10 research outputs found
Damage assessment of the equine sperm membranes by fluorimetric technique
To validate a practical technique of simultaneous evaluation of the plasma, acrosomal and mitochondrial membranes in equine spermatozoa three fluorescent probes (PI, FITC-PSA and MITO) were associated. Four ejaculates from three stallions (n=12) were diluted in TALP medium and split into 2 aliquots, 1 aliquot was flash frozen in liquid nitrogen to induce damage in cellular membranes. Three treatments were prepared with the following fixed ratios of fresh semen: flash frozen semen: 100:0 (T100), 50:50 (T50), and 0:100 (T0). A 150-”L aliquot of diluted semen of each treatment was added of 2 ”L of PI, 2 ”L of MITO and 80 ”L of FITC-PSA; incubated at 38.5ÂșC/8 min, and sperm cells were evaluated by epifluorescent microscopy. Based in regression analysis, this could be an efficient and practical technique to assess damage in equine spermatozoa, as it was able to determine the sperm percentage more representative of the potential to fertilize the oocyte.<br>Para validar uma tĂ©cnica prĂĄtica de avaliação simultĂąnea das membranas plasmĂĄtica, acrossomal e mitocondrial em espermatozĂłides eqĂŒinos trĂȘs sondas fluorescentes (PI, FITC-PSA e MITO) foram associadas. Quatro ejaculados de trĂȘs garanhĂ”es (n=12) foram diluĂdos em meio TALP e divididos em duas alĂquotas, uma alĂquota foi submetida a flash frozen em nitrogĂȘnio lĂquido para induzir danos nas membranas celulares. TrĂȘs tratamentos foram preparados com as seguintes proporçÔes de sĂȘmen fresco: sĂȘmen flash frozen: 100:0 (T100), 50:50 (T50), e 0:100 (T0). Uma amostra de 150 ”L de sĂȘmen diluĂdo de cada tratamento foi adicionada de 2 ”L de PI, 2 ”L de MITO e 80 ”L de FITC-PSA; incubadas Ă 38,5ÂșC/8 min, e as cĂ©lulas espermĂĄticas foram avaliadas por microscopia de epifluorescĂȘncia. Baseados na anĂĄlise de regressĂŁo esta Ă© uma tĂ©cnica eficiente e prĂĄtica para determinar danos em espermatozĂłides eqĂŒinos, capaz de determinar a porcentagem de espermatozĂłides mais representativa do potencial fertilizante do ovĂłcito