation of Protein A, please see the ProteinSEQ™ Protein A Quantitation Kit (Cat. No. 4469343).

• Quantitate leeched protein contaminants with ultra-high sensitivity
• Accelerate process development with more valid test results per run
• Reduce assay hands-on time with less sample dilution and preparation
• Achieve unprecedented productivity with near-5-log assay dynamic range

ProteinSEQ Protein A quantitation
The ProteinSEQ assay platform uses qPCR technology to measure process contaminants with extraordinary sensitivity and unparalleled dynamic range. Protein A or other purification ligands leeching from purification columns can be accurately measured with outstanding spike recovery and dilutional linearity.

Streamlined assay worflow
The ProteinSEQ system is the only instrument platform-based Protein A and purification ligand quantitation solution to offer single-digit picogram sensitivity and nearly 5 logs of dynamic range. Laboratory workflow is greatly accelerated due to the reduction in the number of sample dilutions and replicates in each run.

Pierce™ Bovine Serum Albumin Standard Pre-Diluted Set (Thermo Scientific™)

Thermo Scientific Pierce BSA Protein Assay Standards are high-quality reference samples for generating accurate standard curves and calibration controls in total protein assays.

Features of BSA Protein Assay Standards:

Convenient—complete set containing seven ready-to-use dilutions
Universal—recognized as the industry standard for protein quantitation in colorimetric protein assays
Pure and stable—supplied in ultrapure 0.9% saline solution with 0.05% sodium azide

These bovine serum albumin (BSA) solutions are protein concentration reference standards for use in BCA, Bradford and other protein assay protocols. BSA is the universally accepted reference protein for total protein quantitation. The set of standards is prepared from a stock solution that is calibrated by direct comparison to purified BSA (Fraction V) from the National Institute of Standards and Technology (NIST).

Applications:
• Protein assay quantitation standard (BCA Protein Assay, Coomassie-Bradford Assay, etc.)
• Protein recovery control for desalting and other column procedures
• General calibration of spectrophotometer UV-lamp (absorbance at 280nm)

Selection of a protein standard is potentially the greatest source of error in any protein assay. The best choice for a standard is a purified, known concentration of the most abundant protein contained in the samples being tested. Often, a highly purified, known concentration of the protein of interest is not available or it is too expensive to use as the standard, or the sample itself is a mixture of many proteins (e.g., cell lysate). In such cases, the best standard is one that will produce a normal (i.e., average) color response curve with the selected protein assay method and is readily available to any researcher. BSA is such a protein, and the Pierce Albumin Standards are the most convenient source of ready-to-use BSA standard.

For greatest accuracy in estimating total protein concentration in unknown samples, it is essential to include a standard curve each time the assay is performed. This is particularly true for the protein assay methods that produce non-linear standard curves. Deciding on the number of standards and replicates used to define the standard curve depends upon the degree of non-linearity in the standard curve and the degree of accuracy required. In general, fewer points are needed to construct a standard curve if the color response is linear. Typically, standard curves are constructed using at least two replicates for each point on the curve.

Related Products
Pierce™ Bovine Serum Albumin Standard Ampules, 2 mg/mL

Compat-Able™ Protein Assay Preparation Reagent Kit (Thermo Scientific™)

The Thermo Scientific Compat-Able Protein Assay Preparation Set eliminates interfering substances from protein assay samples to make them compatible with BCA and Bradford assays.

The Compat-Able Reagents remove salts, detergents, reducing agents and other substances from protein samples to eliminate interference with protein assays. The protocol selectively precipitates all of the protein, thereby allowing the non-protein components to be easily decanted and removed. The purified, precipitated protein is redissolved in water, making it ready for accurate determination of the protein concentration by standard protein assay methods. The procedure is accomplished in less than 10 minutes with just a few simple steps. The Compat-Able Preparation Set is available individually or conveniently bundled with either the Thermo Scientific Pierce BCA Protein Assay Kit or the Thermo Scientific Pierce Coomassie Plus Protein Assay Kit.

