Purpose: Streptavidin-coated plates are a widely-used tool for use in biomarker measurements including traditional sandwich immunoassays, bridging immunogenicity assays, and pharmacokinetic (PK) assays. To assure reproducible results for long-term studies, it is essential that streptavidin plates are rigorously characterized for consistent performance and stability. MSD offers streptavidin-coated plates in two different 96-well formats, MSD GOLD Streptavidin and MSD GOLD Small Spot Streptavidin. Here we review the quality control performance of both plate formats spanning multiple years, as well as the results of a 4°C stability study for the MSD GOLD Streptavidin plate format. We also demonstrate the utility of the MSD GOLD Small Spot Streptavidin plate as a tool in the early development phase of a pharmacokinetic (PK) assay for the biopharmaceutical product Adalimumab.
Methods: Precision, accuracy, binding capacity, and uniformity were evaluated for 96-well MSD GOLD Streptavidin and MSD GOLD Small Spot Streptavidin plates. Quality control tests used Biotin and SULFO-TAG™ label conjugated to IgG (BTI) as a surrogate for an assay using a biotinylated capture reagent. Intra-plate reproducibility was measured by testing the entire plate with a constant amount of BTI near the plate’s binding capacity (0.2 pmole of IgG) and inter-lot reproducibility was determined from BTI titration measurements across multiple plate lots. Real time stability was determined by conducting quality control tests at regular intervals over an extended period of storage time: 57+ months at 2-8°C. For the PK application, multiple antibody pairs were screened by testing each candidate antibody as both a capture and detection reagent with calibrator curves and samples.
Results: MSD GOLD Streptavidin: For uniformity testing, the intra-plate %CV for 407 plate lots was below 6%, with an average intra-plate %CV of less than 3% (14,917 plates tested). For titration testing, the inter-lot %CVs were less than 10% and all plates showed consistent binding capacity and signal performance across lots. In stability studies, BTI signal at several levels across the assay range remained constant, indicating equivalent performance through the duration of the 2-8°C storage condition. MSD GOLD Small Spot Streptavidin: For uniformity testing, the intra-plate %CV for 51 plate lots was below 6%, with an average intra-plate %CV of less than 3% (1,919 plates tested). For the PK application, 64 potential antibody pairs were pared down to 3 pairs by selecting top performers for signal intensity, sensitivity, and dilution linearity.
Conclusion: The reproducibility of the MSD GOLD plate format, shown both across and within lots, highlights its utility as a key component in developing robust and reliable assay applications. Both the flexibility and stability of the format makes it an ideal development tool, as demonstrated by the PK application for Adalimumab.
Seth B. Harkins– Rockville, Maryland
Angelina Anderson– Rockville, Maryland
John Joern– Rockville, Maryland
Samarth Chugh– Rockville, Maryland
Jon Buhrman– Rockville, Maryland
Laure Moller– Rockville, Maryland
Brunah Otieno– Rockville, Maryland
David Stewart– Meso Scale Discovery, Rockville, Maryland
Jacob Wohlstadter– Rockville, Maryland
Seth Harkins– Assoc. Director, Meso Scale Discovery