I don't think using a biotinylated antigen to coat the plate will be a problem as far as the antigen binding to the plate. The potential issue I see is that the biotinylation may interfere with epitope recognition by the antibodies in the sera if the biotin is attached in or near the epitope.

If your antigens are small (i.e. peptides or small molecules), don't coat them directly, as most of them will adsorb to the surface of the plate and be quite inaccessible for antibodies, and, in addition, the coating efficiency usually will be very heterogenous from well to well.

Better coat your plates with streptavidin first and attach the antigens in a limiting way (not saturating the plate) via their biotin label. Of course, there is a residual risk, that some antibodies in the sera you're testing won't recognize the antigen, as Adam B Shapiro has mentioned. Depending on the structure of your antigens, biotinylation usually is not uniform (except there is only one lysine present, when using the conventional labeling approach), so just go ahead.

Thanks for the answers! We decided indeed not to use the antigen.

hi

it is also possible to use a Bead-Based ELISA for the Detection of SARS-CoV-2 Antibodies using the magnetic Dynabeads. The beads are pre-conjugated with trimeric spike protein (either as a stand-alone bead or as part of an ELISA kit with all buffers and ctrls). The use of a bead based approach ensures a very rapid protocol - only 10 min incubation with the target (in serum or plasma) is required followed by 15 min with HRP conjugated Abs and 10 min with the substrate with a few washing steps in between (45 min in total). As the beads are magnetic, the protocol can be automated on the KingFisher platform (scripts on Apex or Flex) for a hands off solution.

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ketil