CHARACTERIZATION OF MONOCLONAL-ANTIBODIES AGAINST HUMAN INTERLEUKIN-10 AND THEIR USE IN AN ELISA FOR THE MEASUREMENT OF THIS CYTOKINE

Biological and biochemical characteristics of monoclonal antibodies (M ABs) raised against human interleukin-10 (IL-10) are described as well as their use in the design of a specific ELISA for the measurement of the cytokine. 21 murine anti-human interleukin-10 (IL-10) MABs were o btained by fusion of splenocytes from mice immunized against human rec ombinant IL-10 with SP2/0 myelomatous cells. These antibodies define t hree major antigenic areas on the IL-10 molecule, one of which compris es epitopes involved in receptor binding and induction of biological a ctivity. They recognize recombinant human IL-10 with affinities rangin g from 1.3 x 10(-7) to 3 x 10(-11), as well as natural IL-10. Most of them also recognize viral IL-10 (vIL-10) encoded by the Epstein-Barr v irus (EBV). A specific human-IL-10 ELISA has been developed using two MABs (18 and 19) as capture antibody and one MAB (17) as detector. The sensitivity (3 pg/ml), precision (intra-assays < 4%), reproducibility (interassay < 3%), and accuracy (recoveries, ranging between 84 and 1 07%, in several fluids) of the assay, plus its excellent performance i n dilution tests, and the lack of interference when in the presence of possible cross-reactive substances, permits accurate cytokine measure ment in biological fluids such as serum, plasma, bronchoalveolar lavag e, urine and culture supernatants. Using the assay, IL-10 was measurab le in the plasma of patients with septic shock (range 11-2740 pg/ml) w hereas IL-10 plasma levels were < 7.8 pg/ml in healthy volunteers.