Differential determination of recombinant human interleukin-1α and its deamidated derivative by two sandwich enzyme immunoassays using monoclonal antibodies: Comparison with a polyclonal antibody-based competitive enzyme immunoassay

Using three different monoclonal antibodies (McAb no. 374, no. 964 and no. 1190) to human interleukin-1α (rHu-IL-1α), we have established two sandwich enzyme immunoassays (EIA) to differentiate rHu-IL-1α and its deamidated derivative (rHu-Asp 36-IL-1α) where the asparagine at position 36 (counting from the N-terminus) of rHu-IL-1α is converted to Asp. The McAb no. 1190 reacts specifically with rHu-IL-α and not with the rHu-Asp 36-IL-1α whereas both no. 374 and no. 964 can react with the two different forms of rHu-IL-1α. The first EIA (S-EIA I) which uses the McAb no. 964 labelled with horse-radish peroxidase and the McAb no. 1190 fixed to the microtiter plate, only measure rHu-IL-1α. The second EIA (S-EIA II) which uses enzyme labelled no. 964 and no. 374 fixed to the plate, can detect both rHu-IL-1α and rHu-Asp 36-IL-1α and this assay of total rHu-IL-1α is comparable to a competitive EIA using an enzyme-labelled rHu-IL-1α and an anti-rHu-IL-1α polyclonal antibody. Thus, the level of rHu-Asp 36-IL-1α in the samples containing the two IL-1αs can be calculated by subtracting the level measured by S-EIA I from that measured by S-EIA II. The two EIA systems with an assay range of 1.5–100 ng/ml do not recognize IL-1β, IL-2, rHu-TNFα, IFN-α and IFN-γ of human origin. The assays may be suitable for quality control, stability testing and serokinetic investigations of rHu-IL-1α.