CHARACTERIZATION OF MOUSE-HAMSTER-HUMAN CROSS-REACTING ANTIOOCYTE MONOCLONAL-ANTIBODIES PRODUCED BY INTRASPLENIC IMMUNIZATION OF MICE WITH 12 ZONA-FREE MOUSE OOCYTES

Several intrasplenic immunizations with batches of approximately 15 or approximately 30 zona-free, unfertilized mouse oocytes resulted in 20 0-300 hybrids, respectively, among which about 20 positive clones were selected from each fusion between splenic plasma cells and SP2/0 myel oma cells. When nonimmunized splenic plasma cells were used, only one antibody, showing weak immunoreaction, was obtained from approximately 370 hybrids collected from 2 fusions. From one immunization with a to tal of 12 zona-free, unfertilized mouse oocytes, 15 positive clones we re selected for further study. Eleven of these 15 antibodies reacted w ith antigens only in unfertilized oocytes but not in fertilized, pronu clear stage oocytes. Three antibodies, which recognized antigens in pa raffin-embedded oocyte sections, did not label growing ovarian oocytes , indicating that the antibodies were specific to ovulated, unfertiliz ed oocytes. These antibodies did not detect any antigen epitopes in th e panel of tissues examined. The molecular weight of one antigen, corr esponding to a IgM antibody that is present both in ooplasma and zona pellucida, was approximately 116 kDa. Cross-reactivity to blots of unf ertilized zona-free hamster oocytes was demonstrated by 6 antibodies a nd to unfertilized human oocytes by 7 antibodies. Three antibodies cro ss-reacted with both hamster and human oocytes. The study indicates th at the intrasplenic immunization is an appropriate means of raising an tibodies against unfertilized, zona-free mouse oocytes and that the me thod applied offers an easy way to select antibodies against human ooc ytes for functional studies. (C) 1994 Wiley-Liss, Inc.