REMOVAL OF SPERM COATING ANTIGENS BY PERCOLL GRADIENT CENTRIFUGATION.
1B. R. Gilbert, 2G. W. Cooper and 2D. L. Rosenfeld, 1The James Buchanan Brady Foundation, Div. of Urology, The New York Hospital-Cornell University Medical Center, N.Y. and 2Div. of Human Reproduction, Dept. of Ob/Gyn, The North Shore University Hospital, Manhasset, N.Y.
The role human sperm surface coating antigens play in sperm transport and fertilization is unknown. Evidence will be presented that discontinuous gradient centrifugation through Percoll removes at least two types of sperm coating antigens acquired from seminal fluid . These include: 1) secretory blood group A antigens which can be passively transferred to sperm obtained from blood type O donors and 2) non-blood group antigens of unknown character which react with isotypic IgG antisperm antibodies from women. A subset of antisperm antibody (ASA), primarily of the IgM class has been shown previously to exhibit differential binding to spermatozoa of blood group A and AB men when compared to blood group O spermatozoa. We have labeled spermatozoa from blood group O donors with either semen antigens (1/10 dilution) from blood group A patients or their own semen. These spermatozoa were then washed in standard fashion or placed on a Percoll discontinuous gradient (90%, 70%, 40%) and centrifuged for 35 minutes at 1300 RPM (300G) to remove excess antigen and then labeled with IgM antibody. Excess antibody was removed by washing and the presence of antibody evaluated by the immunobead binding assay (IBB) of motile sperm. Reduction in antibody binding to background levels was found only in the Percoll processed spermatozoa. The antigens which were removed by Percoll centrifugation were shown to have terminal sugars of secreted blood group oligosaccharides, since antibody attachment to the blood group antigen was blocked by co-incubation with the monosaccharide N-acetyl galactosamine. These results show that certain sperm coating antigens can be removed form the sperm surface by Percoll gradient centrifugation. Seminal antigens reacting with IgG isotypic antibodies were demonstrated by the marked reduction in IBB of motile sperm, which occurred after incubation in female sera following Percoll treatment as compared to washing in F-10 alone. The ability to remove sperm surface coating antigens now allows investigation of their role in sperm transport, in women known to have antisperm antibodies and in unexplained infertility.