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EVALUATION OF CHROMOMYCIN A3 ASSAY IN HUMAN SPERM AFTER SIMULATED OVERNIGHT SHIPMENT
Citation:
Jeffay, S C., R A. Morris, L F. Strader, Olshan. AF, D. P. Evenson, AND S P. Darney. EVALUATION OF CHROMOMYCIN A3 ASSAY IN HUMAN SPERM AFTER SIMULATED OVERNIGHT SHIPMENT. Presented at American Society of Andrology, Baltimore, MD, April 17-20, 2004.
Description:
EVALUATION OF CHROMOMYCIN A3ASSAY IN HUMAN SPERM AFTER SIMULATED OVERNIGHT SHIPMENT.
SC Jeffay1, R Morris Buus1, LF Strader1, AF Olshan2, DP Evenson3, SD Perreault1. 1US EPA/ORD, RTP, NC;2UNC-CH, Chapel Hill, NC;3SDSU, Brookings, SD.
Semen collection kits that allow men to collect a sample at their convenience and send it via overnight mail to the laboratory facilitate sampling from different geographic locations and minimize variability through analysis by a centralized lab. We are investigating temperature requirements for sample stability during shipment. Previously, we found that the % sperm with DNA fragmentation (DFI) according to the Sperm Chromatin Structure Assay (SCSA?) increased significantly with simulated overnight shipment at room temperature (RT), but not at 4oC. Here we report on another sperm chromatin assay,Chromomycin A3 (CMA), using the same samples. CMA competes with protamine for DNA.Therefore, protamine-deficientsperm take up the dye and fluoresce brightly (CMA+), while protamine-richsperm do not. Aliquots of semen samples from 24 fertile men were frozen within 2 hours of collection (Dy1), or after overnight storage at either RT (Dy2-RT), or at 4oC to simulate shipment with cold packs (Dy2-4oC). The mean % CMA+ sperm for Dy1 (53.5?3.2, SEM) was not different from Dy2-RT (54.0?3.6) or Dy2-4oC (57.7?3.0)by a Repeated Measures ANOVA with Tukey-Kramer Multiple Comparisons Test on data paired by subject. Protamine-deficient sperm would be expected to be less condensed and therefore might also exhibit?High DNA Stainability? (HDS) in the SCSAa. The %HDS for these samples was also not different by treatment (7.3?0.7, 6.4?0.7, 7.0?0.8, respectively). As expected, %HDS and %CMA+ werecorrelated, p<0.0001, r = 0.6730. These results suggest that sperm nuclear proteins are relatively stable for up to 24 hr at RT or 4oC; therefore, CMA can be performed on semen shipped overnight at either temperature. This abstract does not reflect EPA policy. Supported by CR829327.