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Solid-phase enzyme-immunoassay of anti-insulin antibodies: effect of labeling site in insulin and of labeled number of horseradish peroxidase on the assay sensitivity.

Three horseradish peroxidase (HRP)-labeled porcine insulins which have definite labeling site(s) were compared regarding sensitivity in a solid-phase enzyme-immunoassay (EIA) of anti-insulin antibodies. The standard curves obtained with LysB29-HRP-insulin and GlyA1-HRP-insulin were steeper than that with GlyA1,LysB29-diHRP-insulin for both polyclonal and monoclonal antibodies. Thus, the mono-HRP-labeled insulins can afford higher sensitivities in the EIA. The importance of the HRP-labeling site in insulin and of the number of labeled HRP was first demonstrated by using HRP-labeled insulins having definite labeling site(s).