title: Influence of staining and sampling procedures on goat sperm morphometry using the Sperm Class Analyzer.
abstract: Computer-assisted sperm morphometry analysis (ASMA) has improved the assessment of sperm morphology, but the results depend on the use of adequate sampling and staining procedures of spermatozoa from individual species. In this study, the Sperm Class Analyzer ASMA system was used for the morphometric analysis of goat sperm heads. Semen samples, obtained from four bucks, were used to evaluate the influence of three staining procedures (Diff-Quik, Hemacolor and Harris' Haematoxylin) on the accuracy of image processing and sperm morphometry, the effect of the number of cells analysed and the repeatability of the method. These experiments were performed to obtain objective, accurate and reliable sperm morphometric measurements of goat spermatozoa. Diff-Quik and Harris' Haematoxylin were significantly (p<0.05) more accurate than Hemacolor. However, Diff-Quik obtained the highest proportion of correctly analysed sperm heads (86.06%) and the lowest coefficients of variation on the image processing and morphometric measurements. The staining methods affected significantly the sperm dimensions (p<0.001) with increased values from Diff-Quik than Hemacolor and Harris' Haematoxylin, respectively (Diff-Quik>Hemacolor>Harris' Haematoxylin). No differences in morphometric parameters were found when 100, 150, 175 or 200 spermatozoa were analysed. The repeatability of results obtained was very high since no differences were found when measuring the same sperm on multiple attempts. In conclusion, to obtain objective, accurate and repeatable sperm morphometric measurements by the Sperm Class Analyzer system in goats, the analysis of 100 spermatozoa from slides which have been previously stained with Diff-Quik is recommended.
authors.affiliation: Reproduction and Obstetrics Unit, Faculty of Veterinary Science, University of Cordoba, 14071 Cordoba, Spain. mhidalgo@uco.es
authors.forename: Manuel
authors.initials: M
authors.lastname: Hidalgo
authors.forename: Inmaculada
authors.initials: I
authors.lastname: Rodríguez
authors.forename: Jesús
authors.initials: J
authors.lastname: Dorado
mesh_terms: D000818:Animals; D004396:Coloring Agents / Q000494:pharmacology; D006041:Goats / Q000502:physiology; D007091:Image Processing, Computer-Assisted / Q000379:methods; D008297:Male; D008854:Microscopy, Electron / Q000295:instrumentation*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D053625:Sperm Retrieval; D013094:Spermatozoa / Q000166:cytology* / Q000187:drug effects / Q000648:ultrastructure; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D004396:Coloring Agents