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Critical Reagents Usage in Bioanalytical Labs: Q+A with Dr. Santosh Shah

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Santosh Shah

Dr. Santosh Shah

Director, Biologics, Frontage Laboratories

Dr. Santosh Shah is the Subject Matter Expert and Director of Biologics, at Frontage Laboratories. He has been providing scientific leadership and has been contributing to the development of the Global Biologics Strategy. He works as a liaison between business development, operations, and various other departments. He is also involved with communicating, auditing, and presenting bioassay capabilities (PK, ADA, NAb, Biomarker) to the pharma sponsors. He has been leading the automation development team at Frontage.   Dr. Shah discovered several highly potent, copper-mediated, drug compounds against M. tuberculosis and S. aureus during his Postdoctoral Fellowship at the University of Alabama at Birmingham. During his Ph.D. at the University of Arizona, he dedicated himself to understanding the structure-function relationships of DNA nucleases.

Critical reagents for ligand binding assays (LBA) are essential components of bioanalytical testing. These include but are not limited to antibodies used as positive controls, capture, or detection reagents; peptides as drug targets; reference standards as test analytes for pharmacokinetics; positive controls for immunogenicity studies and matrices.

Critical reagents are crucial for assay performance and data comparability across different laboratories and countries since they directly impact the results of an assay. Thus, they need to be well characterized and validated for the proper execution of preclinical and clinical studies. Critical reagent stocks must also be maintained, and their sources recorded. Below is a snapshot of some of the key questions asked on the webinar, General Practices for Critical Reagents Usage in Bioanalytical Labs.

In the following Q+A, Dr. Santosh Shah, Director of Biologics, answers key questions on critical reagent usage in bioanalytical laboratories.

What approach would you use to bridge reference standards if you want to compare head-to-head, old, and new curves and full complement of quality controls (QC): LLOQ, LQC, MQC, HQC, ULOQ?

We set up two sets of plate controls (i.e., one set = one set of standards and two sets of QCs) each prepared with two different lots separately, and check the following results:

For each set, if standards and QCs meet the acceptance criteria.

If one lot of prepared QC meets the acceptance criteria in comparison to another lot of prepared standards.

Preparing from an old and new batch of reference standards will not fit on a 96-well plate, if there is the need to include three replicates of each. Is just running one or two replicates a valid approach so all five levels of QCs can be tested?

Our most common practice is to use two replicates. If three replicates are required, we use two plates to complete the comparison of reference standards.

Do you have a standard expiry assigned to different types of critical reagents, since often the stability is not known initially? Would you be willing to share your approach for being able to assign the expiry or retest/recertification dates initially?

For critical reagents acquired from external sources, we use expiry dates from certificates of analysis (COAs) provided by the sponsor/vendor. For critical reagents prepared in-house, we usually follow our standard operating procedure (SOP) based on the type of reagent, e.g., for an antibody-based critical reagent, we assign a 1-year expiry and retest annually.

How do you apply the correction factor, as you see >20% QC bias between using two different lots of reagents? In other words, will the correction factor be applied to assay modification or study data?

The correction factor is usually used when bridging sample analysis kits, in which standards and buffers are included for standard calibrators and QCs preparation. From kit to kit, the buffer and standard material are different to a certain extent and the overall comparison in QCs will be greater than individual standard material lots. The correction factor will be applied to the study data, not the assay.

Can you speak on extending expiration dates via retesting?

We perform one run with one set of standards and two sets of QCs (HQC, MQC, and LQC) loaded to plates in duplicate. If the run passes, the expiration may be extended for a time equal to the original time interval.

What is the purpose of BIO-006 Change Control Procedure?

BIO-006 Change Control Procedure SOP provides a procedure for managing document and equipment changes at Frontage Laboratories. It outlines the method of documenting and controlling changes to issued documents and equipment, as well as tracking changes.

What should the re-testing frequency for in-house conjugated reagents, e.g., biotinylated drug or digoxin drug, be?

We usually retest once a year since the labeled reagents tend to become aggregated, leading to a higher signal over time. This change is more detrimental to ADA studies in which the raw data are evaluated for positiveness and titer of the ADA confirmed. We would rather re-label reagents than re-testing the regents.

Frontage’s biologics teams have nearly 15 years of experience in complex drug development and large molecule applications throughout its evolution in product development. We have handled projects for peptides, proteins, monoclonal antibodies, bispecific antibodies, biosimilars, oligonucleotides, biomarkers, and antibody-drug conjugates. Contact our sales team for your biologics projects.

Note: This feature includes the webinar, panel discussion, and Q+A, and was done in association with Bioanalysis Zone.