ELISA Blocking Buffer SuperBlock

Select from a wide variety of easy-to-use and reliable blocking buffers and reagents for ELISA applications. ELISA buffers and reagents are important components to develop the best assay performance for high sensitivity, low background, and blocking non-specific binding. Find below blocking buffers, coating buffers, wash buffers, and recommended lysis buffers for success in ELISA applications.

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Invitrogen ELISA Buffer Kit

For a complete set of ELISA reagents, Invitrogen ELISA Buffer Kit, Cat. No. CNB0011, includes: 2 Coating Buffers (pH 7.4 and pH 9.4), Assay Buffer (Blocking Buffer), Wash Buffer, Stabilized TMB, and Stop Solution. Pair this with Matched Antibody Pair kits that come with matched antibody pairs, standards, and Streptavidin-HRP.

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Recommended ELISA blocking buffers

An ELISA blocking buffer is a solution of irrelevant protein, mixture of proteins, or other compound that passively adsorbs to all remaining binding surfaces of the plate that are not occupied by the coated protein. The blocking buffer is effective if it improves the sensitivity of an assay by reducing background signal and improving the signal-to-noise ratio. The ideal blocking buffer will bind to all potential sites of nonspecific interaction, eliminating background altogether, without altering or obscuring the epitope for antibody binding. Select from easy-to-use and reliable blocking buffers for ELISA applications below.

Blocking buffer Blocking agent Highlights When to use Available formats (Catalog No.)
ELISA Diluent BSA
  • Single purified protein provides fewer chances of cross-reaction with assay components than serum or milk solutions
  • Performs well with a wide range of antibodies and antibody combinations
Concentrate (DS98200)
SuperBlock Blocking Buffer Serum and biotin-free single purified glycoprotein
  • Protein-based formulation does not contain any immunoglobulins, albumin, or endogenous biotin, making it compatible in many situations where traditional blocking agents fail
  • Compatible with streptavidin systems
  • Blocks in less than 10 minutes
  • May be used as a protein stabilizer for drying antigen or antibody coated microplates
  • As a protein stabilizer when storing coated plates
  • With streptavidin systems
PBS (37515)
TBS (37535)
PBST (37516)
TBST (37536)
Pierce Protein-Free Blocking Buffers Non-protein blocking compound
  • Helps minimize or eliminate crossreactivity associated with protein based blocking buffers
  • Sample-and-antibody combinations require the elimination of all possible exogenous animal proteins in the assay system to avoid cross reaction or quenching of the desired probe function
  • Use when protein-based blockers cause high background
PBS (37572)
TBS (37570)
PBST (37573)
TBST (37571)


Recommended ELISA plate coating buffer, wash buffer, and stop solution

Buffer Composition When to use Available Formats (Catalog No.)
ELISA Plate Coating Buffer 10 mM phosphate buffer, pH 7.4 Recommended for most proteins Concentrate (CB07100)
50 mM carbonate buffer, pH 9.4 High pH aids solubility of some proteins and peptides and ensures that most proteins are unprotonated with an overall negative charge, which helps when binding to positively charged plates. Concentrate (CB01100)
Powder (28382)
ELISA Wash Buffer Tris-buffered saline with 0.05% Tween 20   Concentrate (28360)
Powder (28379)
Phosphate-buffered saline with 0.05% Tween 20   Concentrate (28352)
Powder (00-0400-46)
ELISA Diluent Phosphate-buffered solution with BSA ELISA Diluent is designed to be used as (1) a blocking reagent for ELISA plates, (2) a diluent for ELISA standards and samples, (3) as a diluent for the detector antibody, and (4) as a diluent for the HRP conjugate Concentrate (00-4202-56)
Stop Solution 0.16 M sulfuric acid Use as a stop solution when horseradish peroxidase and TMB substrate solution are used Ready-to-use (N600)


