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.
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 |
|
|
Concentrate (DS98200) |
SuperBlock Blocking Buffer | Serum and biotin-free single purified glycoprotein |
|
|
PBS (37515) TBS (37535) PBST (37516) TBST (37536) |
Pierce Protein-Free Blocking Buffers | Non-protein blocking compound |
|
|
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
Protocols and technical guides
- General sandwich ELISA Protocol
- ELISA technical guide
- ELISA technical manual for Invitrogen ELISA Kits
- Biomarker quantitation assay guide
Blocking buffers
ELISA lysis buffers
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 |
|
|
Concentrate (DS98200) |
SuperBlock Blocking Buffer | Serum and biotin-free single purified glycoprotein |
|
|
PBS (37515) TBS (37535) PBST (37516) TBST (37536) |
Pierce Protein-Free Blocking Buffers | Non-protein blocking compound |
|
|
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
Protocols and technical guides
- General sandwich ELISA Protocol
- ELISA technical guide
- ELISA technical manual for Invitrogen ELISA Kits
- Biomarker quantitation assay guide
Blocking buffers
ELISA lysis buffers
Related products and resources
Sandwich ELISA protocol
Detailed protocol for colorimetric and chemiluminescent ELISA.
ELISA kits and antibody pairs
Quantitate single analytes with confidence with highly verified Invitrogen ELISA kits.
ELISA Instrumentation and Equipment
Plate washers, plate readers, pipettes.
Immunoassay guide
Immunoassay guide to protein quantitation.
ELISA Microplates and Plasticware
Hydrophobic, hydrophilic plates, strip plates.
ELISA substrates
Find TMB and other enzyme substrates for ELISA.
Bulk and custom quantities
Scale-up your process or product development pipeline with bulk quantities of high-quality immunoassay reagents.
For Research Use Only. Not for use in diagnostic procedures.