Objective: Male factor infertility accounts for 30% to 50% of the total infertile couples seeking for infertility treatment. In about 40-60% of these men, a specific etiology can not be found. The aim of this study was to confirm the presence of eotaxin and interleukin-4 (IL-4) in human seminal plasma, to show the differences between eotaxin and IL-4 concentrations in fertile and infertile men, and to show the potential relationship between eotaxin and IL-4 levels in semen and spermiogram parameters. In literature, this is the first study that evaluates eotaxin in the human seminal plasma. Material and Methods: The participant of the study was 55 infertile males with abnormal semen parameters as study group and 16 healthy volunteers with normal sperm parameters as the control group. Semen samples were classified according to criteria of the World Health Organization Laboratory Manual. The Morphology of the smears was scored using Kruger's strict criteria. Seminal eotaxin and IL-4 levels were measured by bead based immunoassay multiplex methods. Results: Seminal eotaxin levels were significantly higher in infertilite group compared to fertile donors. There were negative correlations between eotaxin concentrations and parameters such as motility (r= -0.293, p< 0.01) and, +4 motility (r= -0.307, p< 0.01) in the study group. IL-4 levels were similar in fertile and infertile seminal plasma. There were positive correlations between eotaxin and IL-4 levels (r= 0.436 p< 0.01). The receiver operating characteristic curve (ROC) analysis revealed a diagnostic value for eotaxin activity with respect to male factor infetility in case group, with an area under curve of 0.69 (95% confidence interval= 0.55-0.84). Conclusion: Increased levels of eotaxin may play a role in the pathogenesis of male infertility.