Comparison of two direct neutralizing assay formats using recombinant follicle-stimulating hormone as agonist.

Characterizing anti-drug antibodies for neutralizing activity is commonly part of the immunogenicity testing package for most therapeutic proteins. Cell-based neutralization assays can generally be categorized as direct- or indirect assays depending on whether they are associated with therapeutics with agonistic- or antagonistic properties. This paper's aim is a comparison of the two direct neutralization assay formats; the variable- and fixed concentration assay format, using recombinant follicle-stimulating hormone as drug agonist. Essential validation- and performance parameters, such as sample through-put, cut-point, precision, sensitivity and drug tolerance, were compared. The fixed concentration assay format offers superior sample through-put (40 versus 6 samples), precision (coefficient of variation of ≤14% versus 34%) and almost 6 times better sensitivity and is generally recommended as the better option particularly for quasi-quantitative assessments of neutralizing antibodies.