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-hepatocyte growth factor (HGF) IgG for human HGF, also known as the scatter factor, has previously been developed for determining increases in serum HGF levels in various liver diseases. The sensitivity limit of the ELISA is, however, approximately 0.2 ng/ml sample, and HGF concentrations in about 50% of 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 the present study, chicken Fab' from egg yolk anti-HGF immunoglobulin Y and rabbit Fab' from rabbit anti-HGF IgG were conjugated with beta-D-galactosidase. With these conjugates as the second antibodies, we developed two sandwich 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 concentration in sera from 138 normal subjects determined by the ELISA with the rabbit conjugate was 244+/-65 (SD) pg/ml serum, and it correlated very well with the number of leukocytes. Moreover, the ELISA with the rabbit conjugate permitted the determination of HGF levels in urine from normal subjects without first concentrating the sample. The determination of HGF in various biological fluids other than blood and urine by these ELISAs may aid the diagnosis and prognosis of various diseases.
Ohnishi
T
T
Department of Biochemistry, Kagoshima University Dental School, 35-1 Sakuragaoka-8, 890-8544, Kagoshima, Japan.
Kakimoto
K
K
Hashida
S
S
Fujii
M
M
Hirono
S
S
Nishiyama
K
K
Amita
Y
Y
Ishikawa
E
E
Tsubouchi
H
H
Daikuhara
Y
Y
D000328:Adult; D000818:Animals; D002645:Chickens; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D017228:Hepatocyte Growth Factor / Q000097:blood* / Q000652:urine*; D006801:Humans; D018796:Immunoconjugates; D007128:Immunoglobulin Fragments; D007136:Immunoglobulins; D008297:Male; D008875:Middle Aged; D011817:Rabbits; D012016:Reference Values; D012680:Sensitivity and Specificity; D001616:beta-Galactosidase
C016512:IgY; D018796:Immunoconjugates; D007128:Immunoglobulin Fragments; D007136:Immunoglobulins; D017228:Hepatocyte Growth Factor; D001616:beta-Galactosidase