In recent decades, humans have faced greater challenges in reproduction. Assisted reproductive technology is the most prominent approach for addressing this problem. Current clinical screening methods simply consider the motility or morphology of the sperm. However, as the spermatozoa need to navigate over a long distance in the female reproductive tract and survive the natural screening processes therein, these methods are imperfect. Many approaches have been undertaken to study the chemotaxis and thermotaxis navigation behavior of spermatozoa, but few of these have involved integrated screening that considers motility, chemotaxis, and thermotaxis based on the biological environment of the human body. Current routine sperm evaluation techniques are inadequate and fail to simultaneously provide conclusive evidence for the thermotactic and chemotactic characteristics of sperm. Thus, such screening of functional spermatozoa will be an advancement in assisted reproduction. In this study, we developed a fully integrated biomimetic microfluidic system for screening sperm for their characteristics when exposed to temperature and chemical gradients. Based on our results, we showed that spermatozoa were attracted by temperature and chemical gradients in the physiological range. Moreover, we ascertained a suitable temperature gradient range for thermotaxis and statistically proved that the thermotactic and chemotactic responses are not linked. Here, we report the first quantitative study of functional sperm during thermotaxis and chemotaxis, and our analysis of the difference in motility caused by different conditions. More broadly, we foresee the clinical application of these biologically motivated parameters and characteristics in assisted reproduction in humans.