Production and Characterization of Monoclonal Antibodies Against 17α-Hydroxyprogesterone

Abstract

Hybridomas secreting monoclonal antibodies (MAbs) against 17alpha-hydroxyprogesterone (17OHP) have been generated. These MAbs are highly specific and have an affinity of 7-12 x 10(7) M(1). The hybridomas were obtained by fusion of spleen cells from immunized mice with mouse myeloma P3X63 Ag8.653 cells. The antigen used for immunization was 17OHP conjugated to bovine serum albumin (17OHP:BSA). Fused cells were plated and cloned in 96-well microtiter plates. Wells containing hybridomas were screened simultaneously for specific gamma globulin (IgG) and anti-17OHP activity using an enzyme-linked immunosorbent assay (ELISA)-based method, which is faster than the conventional radioimmunoassay (RIA) screening procedure. Limiting dilution methods were used to obtain single hybridoma clones producing MAb. The stable hybridomas secreting anti-17OHP MAbs were expanded into bioreactors or ascites fluid for large-scale production of the required antibodies. These MAbs will be used in the formulation of a 17OHP assay kit to screen for congenital adrenal hyperplasia (CAH) in local newborn human population.

Production and Characterization of Monoclonal Antibody Against Human Serum Albumin

“…Antibody-based immunoassays are the most commonly used type of diagnostic assay and still one of the fastest growing technology for the analysis of biomolecules. (14)(15)(16)(17)(18)(19) High-affinity MAbs specific for the protein HSA are a useful system for the study of antigen antibody interaction, and its clinical significance has been improved because of two reasons. First, as mentioned before, MAU is one of the earliest markers of renal disease and its progression in diabetes, cardiovascular disease and hypertension as well as in the general population.…”