Do immunoassays differentially detect different acidity glycoforms of FSH?

OBJECTIVE The possibility of the carbohydrate residues of glycoproteins aff ecting their recognition in immunoassays is an important and unresolved iss ue. This study looked for evidence of differential recognition of FSH glyco form preparations, of variable isoelectric point (pl) and known molarity, u sing three routine assays employing different antibody configurations. DESIGN Seven glycoform preparations with differing pi bands (between 3.8 an d 5.5) were produced by isoelectric focusing of recombinant human FSH and t he molecular weights determined by mass spectroscopy, Three concentrations of each glycoform were assayed and the results expressed relative to unfrac tionated material. From the relative responses, recognition differences bet ween the assay methods and between the glycoform preparations were investig ated. MEASUREMENTS Three routine assays were employed: the commercially available Amerlite(R) enzyme immunoassay and Delfia(R) immunofluorometric assay, tog ether with an in-house competitive two-site radioimmunoassay (RIA). RESULTS Overall, the three assays gave the same relative responses for equi valent glycoforms, with the only exceptions involving small differences bet ween some assay pairs for the fractions at the extremes of the pi range inv estigated. Within each assay type, differences (P <0.05) of up to 33% exist ed between glycoforms of different pl, however, these differences showed no patterns or trends across the entire acidity range examined. CONCLUSIONS Between the assay methods investigated in this study, few diffe rences exist in the recognition of individual pi bands of FSH when expresse d relative to a common unfractionated standard. Differences were apparent i n the recognition of the different acidity glycoforms within each assay met hod, however, these were small and unlikely to be of clinical significance.