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Purification of human seminal plasma no. 7 antigen by using immunoaffinity chromatography bound monoclonal antibody
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antibody response which is indistinguishable from hyperimmune sera. The primary and secondary alloantibody responses in the allogeneically pregnant rat differ dramatically with regards to the kinetics of the response and with regards to the qualitative nature of the alloantibodies produced.
ANTIFERTILITY EFFECTS FROM IMMUNIZATION OF FEMALE BABOONS WITH C-TERMINAL PEPTIDES OF HCG BETA SUBUNIT. Vernon Stevens, John Powell, Arthur Lee, and P..D-Griffin., Department of Obstetrics and Gynecology, Ohio State University, Columbus, Ohio and World Health Organization, Geneva, Switzerland. The effects of immunization of adult,female baboons with a synthetic carboxy-terminal peptide of hCG beta subunit on f e r t i l i t y was evaluated by comparing the pregnancy rate in 15 immunized animals with the rate observed in 15 control female baboons. The control group was immunized with tetanus toxoid at 28 day i n t e r v a l s , and the hCG vaccine group was immunized with a conjugate containing equal weights of tetanus toxoid and hCG beta subunit peptide I09-145. All animals received the primary i n j e c t i o n in CFA between days 1-5 of t h e i r menstrual cycle and matings with males of proven f e r t i l i t y begun during the t h i r d cycle of immunization. Those females not pregnant were mated in three consecutive cycles. In the cont r o l group, pregnancies among the 15 animals were: 10 in the 1st mating, 4 in the 2nd, and 1 in the 3rd mating. In the conjugate immunized group, pregnancies among the 15 baboons were: 1 in the 1st mating, 1 in the 2nd, and 2 in the 3rd mating. The pregnancy rate f o r controls ( 15 pregnancies / 21 matings ) was 71.4%, while the f e r t i l i t y rate in the conjugate immunized baboons ( 4 pregnancies/42 matings ) was 9.5%. The difference in f e r t i l i t y rates between controls and conjugate groups was highly s i g n i f i c a n t upon s t a t i s t i c a l analysis ( p <0.0005 ). PURIFICATION OF HUMAN SEMINAL PLASMA NO.7 ANTIGEN BY USING IMMUNOAFFINITY CHROMATOGRAPHY BOUND MONOCLONAL ANTIBODY. Shinzo Isojima, Koji Koyama and Keiko Fujiwara. Department of Obstet. and Gynecol. Hyogo Medical College Nishinomiya, 663, Japan Recently the monoclonal antibody IgG to human seminal plasma (HSP) antigen No.7 from the cloned hybridoma was successfully obtained in our laboratory. The HSP No.7 antigen is contained in both seminal plasma and milk protein and its antibody could immobilize the human spermatozoa
$39
with complement. The human seminal plasma protein was passed through the column of CNBr activated Sepharose 4B which bound the monoclonal antibody y-globulin. After washing with borate buffer, 0.2M glycine-HCl buffer (pH 2.5) was applied to elute the corresponding antigen. In several experiments, the recoveries of bound antigen were from 0.7 to 1.0% but the antigenic activity to neutrize the sperm immobilizing antibody was increased to 77.3 times. After rechromatography by the same procedures, the obtained fraction indicated as 196.2 times as neutralizing activity for sperm immobilizing antibody. This high purity gives us the hope that we can prepare the vaccine of seminal plasma for contraception by using the same procedure.
BINDING CHARACTERISTICS OF ANTIBODIES TO THE~-SUBUNIT OF OVI~E LUTEINIZING HORMONE (oLH~) WITH HUMAN AND RHESUS GONADOTROPINS. Y.yamamoto, K.Sundaram and R.B.Thau, Population Council, New York, N.Y. 10021 We have reported that active immunization with oLH~ induced infertility in rhesus monkeys but did not interfere with regular menstrual cycles and ovulation. Cross-reaction of antisera to oLH~ with LH and chorionic gonadotropin (CG) have not previously been studied in primates. We therefore determined the binding characteristics of oLH~-antisera with hLH and hCG in homologous Scatchard plots using iodinated hCG or hLH and increasing amounts of non-radioactive hormone in monkeys with high and low antibody titers 4 wks after the last booster. Association constants were of the same order of magnitude for all groups, although for high titer animals the mean value was significantly lower for hLH as compared to hCG. In contrast, the number of binding sites differed by a factor of i0 for high and low titer monkeys. In one high titer monkey, association constants for rhLH and rhCG were also measured, using iodinated hCG and increasing doses of rhLH or rhCG. The affinities for rhLH and rhCG were similar to those for the human hormones. These results show that in all monkeys with antibody titers above 30%, oLH~-antisera have high affinities to hLH and hCG. The number of binding sites correlate well with antibody titers. Because of the similarities of the binding characteristics of oLH~ antisera with hormones from the human and the rhesus monkey, the monkey is a good animal model to study an infertility vaccine based on oLH~-antibodies for possible application in the human.
antibody response which is indistinguishable from hyperimmune sera. The primary and secondary alloantibody responses in the allogeneically pregnant rat differ dramatically with regards to the kinetics of the response and with regards to the qualitative nature of the alloantibodies produced.
