20-alpha-Hydroxysteroid dehydrogenase from pseudopregnant rat ovary: obtention and characterization of a monoclonal antibody against the enzyme activity.
The enzyme 20-alpha-hydroxysteroid dehydrogenase (20-alpha-HSD) was purified from pseudopregnant rat ovaries and used as antigen for the development of a monoclonal antibody by the hybridoma technique. Spleen cells of BALB/c mice immunized with purified 20-alpha-HSD were fused with SP2/0 mouse myeloma cells. Among the colonies of hybrid cells, one (designated mAb-HSD 11) was found to be secreting antibodies (IgM) able to inhibit 20-alpha-HSD activity. The antibody-secreting hybridome was amplified by ascitic fluid production and the monoclonal antibody purified by Bakerbond ABx procedure. Purified mAb-HSD 11 was able to inhibit 20-alpha-HSD activity in a dose-dependent manner. Studies of Michaelis constants of 20-alpha-HSD indicate that this monoclonal antibody increases the Km for 20-alpha-dihydroprogesterone and decreases the Vmax.
De La Llosa-Hermier
M P
MP
Laboratoire de Biochimie des Hormones, CNRS, Gif-sur-Yvette, France.
Nocart
M
M
Paly
J
J
Hermier
C
C
D015089:20-Hydroxysteroid Dehydrogenases / Q000037:antagonists & inhibitors / Q000276:immunology* / Q000302:isolation & purification; D043607:20-alpha-Hydroxysteroid Dehydrogenase; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology* / Q000302:isolation & purification; D005260:Female; D007075:Immunoglobulin M / Q000276:immunology; D007700:Kinetics; D051379:Mice; D008807:Mice, Inbred BALB C; D010053:Ovary / Q000201:enzymology*; D011555:Pseudopregnancy / Q000201:enzymology; D051381:Rats; D017208:Rats, Wistar
D000911:Antibodies, Monoclonal; D007075:Immunoglobulin M; D015089:20-Hydroxysteroid Dehydrogenases; D043607:20-alpha-Hydroxysteroid Dehydrogenase