Features of the Compat-Able Protein Assay Preparation Reagent Kit:

Eliminate incompatibilities—removes nearly any substance that interferes with colorimetric protein assays: salts, detergents, denaturants, reducing agents, and even IEF sample buffer
Simple and fast—prepare samples in less than 10 minutes with four simple steps
Convenient—reagent set is easier to use than homemade TCA or acetone precipitation reagents, thereby producing more consistent results
Scalable—microcentrifuge method is easily scaled to larger centrifuge tube volumes for special needs
Validated—tested and offered together with BCA and Coomassie Plus Protein Assay Kits; protocol integrates control and normalization procedures

Related Products
Compat-Able™ BCA Protein Assay Kit
Compat-Able™ Coomassie Plus Protein Assay Kit

Pierce™ Detergent Compatible Bradford Assay Kit (Thermo Scientific™)

The Thermo Scientific™ Pierce™ Detergent Compatible Bradford Assay Kit is a quick and ready-to-use modification of the well-known Bradford coomassie dye-binding, colorimetric method for total protein quantitation. Proprietary additives to the Bradford Reagent make it compatible with up to 1% or higher of detergents and lysis reagents that are commonly used in life science research, including Triton® X-100 and NP-40.

• Convenient— detergent-free standard curve
• Flexible— compatible with samples both with and without detergent
• Minimal sample—requires only 10 µL for microplate procedure
• Colorimetric—measure with a standard spectrophotometer or plate reader (595 nm)
• Easy to use— single reagent, no working reagent preparation required
• Fast—10 minute incubation at room temperature
• Broad range—detects protein concentration in the ranges of 2 to 1500 µg/mL

Similar to the Bradford method, coomassie dye binds protein in an acidic medium causing an immediate shift in absorption maximum from 465 nm to 595 nm with a concomitant color change from green to blue. In addition, the assay is complete in just 10 minutes.

The protein assay can be performed in either test tube or microplate format. The standard working range is 100-1500 µg/mL with up to 1% detergent (or higher in some cases). Protein concentrations are estimated by reference to absorbances obtained for a series of standard protein dilutions, typically bovine serum albumin (BSA), which are assayed alongside the unknown samples. Because the color response with coomassie dye is non-linear with increasing protein concentration, a standard curve must be completed with each assay. Standards can be used directly without preparing them in the same detergent found in the test samples.

Related products
Ampule Breakers

Pierce™ BCA Protein Assay Reagent A (Thermo Scientific™)

BCA Protein Assay Reagent A is a component of the Thermo Scientific Pierce BCA Protein Assay Kit, a two-component, high-precision, detergent-compatible assay reagent set to measure (A562nm) total protein concentration compared to a protein standard.

Used in more labs than any other detergent-compatible protein assay, Pierce BCA Reagents provide accurate determination of protein concentration with most sample types encountered in protein research. The Pierce BCA Assay can be used to assess yields in whole cell lysates and affinity-column fractions, as well as to monitor protein contamination in industrial applications. Compared to most dye-binding methods, the BCA Assay is affected much less by protein compositional differences, providing greater protein-to-protein uniformity.

Features of the Thermo Scientific Pierce BCA Protein Assay Kit (reagents A and B) :

Colorimetric—estimate visually or measure with a standard spectrophotometer or plate reader (562nm)
Excellent uniformity—exhibits less protein-to-protein variation than dye-binding methods
Compatible—unaffected by typical concentrations of most ionic and nonionic detergents
Moderately fast—much easier and four times faster than the classical Lowry method
High linearity—linear working range for BSA equals 20 to 2000 µg/mL
Sensitive—detect down to 5 µg/mL with the enhanced protocol

Applications:
• Studying protein:protein interactions
• Measuring column fractions after affinity chromatography
• Estimating percent recovery of membrane proteins from cell extracts
• High-throughput screening of fusion proteins

How the BCA Protein Assay Detects Protein:
The BCA Protein Assay combines the well-known reduction of Cu2+ to Cu1+ by protein in an alkaline medium with the highly sensitive and selective colorimetric detection of the cuprous cation (Cu1+) by bicinchoninic acid. The first step is the chelation of copper with protein in an alkaline environment to form a light blue complex. In this reaction, known as the biuret reaction, peptides containing three or more amino acid residues form a colored chelate complex with cupric ions in an alkaline environment containing sodium potassium tartrate.

