Motility is an indicator of sperm cell viability due to higher probability in swimming through the female reproductive tract and undergo fertilization with the egg cell. Centrifugation method is a technique to process high volume semen and isolate motile sperm cells but decreases the biochemical integrity of spermatozoa due to the contact with reactive oxygen species (ROS) from dead cells released during centrifugation. This study uses solution electrospun poly(ε-caprolactone) membranes as an alternative in isolating motile spermatozoa by utilizing a rationally designed 3D printed module set up, providing the same benefits as commercially available techniques with minimal processing time, and bypassing the centrifugation step to provide higher quality sperm cells. The membranes, with nominal pore size distributions ranging from 5 to 6 µm are highly porous structures suitable for establishing baseline data for sperm cell sorting by motility. The proposed method allows for isolation of motile sperm cells with 74% purity, while decreasing the processing time by 98% when compared to centrifugation techniques. This novel approach provides a facile method for isolating motile spermatozoa directly from frozen semen samples without any pretreatments and is easily scalable for small and medium scale farms as well as larger industries.