A Systematic Evaluation of Methods to Separate X- and Y- Bearing Sperm
This project was initiated to determine if was possible to enrich either X- or Y- bearing sperm, and hence to preselect the sex of a child. Two of the possible reasons why couple might want to select the sex of a child are firstly because of a family history of an X-linked recessive genetic disorder, which usually only affect sons, and secondly families who have had several children of one sex. For this study, men with three or more children of the same sex were recruited following the publication of an article in The West Australian newspaper. The percentage of X- and Y- bearing sperm within the semen samples of men with three or more children of the same sex was determined using dual colour fluorescence in situ hybridisation (FISH). The aim of the investigation was to determine if these men had an altered ratio of X- to Y- bearing sperm, which would explain why these men had a predominance of children of one sex. Comprehensive analyses were also carried out on the semen samples. The reliability of the dual colour FISH technique was established using a number of standard metaphase spreads; from male and female subjects and an individual with Klinefelters syndrome. It was determined that dual colour FISH was a suitable technique for determining the percentage of X- or Y- bearing sperm within a sample. The semen samples were processed using one of two protocols. Samples from men with three or more daughters were treated using Human serum albumin columns, with the intention of increasing the percentage of Y-bearing sperm within the final fraction. It has been suggested that this method enriches the Y- bearing sperm from a sample due to the differential motility exhibited by the X- and Y- bearing sperm, although this characteristic has not been proven. Samples from men with three or more sons were processed using 8-layer ISolate® discontinuous gradients, with, the aim of enhancing the amount of X- bearing sperm within the final fraction. This method is based on the formation of the discontinuous gradients because Percoll has not been approved for the production of sperm fractions for human insemination. It has been suggested that the X- and Y- bearing sperm can be enriched using such gradients either as a result of differences in their velocity of sedimentation or due to a greater nett negative charge on the surface of X- bearing sperm. However, neither of these theories have been validated. As it has also been proposed that the survival rate of X- bearing sperm is slightly longer than that for Y- bearing sperm, this was also investigated. In summary no statistically significant enrichment of X- or Y- bearing sperm was observed following the treatment of the semen samples with either the ISolate® discontinuous gradient or the Human serum albumin column protocols. Nor was there any enrichment in X- bearing sperm due to their suggested greater survival time.
Sex chromosomes, Sex preselection, Spermatozoa, Analysis, Medical Sciences, Medicine and Health Sciences
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Edith Cowan University, Research Online, Perth, Western Australia