Development of highly sensitive enzyme-linked immunosorbent assays for hepatocyte growth factor/scatter factor (HGF/SF): determination of HGF/SF in serum and urine from normal human subjects

A sandwich enzyme-linked immunosorbent assay (ELISA) using rabbit anti-hepa tocyte growth factor (HGF) IgG for human HGF, also known as the scatter fac tor, has previously been developed for determining increases in serum HGF l evels in various liver diseases. The sensitivity limit of the ELISA is, how ever, approximately 0.2 ng/ml sample, and HGF concentrations in about 50% o f normal subjects are not accurately measurable by this method, because the mean level of HGF in normal serum is close to the sensitivity limit. In th e present study, chicken Fab' from egg yolk anti-HGF immunoglobulin Y and r abbit Fab' from rabbit anti-HGF Ige were conjugated with beta -D-galactosid ase. With these conjugates as the second antibodies, we developed two sandw ich ELISAs for human HGF and found that the sensitivities were about 20 pg/ ml with the former conjugate and 2, pg/ml with the latter. The HGF concentr ation in sera from 138 normal subjects determined by the ELISA with the rab bit conjugate was 244+/-65 (SD) pg/ml serum, and it correlated very well wi th the number of leukocytes. Moreover, the ELISA with the rabbit conjugate permitted the determination of HGF levels in urine from normal subjects wit hout first concentrating the sample. The determination of HGF in various bi ological fluids other than blood and urine by these ELISAs may aid the diag nosis and prognosis of various diseases. (C) 2000 Elsevier Science B.V. All rights reserved.