Nuclear chromatin variations in human spermatozoa undergoing swim-up and cryopreservation evaluated by the flow cytometric sperm chromatin structure assay

The sperm chromatin structure assay (SCSA) is a flow cytometric (FCM) techn ique which exploits the metachromatic properties of Acridine Orange to moni tor the susceptibility of sperm chromatin DNA to in-situ acid denaturation. SCSA was used to study the chromatin structure variations of human spermat ozoa in semen, both before and after swim-up and after cryopreservation. Se men samples were provided by 19 healthy normozoospermic subjects attending pre-marriage checks. Each sample was divided into three aliquots: the first aliquot was evaluated without further treatment, the second underwent swim -up, and the third was stored according to standard cryopreservation techni ques in liquid nitrogen at -196 degrees C. Samples were also analysed by li ght and fluorescence microscopy (after Acridine Orange staining to evaluate the number of green fluorescent sperm heads), and by computer-assisted sem en analysis. The results showed that post-rise spermatozoa represent a subp opulation characterized by a general improvement of the morphological (redu ction of the percentage of abnormal forms and heads, increase of the green head sperm percentage) and kinetic parameters. This subpopulation also exhi bited improved chromatin structure properties, confirming that these cells have the best structural and functional characteristics, indicative of opti mal fertilizing ability. On the other hand, overall sperm quality deteriora tes after cryopreservation. When thawed spermatozoa underwent an additional swim-up round, a general improvement of nuclear maturity was seen in the p ost-rise spermatozoa.