The induction and characterization of antiserum against human placenta 17-beta-hydroxysteroid dehydrogenase.
17-beta-Hydroxysteroid dehydrogenase was purified from human placenta. The purification process included the following steps: 50% saturated ammonium sulfate precipitation; heat treatment; affinity column chromatography; and preparative SDS gel electrophoretic elution. This enzyme was also characterized by HPLC gel filtration and two dimensional gel electrophoresis with isoelectrofocusing. The results indicate that the molecular weight of these enzymes in their native condition is around 68 kDa and 34 kDa in SDS PAGE. Therefore, the active form of the enzyme is an identical dimmer in nature. There are three charged isomers as demonstrated by isoelectrofocusing. The antiserum against the 17-beta-HSD was induced by injection of purified human placenta 17-beta-HSD in rabbits. The antiserum was collected and characterized by ELISA, immunohistochemistry, immunoblot and immunoprecipitation tests. The results showed that the antibody titer was over 1:12,800, and specificity against human placenta 17-beta-HSD was also observed.
Chen
J Y
JY
Department of Biology and Anatomy, National Defense Medical Center, Taipei, Taiwan, Republic of China.
Chang
W C
WC
Chao
C F
CF
D015067:17-Hydroxysteroid Dehydrogenases / Q000276:immunology* / Q000302:isolation & purification; D000818:Animals; D000918:Antibody Specificity; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005260:Female; D006801:Humans; D007106:Immune Sera / Q000276:immunology*; D015151:Immunoblotting; D007150:Immunohistochemistry; D007163:Immunosorbent Techniques; D010920:Placenta / Q000201:enzymology*; D011817:Rabbits; D014327:Trophoblasts / Q000201:enzymology
D007106:Immune Sera; D015067:17-Hydroxysteroid Dehydrogenases