Q&A Part 1: Industry Leaders Discuss Antibody Manufacturing

Antibody specificity to the desired protein or epitope in the stated application, testing in relevant positive and/or negative cells including primary models, application of the antibody in different immunoassays, and lot-to-lot consistency in performance.

Our approach is to focus on the needs of the researcher in order to provide highly specific, rigorously validated antibodies across a broad range of scientific research areas. That translates to the following key manufacturing considerations:

• Specificity—Ensure the performance of the antibody for the intended target. This is accomplished in part by including multiple experimental controls as well as panels of cell lines with known target expression. These results are corroborated using a variety of complementary, orthogonal assays to verify on- and off-target interactions.
• Fit for purpose/functionality—Validation of antibody performance in a particular application including immunohistochemistry, immunofluorescence, chromatin immunoprecipitation, flow cytometry, immunoprecipitation, and western blot.
• Transparency—Providing relevant information, including methods, working dilutions, and data, to support all claims of antibody performance to enable the customer to reproduce results.
• Consistency—Compare each new lot side-by-side to ensure lot-to-lot performs identically to previous validations for a particular target or application. This is especially necessary for the development of polyclonal antibodies. Transparency in revealing this information by any manufacturer is paramount.

The most important factor to consider when making an antibody is how the antibody is intended to be used. Knowledgeable antibody producers can affect the performance of antibodies in various immunoassays by selecting the appropriate immunogen, varying the immunization protocol, and/or selecting the appropriate screening strategy to ultimately yield an antibody that is fit-for-purpose. For instance, a peptide or denatured immunogen may result in an antibody functional for conventional Western blotting where the target protein is itself denatured, eliminating higher order protein structures that exist in native proteins. In contrast, a native immunogen is preferred if the immunoassay is intended to detect native targets, such as capture ELISA or fresh-frozen section immunohistochemistry.

The keys to developing a great antibody product are in designing appropriate immunogens, screening for clones in relevant systems, and thorough validation of the clones and materials they produce. This is where the vast majority of effort must be spent. The failure to invest resources in this portion of product development is a key reason some companies fail to achieve a reputation for quality, despite having produced a large collection of antibodies.

The antibody product must be highly specific, highly sensitive, and highly reproducible in all intended applications. The provider must take care that all these criteria are met. In addition, the vendor should describe exactly how the product is to be used. An antibody product is not per se highly specific and highly sensitive, but only in the context of proper use.

When making a new antibody or manufacturing a new lot there are four important focus areas that need to be addressed by the manufacturer. These focus areas are high specificity, sensitivity, reproducibility, and application specificity.