Development and characterization of anti-PMSG and its effect on superovulation in mice

In the present investigation, efforts have been made to raise antibodies (antisera) against commercial pregnant mare serum gonadotrophin (PMSG). The antibodies (raised in rabbit) were characterised by polyacrylamide gel electrophoresis (PAGE) and immunodiffusion (ID) and were tested on mice. The PMSG was mixed with Freund's complete adjuvant (FCA, Difco, USA) in the ratio of 2:1. On 0, 7 and 15 day the rabbit was immunized with the PMSG + FCA suspension, containing 220, 220 and 440 mu g of protein, respectively followed by an intravenous injection (560 mu g protein) on day 21. Blood serum harvested before (0 day) as well as after (28 days) immunization was tested for a total protein, immunoglobulins (Ig) and antibody titre. The presence of a high molecular weight protein (anti-PMSG) in the antiserum (partially purified) was comfirmed by the presence of a single precipitation line in the ID plates. PAGE of PMSG indicated that this hormone/product is a combination of six polypeptides of 185, 160, 130, 70, 30 and < 10 kDa. The comparison of PAGE patterns of normal serum/normal Ig and antiserum/specific Ig indicated that polypeptides of > 240, 225, 180 and < 10 kDa are produced in the anti-PMSG as a result of immunization. While investigating the effect of anti-PMSG on superovulation with PMSG in the mice, it was observed that 100 mu l (10 i.u.) of 20 times diluted antiserum was an effective dose to neutralize the effect of residual PMSG.