Recommended cell lysis buffers

Buffer Composition When to use Available Formats (Catalog No.)
NP40 Cell Lysis Buffer 50 mM Tris, pH 7.4, 250 mM NaCl, 5 mM EDTA, 50 mM NaF, 1 mM Na3VO4, 1% NP40, 0.02% NaN3 Most popular and recommended for lysis of cell membrane and cytoplasmic proteins to measure intracellular proteins using ELISA Ready-to-use (FNN0021)
RIPA Cell Lysis Buffer 10 mM Tris, pH 7.4, 100 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1 mM NaF, 20 mM Na4P2O7, 2 mM Na3VO4, 1% Triton X-100, 10% glycerol, 0.1% SDS, 0.5% deoxycholate Need a harsher buffer than NP40 or when nuclear disruption is also needed Ready-to-use (FNN0011)
Tissue Extraction Reagent I 50 mM Tris, pH 7.4, 250 mM NaCl, 5 mM EDTA, 2 mM Na3VO4, 1 mM NaF, 20 mM Na4P2O7, 0.02% NaN3, and detergent For total protein extraction from tissue samples or alternate formulation for total protein lysis if NP40 or RIPA is not sufficient Ready-to-use (FNN0071)


AP and HRP conjugates

Select from a wide variety of conjugates and detection probes for use in direct or indirect sandwich ELISA applications. Below are recommended biotin-binding probes including streptavidin and NeutrAvidin protein. Each protein binds four biotins per molecule with high affinity and selectivity. Streptavidin is most commonly used—it is non-glycosylated and exhibits low levels of nonspecific binding. NeutrAvidin has been processed to remove the carbohydrate and lower its isoelectric point, resulting in reduced nonspecific background staining.

For directly conjugated primary and secondary antibodies, search antibody enzyme conjugates


Recommended HRP Conjugates and AP Conjugates for ELISA applications

Conjugate Enzyme Recommended concentration* Benefits, when to use Available format (Catalog No.)
HRP- Streptavidin HRP 0.1 µg/mL Most popular 2 mg (21124)
1 mg (21126)
5 mg (21127)
Poly-HRP Streptavidin HPR 1:2,000 - 1:20,000 Use when exhibiting weak signal with typical HRP conjugates. Poly increases signal amplification and improves upon the signal provided by typical (non-Poly) streptavidin-HRP conjugates 0.25 mL (N200)
HRP-NeutrAvidin HRP 0.1 µg/mL Specially prepared form of avidin that decreases background in biotin-binding 2 mg (31001)
HRP-NeutrAvidin, High Sensitivity HRP 1:8,000 - 1:40,000 Detection of low levels of target without background; provides signal amplification like poly-HRP conjugates 0.5 mL (31030)
AP-Streptavidin AP 0.1 µg/mL   1 mg (21324)
AP-NeutrAvidin AP   Specially prepared form of avidin that decreases background in biotin-binding 2 mg (31002)

* The optimal dilution must be determined empirically for each specific application. Prepare dilutions immediately before use.

Search all available Avidin, Streptavidin, and Biotin conjugates
 


Recommended ELISA blocking buffers

An ELISA blocking buffer is a solution of irrelevant protein, mixture of proteins, or other compound that passively adsorbs to all remaining binding surfaces of the plate that are not occupied by the coated protein. The blocking buffer is effective if it improves the sensitivity of an assay by reducing background signal and improving the signal-to-noise ratio. The ideal blocking buffer will bind to all potential sites of nonspecific interaction, eliminating background altogether, without altering or obscuring the epitope for antibody binding. Select from easy-to-use and reliable blocking buffers for ELISA applications below.

Blocking buffer Blocking agent Highlights When to use Available formats (Catalog No.)
ELISA Diluent BSA
  • Single purified protein provides fewer chances of cross-reaction with assay components than serum or milk solutions
  • Performs well with a wide range of antibodies and antibody combinations
Concentrate (DS98200)
SuperBlock Blocking Buffer Serum and biotin-free single purified glycoprotein
  • Protein-based formulation does not contain any immunoglobulins, albumin, or endogenous biotin, making it compatible in many situations where traditional blocking agents fail
  • Compatible with streptavidin systems
  • Blocks in less than 10 minutes
  • May be used as a protein stabilizer for drying antigen or antibody coated microplates
  • As a protein stabilizer when storing coated plates
  • With streptavidin systems
PBS (37515)
TBS (37535)
PBST (37516)
TBST (37536)
Pierce Protein-Free Blocking Buffers Non-protein blocking compound
  • Helps minimize or eliminate crossreactivity associated with protein based blocking buffers
  • Sample-and-antibody combinations require the elimination of all possible exogenous animal proteins in the assay system to avoid cross reaction or quenching of the desired probe function
  • Use when protein-based blockers cause high background
PBS (37572)
TBS (37570)
PBST (37573)
TBST (37571)