In the second step of the color development reaction, bicinchoninic acid (BCA) reacts with the reduced (cuprous) cation that was formed in step one. The intense purple-colored reaction product results from the chelation of two molecules of BCA with one cuprous ion. The BCA/copper complex is water-soluble and exhibits a strong linear absorbance at 562 nm with increasing protein concentrations. The BCA reagent is approximately 100 times more sensitive (lower limit of detection) than the pale blue color of the first reaction.

The reaction that leads to BCA color formation is strongly influenced by four amino acid residues (cysteine or cystine, tyrosine, and tryptophan) in the amino acid sequence of the protein. However, unlike the Coomassie dye-binding methods, the universal peptide backbone also contributes to color formation, helping to minimize variability caused by protein compositional differences.

For more information, see the article "Chemistry of Protein Assays" in the Protein Methods Library.

Related Products
Pierce™ BCA Protein Assay Kit
Pierce™ BCA Protein Assay Reagent B
Pierce™ BCA Solid

Pierce™ Bovine Serum Albumin Standard Ampules, 2 mg/mL (Thermo Scientific™)

Thermo Scientific Pierce BSA Protein Assay Standards are high-quality reference samples for generating accurate standard curves and calibration controls in total protein assays.

Features of BSA Protein Assay Standards:

Convenient—1 mL ampules
Universal—recognized as the industry standard for protein quantitation in colorimetric protein assays
Pure and stable—supplied in ultrapure 0.9% saline solution with 0.05% sodium azide; room temperature stable
Accurate and consistent—precisely formulated at 2.00 +/-0.03 mg/mL compared to an NIST reference

These bovine serum albumin (BSA) solutions are protein concentration reference standards for use in BCA, Bradford and other protein assay protocols. BSA is the universally accepted reference protein for total protein quantitation. The albumin standard is precisely formulated at 2 mg/mL in an ultrapure 0.9% sodium chloride (saline) solution. The concentration of the stock solution is calibrated by direct comparison to purified BSA (Fraction V) from the National Institute of Standards and Technology (NIST).

Applications:
• Protein assay quantitation standard (BCA Protein Assay, Coomassie-Bradford Assay, etc.)
• Protein recovery control for desalting and other column procedures
• General calibration of spectrophotometer UV-lamp (absorbance at 280nm)

Selection of a protein standard is potentially the greatest source of error in any protein assay. The best choice for a standard is a purified, known concentration of the most abundant protein contained in the samples being tested. Often, a highly purified, known concentration of the protein of interest is not available or it is too expensive to use as the standard, or the sample itself is a mixture of many proteins (e.g., cell lysate). In such cases, the best standard is one that will produce a normal (i.e., average) color response curve with the selected protein assay method and is readily available to any researcher. BSA is such a protein, and the Pierce Albumin Standards are the most convenient source of ready-to-use BSA standard.

For greatest accuracy in estimating total protein concentration in unknown samples, it is essential to include a standard curve each time the assay is performed. This is particularly true for the protein assay methods that produce non-linear standard curves. Deciding on the number of standards and replicates used to define the standard curve depends upon the degree of non-linearity in the standard curve and the degree of accuracy required. In general, fewer points are needed to construct a standard curve if the color response is linear. Typically, standard curves are constructed using at least two replicates for each point on the curve.

For added safety when opening glass ampules, consider using our Ampule Breakers, which are disposable safety devices that protect the fingers when breaking open a glass ampule.

Related Products
Pierce™ Bovine Serum Albumin Standard Pre-Diluted Set

Compat-Able™ BCA Protein Assay Kit (Thermo Scientific™)

The Thermo Scientific Compat-Able BCA Protein Assay Kit eliminates interfering substances from protein assay samples to make them compatible with BCA and Bradford assays.

The Compat-Able Reagents remove salts, detergents, reducing agents and other substances from protein samples to eliminate interference with protein assays. The protocol selectively precipitates all of the protein, thereby allowing the non-protein components to be easily decanted and removed. The purified, precipitated protein is redissolved in water, making it ready for accurate determination of the protein concentration by standard protein assay methods. The procedure is accomplished in less than 10 minutes with just a few simple steps. The Compat-Able Preparation Set is available individually or conveniently bundled with either the Thermo Scientific Pierce BCA Protein Assay Kit or the Thermo Scientific Pierce Coomassie Plus Protein Assay Kit.

