Ligand binding assays (LBAs) are commonly used in the pharmaceutical industry to determine the potency and specificity of drug candidates. These assays rely on the interaction between a ligand and its receptor, often antibodies or antibody pairs. However, the accuracy and reproducibility of LBAs can be compromised by suboptimal development of critical reagents, which are components of the assay that are essential for its performance.
Critical reagents include antibodies, proteins, and other biomolecules, labelled or unlabelled, that are used to detect or capture the analyte. They can be the source of several issues in LBAs, including lot-to-lot variability, poor stability, and non-specific binding. These issues can result in inaccurate or unreliable assay results, which can have serious consequences for drug development and patient safety.
In a recent survey by Bioanalysis Zone, 74% of respondents reported critical reagent development to be the main limitation of ligand binding assays, with 52% reporting critical reagent consistency as a major limitation.
One especially common problem with critical reagents is lot-to-lot variability. This can occur when different batches of a reagent have different properties, such as binding affinity or specificity, or if the labelling conditions used result in inconsistent levels of incorporation. If a reagent with different properties is used in an assay, it can lead to inconsistent or unreliable results. To mitigate this problem, it is important to ensure that not only are the raw materials used to produce the reagent are consistent, but that the bioconjugation and purification processes involved are sufficiently robust to day-to-day variation. All too frequently, reagents developed using off-the-shelf labelling kits fail to meet these criteria.
Whilst use of such kits may give a conjugatesuitable for early assay development work, their lack of control and flexibility means that the assay performance obtained is likely to be far from optimal. By controlling the labelling conditions through careful conjugate development, another important handle is available for optimising the assay.
Typically issues also arise when the initial conjugate batch becomes depleted and a replacement critical reagent lot is required. At that point, unless the bioconjugation process is completely understood and controlled, variable and inconsistent reagents will be produced which then impact assay performance. As regulator demands for greater levels of sensitivity and precision increase, having all the tools necessary to get peak performance from the assay becomes invaluable.
Fleet Bioprocessing Ltd. are well-known worldwide for our expertise in bioconjugation chemistry, smoothly coupling molecules of interest to create powerful new entities such as antibody-enzyme conjugates. Our experts have worked at the cutting edge of this field since the 1980s, and our Managing Director is co-author of Bioconjugation: Protein Coupling Techniques for the Biomedical Sciences (Macmillan, 1998), a standard reference work in the field.
Within the contract R&D sphere, Fleet have unrivalled experience and knowhow in bioconjugate development. We pride ourselves on the reproducibility of our conjugation methods - we routinely employ highly controllable heterobifunctional coupling chemistry allowing the greatest flexibility in achieving your product requirements. Typically we will provide your conjugate as a highly-purified product, in a buffer of your choice, with detailed incorporation information.
Batch after batch (for more than 20 years in some cases!), conjugates developed and manufactured by Fleet have reliably given good performance in immunoassays used all over the world. We would be happy to talk to you about your LBA critical reagent needs – and hopefully help you to avoid some of the issues highlighted in the survey.