Differential determination of recombinant human interleukin-1 alpha 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-1 alpha (rHu-IL-1 alpha), we have established two sandwich enzyme immunoassays (EIA) to differentiate rHu-IL-1 alpha and its deamidated derivative (rHu-Asp36-IL-1 alpha) where the asparagine at position 36 (counting from the N-terminus) of rHu-IL-1 alpha is converted to Asp. The McAb no. 1190 reacts specifically with rHu-IL-alpha and not with the rHu-Asp36-IL-1 alpha whereas both no. 374 and no. 964 can react with the two different forms of rHu-IL-1 alpha. 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-1 alpha. The second EIA (S-EIA II) which uses enzyme labelled no. 964 and no. 374 fixed to the plate, can detect both rHu-IL-1 alpha and rHu-Asp36-IL-1 alpha and this assay of total rHu-IL-1 alpha is comparable to a competitive EIA using an enzyme-labelled rHu-IL-1 alpha and an anti-rHu-IL-1 alpha polyclonal antibody. Thus, the level of rHu-Asp36-IL-1 alpha in the samples containing the two IL-1 alpha s 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-1 beta, IL-2, rHu-TNF alpha, IFN-alpha and IFN-gamma of human origin.(ABSTRACT TRUNCATED AT 250 WORDS)
Sunahara
N
N
Research Laboratories, Dainippon Pharmaceutical Co., Osaka, Japan.
Kawata
S
S
Kaibe
K
K
Furuta
R
R
Yamayoshi
M
M
Yamada
M
M
Kurooka
S
S
D000818:Animals; D000911:Antibodies, Monoclonal / Q000145:classification; D001224:Aspartic Acid / Q000378:metabolism; D001667:Binding, Competitive; D004355:Drug Stability; D005260:Female; D006801:Humans; D007106:Immune Sera; D007124:Immunoenzyme Techniques / Q000592:standards; D007375:Interleukin-1 / Q000031:analogs & derivatives / Q000032:analysis* / Q000493:pharmacokinetics / Q000592:standards; D007700:Kinetics; D051379:Mice; D008807:Mice, Inbred BALB C; D008809:Mice, Inbred C3H; D011786:Quality Control; D011817:Rabbits; D011994:Recombinant Proteins / Q000031:analogs & derivatives / Q000032:analysis* / Q000592:standards
D000911:Antibodies, Monoclonal; D007106:Immune Sera; D007375:Interleukin-1; D011994:Recombinant Proteins; D001224:Aspartic Acid