Features of the Compat-Able Protein Assay Preparation Reagent Kit:

Eliminate incompatibilities—removes nearly any substance that interferes with colorimetric protein assays: salts, detergents, denaturants, reducing agents, and even IEF sample buffer
Simple and fast—prepare samples in less than 10 minutes with four simple steps
Convenient—reagent set is easier to use than homemade TCA or acetone precipitation reagents, thereby producing more consistent results
Scalable—microcentrifuge method is easily scaled to larger centrifuge tube volumes for special needs
Validated—tested and offered together with BCA and Coomassie Plus Protein Assay Kits; protocol integrates control and normalization procedures

Related Products
Compat-Able™ Protein Assay Preparation Reagent Kit
Compat-Able™ Coomassie Plus Protein Assay Kit
Ampule Breakers

Pierce™ BCA Protein Assay Kit (Thermo Scientific™)

The Thermo Scientific Pierce BCA Protein Assay Kit is a two-component, high-precision, detergent-compatible assay reagent set to measure (A562nm) total protein concentration compared to a protein standard. Used in more labs than any other detergent-compatible protein assay, Pierce BCA Reagents provide accurate determination of protein concentration with most sample types encountered in protein research. The Pierce BCA Assay can be used to assess yields in whole cell lysates and affinity-column fractions, as well as to monitor protein contamination in industrial applications. Compared to most dye-binding methods, the BCA Assay is affected much less by protein compositional differences, providing greater protein-to-protein uniformity.

Features of the BCA Protein Assay Kit:
• Colorimetric—estimate visually or measure with a standard spectrophotometer or plate reader (562nm)
• Excellent uniformity—exhibits less protein-to-protein variation than dye-binding methods
• Compatible—unaffected by typical concentrations of most ionic and nonionic detergents
• Moderately fast—much easier and four times faster than the classical Lowry method
• High linearity—linear working range for BSA equals 20 to 2000 µg/mL
• Sensitive—detect down to 5 µg/mL with the enhanced protocol

Applications:
• Studying protein:protein interactions
• Measuring column fractions after affinity chromatography
• Estimating percent recovery of membrane proteins from cell extracts
• High-throughput screening of fusion proteins

How the BCA Protein Assay Detects Protein:
The BCA Protein Assay combines the well-known reduction of Cu2+ to Cu1+ by protein in an alkaline medium with the highly sensitive and selective colorimetric detection of the cuprous cation (Cu1+) by bicinchoninic acid. The first step is the chelation of copper with protein in an alkaline environment to form a light blue complex. In this reaction, known as the biuret reaction, peptides containing three or more amino acid residues form a colored chelate complex with cupric ions in an alkaline environment containing sodium potassium tartrate.

In the second step of the color development reaction, bicinchoninic acid (BCA) reacts with the reduced (cuprous) cation that was formed in step one. The intense purple-colored reaction product results from the chelation of two molecules of BCA with one cuprous ion. The BCA/copper complex is water-soluble and exhibits a strong linear absorbance at 562 nm with increasing protein concentrations. The BCA reagent is approximately 100 times more sensitive (lower limit of detection) than the pale blue color of the first reaction.

The reaction that leads to BCA color formation is strongly influenced by four amino acid residues (cysteine or cystine, tyrosine, and tryptophan) in the amino acid sequence of the protein. However, unlike the Coomassie dye-binding methods, the universal peptide backbone also contributes to color formation, helping to minimize variability caused by protein compositional differences.

For more information, see the article "Chemistry of Protein Assays" in the Protein Methods Library.

Pierce™ Detergent Compatible Bradford Assay Reagent (Thermo Scientific™)

The Thermo Scientific™ Pierce™ Detergent Compatible Bradford Assay Reagent is a quick and ready-to-use modification of the well-known Bradford coomassie dye-binding, colorimetric method for total protein quantitation. Proprietary additives to the Bradford Reagent make it compatible with up to 1% or higher of detergents and lysis reagents that are commonly used in life science research, including Triton® X-100 and NP-40.

