OBJECTIVE: To compare two commonly used sperm-preparation techniques, density gradient centrifugation and swim-up procedures, with respect to their effects on acrosome reaction (AR), hypoosmotic swelling (HOS) and nuclear maturity in men with abnormal and normal semen analyses. STUDY DESIGN: In accordance with World Health Organization criteria, 23 men with abnormal (group I) and 20 men with normal (group II) semen analyses were included in a prospective, controlled study. Each semen specimen was divided into aliquots in order to assess AR, HOS and nuclear maturity, determined with acridine orange staining, in both raw and processed semen samples using the density gradient centrifugation and swim-up techniques. RESULTS: Initial semen samples in group I revealed diminished AR, HOS and nuclear maturity rates in comparison to those in group II. In group I, density gradient centrifugation improved AR, HOS and nuclear maturity rates more than did swim-up. However, in group II it improved only the AR; HOS rates were better than with swim-up. There was a significant positive correlation between sperm concentration and HOS rate in raw semen samples from group I. In the same group, motility and morphology correlated with the nuclear maturity rate but not with AR and HOS rates. Semen samples with better motility (> 20%) or morphology (> 25%) showed better nuclear maturity rates (> 50%) in men with abnormal semen analyses. Motility had a sensitivity of 77% and specificity of 90% in predicting nuclear maturity. Morphology had similar sensitivity but lower specificity (70%). CONCLUSION: Density gradient centrifugation is superior to the swim-up technique in improving AR, HOS and nuclear maturity rates in men with abnormal semen analyses. However, when only nuclear maturity rate is taken into account, the swim-up technique seems to be sufficient for selecting spermatozoa in men with normal semen analyses. The nuclear maturity rate also correlates with sperm morphology and motility.