ANTIFERTILITY EFFECTS FROM IMMUNIZATION OF FEMALE BABOONS WITH C-TERMINAL PEPTIDES OF HCG BETA SUBUNIT. Vernon Stevens, John Powell, Arthur Lee, and P..D-Griffin., Department of Obstetrics and Gynecology, Ohio State University, Columbus, Ohio and World Health Organization, Geneva, Switzerland. The effects of immunization of adult,female baboons with a synthetic carboxy-terminal peptide of hCG beta subunit on f e r t i l i t y was evaluated by comparing the pregnancy rate in 15 immunized animals with the rate observed in 15 control female baboons. The control group was immunized with tetanus toxoid at 28 day i n t e r v a l s , and the hCG vaccine group was immunized with a conjugate containing equal weights of tetanus toxoid and hCG beta subunit peptide I09-145. All animals received the primary i n j e c t i o n in CFA between days 1-5 of t h e i r menstrual cycle and matings with males of proven f e r t i l i t y begun during the t h i r d cycle of immunization. Those females not pregnant were mated in three consecutive cycles. In the cont r o l group, pregnancies among the 15 animals were: 10 in the 1st mating, 4 in the 2nd, and 1 in the 3rd mating. In the conjugate immunized group, pregnancies among the 15 baboons were: 1 in the 1st mating, 1 in the 2nd, and 2 in the 3rd mating. The pregnancy rate f o r controls ( 15 pregnancies / 21 matings ) was 71.4%, while the f e r t i l i t y rate in the conjugate immunized baboons ( 4 pregnancies/42 matings ) was 9.5%. The difference in f e r t i l i t y rates between controls and conjugate groups was highly s i g n i f i c a n t upon s t a t i s t i c a l analysis ( p <0.0005 ). PURIFICATION OF HUMAN SEMINAL PLASMA NO.7 ANTIGEN BY USING IMMUNOAFFINITY CHROMATOGRAPHY BOUND MONOCLONAL ANTIBODY. Shinzo Isojima, Koji Koyama and Keiko Fujiwara. Department of Obstet. and Gynecol. Hyogo Medical College Nishinomiya, 663, Japan Recently the monoclonal antibody IgG to human seminal plasma (HSP) antigen No.7 from the cloned hybridoma was successfully obtained in our laboratory. The HSP No.7 antigen is contained in both seminal plasma and milk protein and its antibody could immobilize the human spermatozoa
$39
with complement. The human seminal plasma protein was passed through the column of CNBr activated Sepharose 4B which bound the monoclonal antibody y-globulin. After washing with borate buffer, 0.2M glycine-HCl buffer (pH 2.5) was applied to elute the corresponding antigen. In several experiments, the recoveries of bound antigen were from 0.7 to 1.0% but the antigenic activity to neutrize the sperm immobilizing antibody was increased to 77.3 times. After rechromatography by the same procedures, the obtained fraction indicated as 196.2 times as neutralizing activity for sperm immobilizing antibody. This high purity gives us the hope that we can prepare the vaccine of seminal plasma for contraception by using the same procedure.
BINDING CHARACTERISTICS OF ANTIBODIES TO THE~-SUBUNIT OF OVI~E LUTEINIZING HORMONE (oLH~) WITH HUMAN AND RHESUS GONADOTROPINS. Y.yamamoto, K.Sundaram and R.B.Thau, Population Council, New York, N.Y. 10021 We have reported that active immunization with oLH~ induced infertility in rhesus monkeys but did not interfere with regular menstrual cycles and ovulation. Cross-reaction of antisera to oLH~ with LH and chorionic gonadotropin (CG) have not previously been studied in primates. We therefore determined the binding characteristics of oLH~-antisera with hLH and hCG in homologous Scatchard plots using iodinated hCG or hLH and increasing amounts of non-radioactive hormone in monkeys with high and low antibody titers 4 wks after the last booster. Association constants were of the same order of magnitude for all groups, although for high titer animals the mean value was significantly lower for hLH as compared to hCG. In contrast, the number of binding sites differed by a factor of i0 for high and low titer monkeys. In one high titer monkey, association constants for rhLH and rhCG were also measured, using iodinated hCG and increasing doses of rhLH or rhCG. The affinities for rhLH and rhCG were similar to those for the human hormones. These results show that in all monkeys with antibody titers above 30%, oLH~-antisera have high affinities to hLH and hCG. The number of binding sites correlate well with antibody titers. Because of the similarities of the binding characteristics of oLH~ antisera with hormones from the human and the rhesus monkey, the monkey is a good animal model to study an infertility vaccine based on oLH~-antibodies for possible application in the human.