The Sperm Chromatin Dispersion Test: A Simple Method for the Determination of Sperm DNA Fragmentation

摘要: Sperm DNA fragmentation is being increasingly rec- ognized as an important cause of infertility. We herein describe the Chromatin Dispersion (SCD) test, a novel assay for sperm in semen. The SCD test based on principle that with fragmented fail to produce characteristic halo dispersed loops observed non- DNA, following acid denaturation and removal nuclear proteins. This was confirmed by analysis using specific Breakage Detection-Fluorescence In Situ Hy- bridization (DBD-FISH) assay, which allows detection breaks lysed nuclei. suspensions either prepared from semen or isolated gradient centrifugation were embedded agarose microgel slides treated 0.08 N HCl lysing solutions containing 0.8 M dithiothreitol (DTT), 1% sodium dodecyl sulfate (SDS), 2 NaCl. Then, sequentially stained DAPI (49,6-diamidino-2-phenylindole) and/ Diff-Quik reagent, percentages nondis- persed chromatin monitored fluores- cence brightfield microscopy, respectively. results indicate all nondispersed displayed fragmen- tation, measured DBD-FISH. Conversely, dis- had very low undetectable DBD-FISH labeling. values significantly higher patients screened infertility than normozoospermic donors who par- ticipated donor insemination program. coefficient varia- tion obtained different observers, digital image (DIA) microscopy scoring, less 3%. conclusion, simple, accurate, highly repro- ducible, inexpensive method frag- mentation processed sperm. Therefore, could potentially be used routine screening andrology laboratory.