Abstract Background:Sperm selection for ICSI is based on morphology and motility, but these parameters may not relevant chromatin integrity. One of the sperm selection methods for ICSI is based on sperm functional characteristics. This study was designed to compare two sperm selection methods (HA binding and Zeta method) for selection of spermatozoa with normal morphology and intact chromatin. Methods:Semen samples of 70 infertile couples referring to Isfahan Fertility and Infertility Center was assessed during this study. Semen analysis was carried out according to WHO criteria. Semen samples divided into 3 groups. A portion of neat semen considered as control group and remained of semen used for Zeta and HA binding procedures. Sperm morphology, protamine deficiency and DNA fragmentation were assessed by Papanicolaou staining, Chromomycin A3 (CMA3) staining, and SCD test, respectively in 3 group. Findings:Both HA binding assay and Zeta method are efficient to select sperm with normal morphology and lower protamine deficiency (P < 0.05). But in term of DNA fragmentation Zeta method appear to be more efficient to select sperm with low DNA fragmentation (P < 0.05). Conclusion:Our results suggest that these sperm selection methods can select spermatozoa with normal morphology and protamine content for ICSI. But Zeta method may be more efficient to select sperm with low DNA fragmentation. In patient with high DNA fragmentation, Zeta method can be useful to select spermatozoa for ICSI. Key words:Hyaluronic acid (HA), Zeta potential, Intra-Cytoplasmic Sperm Injection (ICSI), Morphology, Protamine deficiency, DNA fragmentation.