Recommended ELISA plate coating buffer, wash buffer, and stop solution

Buffer Composition When to use Available Formats (Catalog No.)
ELISA Plate Coating Buffer 10 mM phosphate buffer, pH 7.4 Recommended for most proteins Concentrate (CB07100)
50 mM carbonate buffer, pH 9.4 High pH aids solubility of some proteins and peptides and ensures that most proteins are unprotonated with an overall negative charge, which helps when binding to positively charged plates. Concentrate (CB01100)
Powder (28382)
ELISA Wash Buffer Tris-buffered saline with 0.05% Tween 20   Concentrate (28360)
Powder (28379)
Phosphate-buffered saline with 0.05% Tween 20   Concentrate (28352)
Powder (00-0400-46)
ELISA Diluent Phosphate-buffered solution with BSA ELISA Diluent is designed to be used as (1) a blocking reagent for ELISA plates, (2) a diluent for ELISA standards and samples, (3) as a diluent for the detector antibody, and (4) as a diluent for the HRP conjugate Concentrate (00-4202-56)
Stop Solution 0.16 M sulfuric acid Use as a stop solution when horseradish peroxidase and TMB substrate solution are used Ready-to-use (N600)


Recommended cell lysis buffers

Buffer Composition When to use Available Formats (Catalog No.)
NP40 Cell Lysis Buffer 50 mM Tris, pH 7.4, 250 mM NaCl, 5 mM EDTA, 50 mM NaF, 1 mM Na3VO4, 1% NP40, 0.02% NaN3 Most popular and recommended for lysis of cell membrane and cytoplasmic proteins to measure intracellular proteins using ELISA Ready-to-use (FNN0021)
RIPA Cell Lysis Buffer 10 mM Tris, pH 7.4, 100 mM NaCl, 1 mM EDTA, 1 mM EGTA, 1 mM NaF, 20 mM Na4P2O7, 2 mM Na3VO4, 1% Triton X-100, 10% glycerol, 0.1% SDS, 0.5% deoxycholate Need a harsher buffer than NP40 or when nuclear disruption is also needed Ready-to-use (FNN0011)
Tissue Extraction Reagent I 50 mM Tris, pH 7.4, 250 mM NaCl, 5 mM EDTA, 2 mM Na3VO4, 1 mM NaF, 20 mM Na4P2O7, 0.02% NaN3, and detergent For total protein extraction from tissue samples or alternate formulation for total protein lysis if NP40 or RIPA is not sufficient Ready-to-use (FNN0071)


AP and HRP conjugates

Select from a wide variety of conjugates and detection probes for use in direct or indirect sandwich ELISA applications. Below are recommended biotin-binding probes including streptavidin and NeutrAvidin protein. Each protein binds four biotins per molecule with high affinity and selectivity. Streptavidin is most commonly used—it is non-glycosylated and exhibits low levels of nonspecific binding. NeutrAvidin has been processed to remove the carbohydrate and lower its isoelectric point, resulting in reduced nonspecific background staining.

For directly conjugated primary and secondary antibodies, search antibody enzyme conjugates


Recommended HRP Conjugates and AP Conjugates for ELISA applications

Conjugate Enzyme Recommended concentration* Benefits, when to use Available format (Catalog No.)
HRP- Streptavidin HRP 0.1 µg/mL Most popular 2 mg (21124)
1 mg (21126)
5 mg (21127)
Poly-HRP Streptavidin HPR 1:2,000 - 1:20,000 Use when exhibiting weak signal with typical HRP conjugates. Poly increases signal amplification and improves upon the signal provided by typical (non-Poly) streptavidin-HRP conjugates 0.25 mL (N200)
HRP-NeutrAvidin HRP 0.1 µg/mL Specially prepared form of avidin that decreases background in biotin-binding 2 mg (31001)
HRP-NeutrAvidin, High Sensitivity HRP 1:8,000 - 1:40,000 Detection of low levels of target without background; provides signal amplification like poly-HRP conjugates 0.5 mL (31030)
AP-Streptavidin AP 0.1 µg/mL   1 mg (21324)
AP-NeutrAvidin AP   Specially prepared form of avidin that decreases background in biotin-binding 2 mg (31002)

* The optimal dilution must be determined empirically for each specific application. Prepare dilutions immediately before use.

Search all available Avidin, Streptavidin, and Biotin conjugates
 


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