Production of monoclonal antibodies against interleukin-1 alpha and -1 beta. Development of two enzyme immunometric assays (EIA) using acetylcholinesterase and their application to biological media.
We describe two series of monoclonal antibodies (mAbs) directed against human interleukin-1 alpha (36 mAbs) and -1 beta (11 mAbs). The binding compatibility of each of mAb was studied using biotin-labelled mAbs in immunometric tests. Among the different pairs of compatible mAbs, we selected one pair for each interleukin-1 (IL-1) with optimal properties for a two-site immunometric assay. In these assays, covalent conjugates of mAb coupled to the tetrameric form of acetylcholinesterase (mAb-AChE) were used as tracers. The tests were performed in 96-well microtiter plates coated with the complementary mAb. Both assays appeared sensitive and specific since minimum detectable concentrations as low as 1 pg/ml were determined for each IL-1 without any significant cross-reactivity (less than 0.01%). The intra-assay precision was also very good with a coefficient of variation of less than 10% over a wide range (between 3 and 500 pg/ml depending on the time devoted to the enzymatic reaction). The high sensitivity and precision of the assays can be ascribed to the high affinities of the mAbs as well as the optimal catalytic properties of AChE. The specificity of the determination performed in culture medium was demonstrated using different validation tests including a comparison with a bioassay and the fractionation of samples by molecular sieve chromatography. Evidence is presented that the assay could be used for the determination of IL-1 levels in biological media such as plasma or serum.
Grassi
J
J
Section de Pharmacologie et d'Immunologie, CEN-SACLAY, Gif-sur-Yvette, France.
Frobert
Y
Y
Pradelles
P
P
Chercuitte
F
F
Gruaz
D
D
Dayer
J M
JM
Poubelle
P E
PE
D000110:Acetylcholinesterase; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000918:Antibody Specificity; D001667:Binding, Competitive; D003470:Culture Media; D004306:Dose-Response Relationship, Immunologic; D006801:Humans; D007124:Immunoenzyme Techniques; D007375:Interleukin-1 / Q000032:analysis / Q000276:immunology*; D011994:Recombinant Proteins / Q000276:immunology
D000911:Antibodies, Monoclonal; D003470:Culture Media; D007375:Interleukin-1; D011994:Recombinant Proteins; D000110:Acetylcholinesterase