Purification and properties of a monoclonal antibody specific to the free beta-subunit of human chorionic gonadotropin (hCG beta) and its use in the isolation of free hCG beta produced by choriocarcinoma cells.
The free beta-subunit of hCG is secreted by several tumors and is reported to be different from native hCG beta. We have developed a monoclonal antibody, B158, specific for free hCG beta to facilitate the detection and isolation of tumor-derived free hCG beta in the presence of intact hCG and hCG alpha. B158 belongs to the immunoglobulin G1 subclass, has high affinity for hCG beta (Ka, 1.05 X 10(9) M-1), and can be obtained in large quantities. The sensitivity of this antibody to detect free hCG beta in a RIA is less than 1 ng. B158 has negligible cross-reactivity with hCG, human LH, and other intact glycoprotein hormones and reacts with the free beta-subunits of deglycosylated hCG, human LH, and ovine LH. The antibody completely inhibits the recombination of hCG beta with hCG alpha indicating the antigenic site to be in the subunit interaction region or its vicinity. Comparison of the amino acid sequences of the various cross-reacting and noncross-reacting hormones indicates that the antigenic site may be discontinuous and conformational. B158 was purified to homogeneity from ascites fluid by DEAE-Affi-Gel blue and hCG beta affinity chromatography. Immobilized pure B158 antibody was used to isolate free hCG beta in a homogeneous form and high yield from BeWo choriocarcinoma cell culture supernatants. This free hCG beta has a slightly higher mol wt than standard hCG beta and recombines normally with hCG alpha. BeWo cells appear to produce only one species of free hCG beta.
D000818:Animals; D000911:Antibodies, Monoclonal / Q000302:isolation & purification*; D002822:Choriocarcinoma / Q000032:analysis*; D006063:Chorionic Gonadotropin / Q000276:immunology* / Q000302:isolation & purification; D018997:Chorionic Gonadotropin, beta Subunit, Human; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D051379:Mice; D008807:Mice, Inbred BALB C; D010446:Peptide Fragments / Q000276:immunology* / Q000302:isolation & purification; D011247:Pregnancy; D011863:Radioimmunoassay; D014594:Uterine Neoplasms / Q000032:analysis*
D000911:Antibodies, Monoclonal; D006063:Chorionic Gonadotropin; D018997:Chorionic Gonadotropin, beta Subunit, Human; D010446:Peptide Fragments