G M Centola1, E Andolina, A Deutsch. 1. Department of OB/GYN, University of Rochester, NY 14618, USA. gcentola@obgyn.rochester.edu
Abstract
PROBLEM: The IBT is considered the gold standard of sperm antibody assays. This test uses polyacrylamide beads labeled with antiglobulins (anti-IgG, anti-IgA, and anti-IgM), which bind to the corresponding antibody on the sperm surface. The IS uses color-coded latex beads of uniform 3.0 microns size coated with the antiglobulins which can be viewed with brightfield light microscopy. The purpose of the present study was to compare the IBT and IS in an indirect test using human serum. METHOD: Serum specimens (n = 42) were tested for the presence of antibody isotypes IgG, IgA, and IgM to sperm using the standard protocol for IBT and IS. Donor sperm was washed in BWW with 5% BSA and diluted to a final concentration of 50 x 10(6) motile sperm/ ml. The sperm were incubated with a 1:10 dilution of test serum for 30 min to 1 h at 37 degrees C and then washed by three cycles of centrifugation. The sperm and beads (IBT, IS) were mixed on a glass slide, covered with a coverslip, and observed within 5 min. At least 100 motile sperm were counted and scored for bead binding. A specimen was considered positive if 20% or more of the sperm were coated with one or more beads. The data were analyzed using calculation of the non-parametric kappa statistic with correction for chance expected agreement, and by calculating the proportion of specific agreement between the two methods. RESULTS: The results are summarized in the following table: [table see text]; The IS was able to detect 94% of IgG antibodies, 91% of IgA antibodies, and 100% of IgM antibodies. One serum specimen was IgG negative by IS (14% binding), but positive by IBT (20%). A second serum specimen was IgA negative by IS (16%) yet positive by IBT (29%). There were no false positives with the IS assay. Of the IgM positives (five of six) occurred alone and not with IgG or IgA, suggesting the necessity for testing all specimens also for IgM. CONCLUSION: Antisperm antibody test results obtained by the IS assay are in agreement with the results obtained with the IBT test. The Immunospheres are monodispersed, color coded, and can be visualized with brightfield microscopy.
PROBLEM: The IBT is considered the gold standard of sperm antibody assays. This test uses polyacrylamide beads labeled with antiglobulins (anti-IgG, anti-IgA, and anti-IgM), which bind to the corresponding antibody on the sperm surface. The IS uses color-coded latex beads of uniform 3.0 microns size coated with the antiglobulins which can be viewed with brightfield light microscopy. The purpose of the present study was to compare the IBT and IS in an indirect test using human serum. METHOD: Serum specimens (n = 42) were tested for the presence of antibody isotypes IgG, IgA, and IgM to sperm using the standard protocol for IBT and IS. Donor sperm was washed in BWW with 5% BSA and diluted to a final concentration of 50 x 10(6) motile sperm/ ml. The sperm were incubated with a 1:10 dilution of test serum for 30 min to 1 h at 37 degrees C and then washed by three cycles of centrifugation. The sperm and beads (IBT, IS) were mixed on a glass slide, covered with a coverslip, and observed within 5 min. At least 100 motile sperm were counted and scored for bead binding. A specimen was considered positive if 20% or more of the sperm were coated with one or more beads. The data were analyzed using calculation of the non-parametric kappa statistic with correction for chance expected agreement, and by calculating the proportion of specific agreement between the two methods. RESULTS: The results are summarized in the following table: [table see text]; The IS was able to detect 94% of IgG antibodies, 91% of IgA antibodies, and 100% of IgM antibodies. One serum specimen was IgG negative by IS (14% binding), but positive by IBT (20%). A second serum specimen was IgA negative by IS (16%) yet positive by IBT (29%). There were no false positives with the IS assay. Of the IgM positives (five of six) occurred alone and not with IgG or IgA, suggesting the necessity for testing all specimens also for IgM. CONCLUSION: Antisperm antibody test results obtained by the IS assay are in agreement with the results obtained with the IBT test. The Immunospheres are monodispersed, color coded, and can be visualized with brightfield microscopy.