| Description |
Microfluidic technology has the unique potential to separate sperm from unwanted debris while improving the effectiveness of assisted reproductive technologies (ART). Limitations of current clinical protocols regarding separation of sperm from other cells and cellular debris can lead to low sperm recovery when the sample contains low concentrations of mostly low motility sperm and a high concentration of unwanted cells or cellular debris, such as occurs with surgical testis dissection samples from nonobstructive azoospermia (NOA) patients who have undergone microsurgical testicular sperm extraction (mTESE), and semen samples from leukospermia patients (high white blood cell (WBC) semen). Over the years, most microfluidic sperm separation approaches have relied on sperm motility for separation with added features through which only highly motile sperm can pass. Thus, these techniques can separate only progressive motile sperm from semen samples, but they lose a significant number of sperm cells including viable nonprogressive motile and nonmotile sperm. This dissertation demonstrates label-free separation of sperm from challenging sperm samples using inertial microfluidics. The approach does not require any externally applied forces except the movement of the fluid sample through the instrument. In this way, it is possible to recover not only any motile sperm, but also viable less-motile and nonmotile sperm with high recovery rates. The results show the usefulness of inertial microfluidics to significantly reduce the concentrations of unwanted cells/cellular debris (Red blood cells/White blood cells) significantly by flow focusing of debris within a spiral channel flow. The majority (∼80%) of sperm cells collect to the designated outlet and ∼98% of debris goes to the waste outlet. The estimated sample process time is more rapid (∼5minutes) and autonomous than conventional methods which may take between ∼1 hour (semen purification) and 10 ∼18 hours (manual mTESE sample search process). The flow focusing results of sperm and blood cells included that sharp flow focusing of RBC and WBC, but not of sperm cell where sharp flow focusing didn't appear. The successful flow focusing of RBC and WBC imply that the spherical model did accurately predict the behavior of RBCs and WBCs, but the lack of definitive focusing of sperm cells imply that the modeling of sperm cells wasn't accurate. This partial success of sperm modeling was caused by a lack of understanding of sperm behavior in the curved channel. This dissertation presents an improved model of sperm cell behavior in curved channels based on both 2D COMSOL® simulations and experimental studies. The results show promising evidence that the proposed method should able to generate more precise sperm separation for mTESE samples. Lastly this dissertation also performed viability, toxicity, and recovery tests on the proposed sperm separation method for biocompatibility verification. These tests should provide initial validation of clinical usefulness. |
