Flow cytometry application for the acrosomal exocytosis evaluation in human sperm

FLORES MONTERO K; BOGAMILSKY, P; RUETE MC; GARCIA SAMARTINO, C

Mendoza

Simposio; II Simposio Internacional de Medicina Traslacional - XIV Jornadas de Investigación de la Facultad de Ciencias Médicas; 2019

FCM - FCEN - UNCuyo

The sperm acrosome reaction (AR) is a regulated exocytosis essential for fertilization. The fusion of the plasma membrane and the secretory granule membrane of the sperm (acrosome), leads to the exit of acrosomal content and subsequent oocyte fecundation. In our laboratory, the technique of staining sperm with PSA lectin bound to fluorescein isothiocyanate is used to evaluate AR by observation in an epifluorescence microscope. Flow cytometry provides a rapid analysis to measure the amount of one or more fluorescent stains associated with cells in an unbiased manner. The aims of this work were: 1) to establish a rapid and efficient assay for the evaluation of AR in human sperm and 2) to correlate it with the fluorescence microscopy technique used in the laboratory. Sperm were permeabilized with streptolysin O and then treated with calcium as an inducer of acrosomal exocytosis in the presence of PSA-FITC lectin in the incubation medium. The AR was evaluated by flow cytometry. The results obtained by flow cytometry and fluorescence microscopy were compared. In the case of cytometry, more than 5000 sperm were evaluated per condition and in the case of fluorescence microscopy, only 200 spermatozoa were evaluated. The obtained results were similar in both procedures, however, the speed, the number of sperm counted and the objectivity were higher through flow cytometry. We can conclude that cytometry is a quick and objective method; therefore, serial sampling can be performed in a more effective way than in the case of fluorescence microscopy analysis for the AR evaluation.