The enzyme-linked immunosorbent assay (ELISA) is a frequently used application which determines the concentration of an analyte in solution. The ELISA method is straightforward and rapid. With automated equipment the assay is easily scalable for set up in high-throughput and can handle a large number of samples in parallel, making it a popular choice for the evaluation of research and diagnostic targets.

ELISA Protocols

General ELISA Protocols

Standard methods for different types of ELISA in 96-well microtiter plate format, including recommended buffers.

  1. Indirect ELISA
  2. Competition (Inhibition) ELISA
  3. Direct ELISA with streptavidin-biotin detection
  4. Sandwich ELISA with streptavidin-biotin detection
  5. Sandwich ELISA with direct detection

Protocols for use with Anti-Biotherapeutic Antibodies

Bio-Rad’s anti-biotherapeutic antibodies are ideal for the measurement of monoclonal antibody drugs and biosimilar products in PK and ADA assays.

We have developed a range of highly specific, recombinant anti-idiotypic antibodies with different binding modes and properties, enabling the development of bridging ELISA and antigen capture ELISA. Protocols are available for each type of PK assay and ADA assay for many of the anti-biotherapeutic products in our catalog.

PK and ADA Bridging ELISA Protocols

Bridging ELISA is a special case of a sandwich ELISA in which a dimeric or oligomeric antigen (most often an antibody in a sample) is detected by a capture and detection antibody. The antigen bridges the two specific antibodies. Bridging ELISAs are most frequently used for the detection of IgG, for example in pharmacokinetic (PK) or anti-drug antibody (ADA) assays.

PK Bridging ELISA measuring free drug protocols

PK Bridging ELISA measuring free drug

Type 1 anti-idiotypic antibodies are inhibitory and are used to detect free drug

  1. Adalimumab
  2. Bevacizumab
  3. Cetuximab
  4. Denosumab
  5. Eculizumab
  6. Etanercept
  7. Golimumab
  8. Infliximab
  9. Ipilimumab
  10. Natalizumab
  11. Nivolumab
  12. Omalizumab
  13. Palivizumab
  14. Panitumumab
  15. Pembrolizumab
  16. Rituximab
  17. Tocilizumab
  18. Trastuzumab (HCA166, HCA168)
  19. Trastuzumab  (HCA169, HCA176P)
  20. Trastuzumab (HCA169, HCA177P)
  21. Trastuzumab (HCA169, HCA270 IgG4)
  22. Ustekinumab  (HCA208, HCA210)
  23. Ustekinumab  (HCA209, HCA210)
  24. Vedolizumab 
PK Bridging ELISA measuring total drug protocols

PK Bridging ELISA measuring total drug

Type 2 anti-idiotypic antibodies are non-inhibitory and enable the measurement of total drug – free, partially and fully bound

  1. Cetuximab
  2. Etanercept
  3. Golimumab
  4. Infliximab
  5. Tocilizumab
ADA Bridging ELISA protocols

ADA Bridging ELISA

Type 1 anti-idiotypic antibodies are neutralizing and fully human; in immunoglobulin format they can be used as a control or calibrator as a surrogate for human ADAs

  1. Adalimumab (IgG1)
  2. Adalimumab (IgE)
  3. Bevacizumab
  4. Certolizumab pegol
  5. Cetuximab
  6. Denosumab
  7. Eculizumab
  8. Etanercept
  9. Infliximab
  10. Ipilimumab
  11. Natalizumab
  12. Nivolumab
  13. Omalizumab
  14. Palivizumab
  15. Panitumumab
  16. Pembrolizumab
  17. Ranibizumab
  18. Rituximab
  19. Tocilizumab
  20. Trastuzumab
  21. Ustekinumab
  22. Vedolizumab


Antigen Capture ELISA Protocols

A third type of antibody that only detects drug bound to its target antigen is available for some of our specificities.

Protocols: PK Antigen Capture ELISA measuring bound drug exclusively

PK Antigen Capture ELISA measuring bound drug exclusively

A Type 3 antibody is used in an antigen capture ELISA to detect bound drug exclusively and avoids the bridging ELISA format

  1. Adalimumab
  2. Golimumab
  3. Ipilimumab
  4. Omalizumab
  5. Ranibizumab HCA304, HCA306, HCA307
  6. Ranibizumab HCA316P
  7. Trastuzumab

Technical Support

Technical Support