Abstract: The present technology relates to a systems for quantifying rheotaxis in a sperm-containing sample. The system includes a microfluidic system having an inlet for charging fluids into a passage. One or more probes defining a confinement region suitable for retaining motile sperm are within the passage. The system further comprises an image processing computing device for obtaining a sequence of images of the confinement region of at least one of the one or more probes having motile sperm retained therein. The sequence of images of the confinement region is processed to determine a signal intensity value for said sequence of images, wherein said signal intensity value is based on a concentration of the motile sperm located in the confinement region at the flow rate. A rheotaxis quality value is determined for said sperm-containing sample based on the signal intensity value. Methods for quantifying rheotaxis in a sperm-containing sample are also disclosed.