A SANDWICH ENZYME-IMMUNOASSAY FOR THE DETERMINATION OF NEUTROPHIL LIPOCALIN IN BODY-FLUIDS

Human neutrophil lipocalin was purified from human buffycoat. A polyclonal antibody was obtained by immunisation of rabbits. The antibody reacted with the free lipocalin as well as with the PMNL-gelatinase bound protein. This antibody was used to establish a sensitive sandwich-ELISA for the determination of the protein in body fluids using the biotin/streptavidin system. The mean intra-assay C.V. was 2.3% and the mean inter-assay C.V. 6.7%. The recovery in human plasma was determined to be 98.8%. The ELISA allowed the determination of the protein in the concentration range 0.2-25 mu g/l. Measurement of the neutrophil lipocalin concentration showed that human plasma of healthy donors contained 97 +/- 81 mu g/l (n = 122) and that the concentrations in serum were significantly higher (P < 0,001) with 133 +/- 90 mu g/l (n = 122). Neutrophil lipocalin was also found in the urine of healthy donors (8.1 mu g/l; n = 9). Very high concentrations of this lipocalin were found in the synovial fluids of patients suffering from inflammatory rheumatoid arthritis (1.7 +/- 1.4 mg/l; n = 37).