• Convenient—detergent-free standard curve
• Flexible—compatible with samples both with and without detergent
• Minimal sample—requires only 10 µL for microplate procedure
• Colorimetric—measure with a standard spectrophotometer or plate reader (595 nm)
• Easy to use—single reagent, no working reagent preparation required
• Fast—10 minute incubation at room temperature
• Broad range—detects protein concentration in the ranges of 2 to 1500 µg/mL

Similar to the Bradford method, coomassie dye binds protein in an acidic medium causing an immediate shift in absorption maximum from 465 nm to 595 nm with a concomitant color change from green to blue. In addition, the assay is complete in just 10 minutes.

The protein assay can be performed in either test tube or microplate format. The standard working range is 100-1500 µg/mL with up to 1% detergent (or higher in some cases). Protein concentrations are estimated by reference to absorbances obtained for a series of standard protein dilutions, typically bovine serum albumin (BSA), which are assayed alongside the unknown samples. Because the color response with coomassie dye is non-linear with increasing protein concentration, a standard curve must be completed with each assay. Standards can be used directly without preparing them in the same detergent found in the test samples.

Easy-Titer™ Rabbit IgG Assay Kit (Thermo Scientific™)

The Thermo Scientific Easy-Titer Rabbit IgG Assay Kit includes antibody-sensitized microspheres to measure the specific concentration of antibodies by an easy and rapid microagglutination technique using standard microplates and UV-Vis plate reader (spectrophotometer). This kit is specific for rabbit IgG and, unlike total protein assays, can specifically measure the concentration of target antibody in samples (e.g., serum, plasma, culture supernatant) that contain other proteins. It is sensitive, requiring very small sample volumes. Antibody concentration is determined from the assay response (absorbance) by comparison to a standard curve prepared using dilutions of a known antibody sample (sold separately).

General features of Easy-Titer Antibody Assay Kits:

Antibody-based specificity—measure concentration of target antibody in a sample, not just total protein; no need to purify antibody to assess its concentration
Faster and easier than ELISA—three-component, homogenous assay; 10 minutes total incubation time
No special equipment needed—uses standard vortex mixer, pipetter, 96-well microplate, plate shaker and reader (measure absorbance at 340nm or 400nm)
Sensitive—assay range (standard curve) 8 to 500 ng/mL; use sample at 15 to 300 ng/mL for optimal results
Reproducible—coefficient of variation < 5%; error depends on dilution and pipetting technique
Antibody standards sold separately—see product list for suggested products; use any antibody standard with proper target identity and known concentration (greater than 10 µg/mL)
Kits for five popular targets—choose a kit specific for a particular species and class of immunoglobulin; no cross-reaction with other species and classes of the target antibody

Easy-Titer Assay Kits detect and measure specific target antibodies using agglutination of microspheres that are coated ("sensitized") with the specific anti-IgG or IgM polyclonal antibodies. In the appropriate aqueous buffer (supplied in kit), the monodispersed antibody-coated microspheres (> 1 µM diameter) have highest absorptivity (λ-max) to incident light having a wavelength (340nm) that is equal to approximately half their diameter. When sample is added, two or more microspheres bind to each antibody target via their coated specific polyclonal antibodies, and this agglutination into effectively larger apparent spheres results in proportional decrease in absorptivity (lower absorbance).

Typical microagglutination assays depend on a change in light-scattering and corresponding change in transmittance, to which absorbance is inversely related. Easy-Titer Assay Kits use a special dilution buffer whose refractive index eliminates the effect of light-scattering on the monodispersed microspheres for the measurement wavelength used. Because of this, the final 10- to 20-fold dilution of the sample for use in the assay must be done using the Dilution Buffer supplied in the kit.

Easy-Titer Antibody Assays are faster and easier than ELISA:
• Prepare standards (5 to 500 ng/mL) by diluting purified antibody in Kit Dilution Buffer.
• Prepare samples by diluting in Dilution Buffer to within assay range (8 to 500 ng/mL).
• Vortex vial of microsphere beads to create homogeneous suspension.
• Pipette 20 µL of bead suspension and 20 µL of each sample and standard into 96-well microplate wells.
• Incubate microplate for 5 minutes with vigorous mixing.
• Add 100 µL of Kit Blocking Reagent.
• Incubate microplate for 5 minutes with vigorous mixing.
• Measure absorbance on standard plate reader (340nm or 405nm).
• Plot standard curve and interpolate samples to determine concentration.

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