Q&A Part 3: Ensuring Antibody Quality

Our antibodies are screened by various quality control assays at multiple points of the manufacturing process. Each time antiserum is collected for a polyclonal antibody, it is tested for antibody performance by multiple immunoassays, such as ELISA and Western blot, and must meet tight acceptance criteria. For qualification of a new antibody, we run controlled assays using overexpressed or recombinant proteins, as well as established positive and negative cell lines for the target protein. This allows us to demonstrate the sensitivity and specificity of the antibody. For all antibodies, testing is conducted at every step of the manufacturing process, including after purification, conjugation, and again before each lot is released. Rockland has strict lot-to-lot release criteria to ensure that the customer obtains the same product with each lot. Rockland takes pride in following this rigorous testing schedule and having full control over the manufacturing process.

BD carefully selects and characterizes antibody content in product development and tests in relevant primary model systems to ensure biological accuracy.

BioLegend manufactures antibody products at our main facility in San Diego, where we are certified to operate under ISO 13485:2003 standards. Our Quality Management System ensures traceability, reproducibility, and continuous improvements in manufacturing.

It is very important to us that every product that leaves our facility performs exactly as promised. This starts with validation of all new antibodies against endogenous targets in the applications that will be most relevant to customers. The confirmation of specificity and lack of off-target binding is confirmed with appropriate cellular controls, comparison with any pre-existing antibody clones, and appropriate performance in multiple relevant assays. Subsequent manufactured lots of any given antibody are characterized through several biochemical and functional QC tests.

Ensuring the quality of our products involves multiple, independent teams in the product development, release, and support process. These scientist-led teams independently test and validate the antibody only when predetermined milestones have been achieved. This process begins at the time of target selection: teams individually review the proposed project, establish testing strategies and determining biologically relevant models for testing, as well as acceptance criteria for each application desired for the antibody product’s use. By following this strategy, no CST antibody product is released without the approval of a development scientist, an application scientist for each approved application, and a group leader who independently reviews all of the data. Then, before the product is available in our catalog, each product must go through an additional round of verification by our strongest critics—the production scientists. They represent the teams that provide technical support, regular quality control testing, and development of all subsequent lots of the same product. If there is any reason to believe that a product cannot be supported both in the short- and long-term, it will not be released. While this extremely cautious approach may result in failed projects, it ensures that only the highest quality, most rigorously validated, consistently produced products make it out our door.

At Miltenyi Biotec we manufacture monoclonal antibodies mainly for cell isolation and flow cytometry applications. This means that we directly conjugate antibodies to magnetic beads or fluorochromes. When we develop a new antibody product for our portfolio, we rigorously evaluate its specificity and sensitivity in all target applications. Furthermore, we make sure that our manufacturing processes are highly consistent and that our quality control procedures ensure all critical quality attributes.

Once the antibody is made it undergoes a multi-application validation process. We have an extensive tissue bank and blot library that allow us to test each antibody in a variety of applications to determine the precise specificity of the antibody. We not only concentrate on the detection of protein in a sample, but also its sub-cellular localization. In addition, we test our antibodies in several applications to assess specificity for the target and provide contextually relevant validation in conditions most likely used by researchers. Some of our antibodies also undergo enhanced validation, which involves the use of multiple antibodies for the same target, parallel knockout or knockdown techniques, and the use of orthogonal techniques whereby immunodetection results are compared with RNA-seq results to obtain an antibody-independent method for measuring the gene or protein of interest.