Detection of sex chromosomal aneuploidies X‐X, Y‐Y, and X‐Y in human sperm using two‐chromosome fluorescence in situ hybridization

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Abstract
Sex chromosome aneuploidy is the most common numerical chromosomal abnormality in humans at birth and a substantial portion of these abnormalities involve paternal chromosomes. An efficient method is presented for using air‐dried smears of human semen to detect the number of X and Y chromosomes in sperm chromatin using two‐chromosome fluorescence in situ hybridization. Air‐dried semen smears were pre‐treated with dithiothreitol and 3,4‐diio‐dosalicylate salt to decondense the sperm chromatin and then were hybridized with repetitive sequence DNA probes that had been generated by PCR and differentially labeled. Hybridizations with X and Y specific probes showed the expected ratio of 50%X:50%Y bearing sperm. Sperm carrying extra fluorescence domains representing disomy for the X or Y chromosomes occurred at frequencies of ∼4 per 10,000 sperm each. Cells carrying both X and Y fluorescence domains occurred at a frequency of ∼6/10,000. Thus, the overall frequency of sperm that carried an extra sex chromosome was 1.4/1,000. The frequencies of sperm carrying sex chromosome aneuploidies determined by hybridization did not differ statistically from those reported from the same laboratory using the human‐sperm/hamster‐egg cytogenetic technique. Multi‐chromosome fluorescence in situ hybridization to sperm is a promising method for assessing sex‐ratio alterations in human semen and for determining the fraction of sperm carrying sex or other chromosome aneuploidies which may be transmissible to offspring.