Novel and sensitive enzyme immunoassay (immune-complex-transfer enzyme immunoassay) for alpha-fetoprotein from hepatocellular carcinoma.
Alpha-fetoprotein in sera from patients with hepatocellular carcinoma was fractionated into three peaks by affinity chromatography on a column of Lens culinaris agglutinin-Sepharose 4B. One peak (the first peak), which passed through the column without adsorption, was found in both healthy subjects and patients with liver cirrhosis and hepatocellular carcinoma. The second and third peaks were reactive with L. culinaris agglutinin and found only in patients with hepatocellular carcinoma. For alpha-fetoprotein in the second and third peaks, a novel and sensitive enzyme immunoassay (immune-complex-transfer enzyme immunoassay) was developed. Alpha-fetoprotein in test serum was reacted with dinitrophenyl affinity-purified anti-alpha-fetoprotein IgG, and the complex formed was trapped onto affinity-purified (antidinitrophenyl bovine serum albumin) IgG-coated polystyrene balls. The polystyrene balls were washed to eliminate substance(s) other than alpha-fetoprotein in the test serum, and the complex was eluted from the polystyrene balls with dinitrophenyl-L-lysine. The eluted complex containing alpha-fetoprotein in the second and third peaks was trapped onto L. culinaris agglutinin-coated polystyrene balls and reacted with affinity-purified anti-alpha-fetoprotein Fab'-beta-D-galactosidase conjugate. Beta-D-galactosidase activity bound to the polystyrene balls was assayed by fluorimetry. The maximal volume of serum that could be used without interference was 20 microliters, which was 100-fold larger than that in the previous enzyme immunoassay.(ABSTRACT TRUNCATED AT 250 WORDS)
Kohno
T
T
Department of Biochemistry, Medical College of Miyazaki, Japan.
Kitamura
T
T
Tsuda
K
K
Ishikawa
E
E
D014408:Biomarkers, Tumor; D006528:Carcinoma, Hepatocellular / Q000032:analysis* / Q000276:immunology; D002846:Chromatography, Affinity; D006801:Humans; D007124:Immunoenzyme Techniques; D008103:Liver Cirrhosis / Q000175:diagnosis; D008113:Liver Neoplasms / Q000032:analysis* / Q000276:immunology; D000509:alpha-Fetoproteins / Q000032:analysis*; D001616:beta-Galactosidase
D014408:Biomarkers, Tumor; D000509:alpha-Fetoproteins; D001616:beta-Galactosidase