Influence of swim-up time on the ratio of X- and Y-bearing spermatozoa

ObjectiveThe objective was to examine the separation of X- and Y-bearing spermatozoa in a modified swim-up procedure using fluorescent in situ hybridization (FISH), and to find out the influence of swim-up time on the ratio of X- and Y-bearing spermatozoa.Study designProspective study.SettingReproductive testing laboratory in a university hospital.PatientsNormal spermatozoa samples were obtained from 10 volunteers by masturbation after sexual abstinence for 3–5 days.InterventionsSpermatozoa were put into 18 tubes with 0.25 ml in each, then mixed with HTF medium and centrifuged for 5 min (400 × g). The supernatant was removed and discarded and 0.5 ml HTF was added slowly along the tube wall. Motile spermatozoa were collected after swimming up in different times (from 5 up to 150 min, with a total of 17 intervals). The X- and Y-bearing spermatozoa were determined using the FISH technique. The X/Y dual-color CEP probes that were marked by fluorescein isothiocyanate (FITC) and Texas red were applied to analyze the ratio of X- and Y-bearing spermatozoa. The FISH staining slides were analyzed under an immunofluorescence microscope. About 1000–1500 spermatozoa were counted per slide.Main outcome measuresThe percentages of X- and Y-bearing spermatozoa were calculated.ResultsThe study results suggested that the total ratio of hybridization was 98.33%. The ratio of X-bearing spermatozoa after swimming up for different amounts of time is 50.03 ± 0.91% at 0 min, 50.45 ± 2.06% after 15 min, 50.61 ± 2.47% after 30 min, 50.16 ± 2.67% after 60 min, 50.72 ± 2.64% after 90 min, and 50.56 ± 2.20% after 150 min. The statistical analysis showed that there were no significant differences among different swim-up times in the ratio of X-bearing spermatozoa.ConclusionsThere was no significant effect of swim-up time on the ratios of X- and Y-bearing spermatozoa using a modified swim-up procedure. No direct evidence was found that the swim-up procedure for separating motile spermatozoa to use for either intrauterine insemination (IUI) or in vitro fertilization (IVF) would lead to an imbalance of boys and girls.