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Semi-automated Scoring of Triple-probe FISH in Human Sperm: Methods and Further Validation1
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    Although the frequency and consequence of sperm chromosomal abnormalities are considerable, few epidemiologic studies in large samples have been conducted to investigate etiologic risk factors. This is, in part, attributable to the labor intensive demands of manual sperm fluorescence in situ hybridization (FISH) scoring. As part of an epidemiologic study investigating environmental risk factors for aneuploidy among men attending a hospital-based fertility clinic, a semi-automated method of slide scoring was further validated and used to estimate sex chromosome sperm disomy frequency in a large number of samples. Multiprobe FISH for chromosomes X, Y, and 18 was used to determine sex chromosome disomy in sperm nuclei. Semi-automated scoring methods were used to quantify X disomy (sperm FISH genotype XX18), Y disomy (YY18), and XY disomy (XY18). The semi-automated results were compared with the results from manual scoring in 10 slides. The semi-automated method was then used to estimate sex chromosome disomy frequency in 60 men. Of 10 slides scored, significant differences between the manual and semi-automated results were seen primarily in one slide that was of poor quality because of over swollen nuclei. Among 60 men analyzed using the semi-automated method, median total sex chromosome disomy frequency was 1.65%, which is higher than seen among normal men but within range with reports from fertility clinic populations. These results further validate that semi-automated methods can be used to score sperm disomy with results comparable to manual methods. This is the largest study to date to provide estimates of sex chromosome disomy among men attending fertility clinics. These methods should be replicated in larger clinic populations to arrive at stable estimates of aneuploidy frequency in men who are members of subfertile couples. © 2011 International Society for Advancement of Cytometry.

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    R00 ES017744/ES/NIEHS NIH HHS/United States
    T42 OH008416/OH/NIOSH CDC HHS/United States
    P30 ES000002/ES/NIEHS NIH HHS/United States
    R01 ES009718/ES/NIEHS NIH HHS/United States
    R01 ES009718-01A1/ES/NIEHS NIH HHS/United States
    P30ES000002/ES/NIEHS NIH HHS/United States
    KO1ES10959/ES/NIEHS NIH HHS/United States
    K01 ES010959-01/ES/NIEHS NIH HHS/United States
    K01 ES010959/ES/NIEHS NIH HHS/United States
    P30 ES000002-50/ES/NIEHS NIH HHS/United States
    R01ES09718/ES/NIEHS NIH HHS/United States
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