Exhaustion of racing sperm in nature-mimicking microfluidic channels during sorting

Abstract

Fertilization is central to the survival and propagation of a species, however, the precise mechanisms that regulate the sperm's journey to the egg are not well understood. In nature, the sperm has to swim through the cervical mucus, akin to a microfluidic channel. Inspired by this, a simple, cost-effective microfluidic channel is designed on the same scale. The experimental results are supported by a computational model incorporating the exhaustion time of sperm.

Keywords: biodesign; fertility; microfluidic sorting; sperm exhaustion; sperm sorting.

Figure 1

A schematic illustration of unloading sperm samples into the microchannels of space-constrained microfluidic sorting (SCMS) system from the inlets. (A) SCMS system with different channel lengths is assessed for effective sperm sorting. Microchannels are prefilled with media prior to loading the sperm sample. The outlet is then covered with mineral oil to avoid evaporation. (B) Microscope image of the channel inlet with a diameter of 0.65 mm under a 2X objective. (C) Image of sperm cells swimming inside a microchannel under a 10X objective. (D) Microscope image of the channel outlet with a diameter of 2 mm under a 2X objective. Scale bars for the channel inlets and outlets are 1 cm. (E) Sperm tracks obtained by ImageJ to evaluate velocities and persistence time of sperm cells. (F) A schematic of the trajectory of a sperm performing a Persistent Random Walk (PRW), where S is the velocity, P is the persistence time, Δt is the time step, and θ is the angle that the trajectory makes with the x-axis.

Figure 2

Comparison of experimental and simulated mouse sperm distributions within the channels of space-constrained microfluidic sorting (SCMS) microchips after varying incubation times. (A) Distribution of sperm within the microchannel after incubation for 1 h. Experimental results are compared with the computational model with different parameters: (1) Persistent Random Walk (PRW), (2) PRW and initially 25% of sperm are dead, (3) PRW including 30 minutes average exhaustion time of sperm (± 15 minutes), and (4) PRW including both exhaustion time and initially 25% dead sperm. (B) Distribution of sperm within the microchannel after an incubation period of 5 min, 15 min, 30 min and 1 h. Experimental results are compared with simulation results from PRW model with both exhaustion time and 25% initially dead sperm population for the corresponding incubation time. Data were presented as average ± standard error.

Figure 3

Evaluation of channel length and incubation time by using space-constrained microfluidic sorting (SCMS) systems for mouse sperm sorting. The effective sorting of the microchannels with varying channel lengths was illustrated through (A) Curvilinear velocity (VCL), (B) Straight-line velocity (VSL), (C) Linearity of sperm (LIN), and (D) percentage of motile sperm at the inlet and the outlet of each channel for 30 min and 1 h of incubation. The statistical significance between channel lengths were marked with *, and between inlets and outlets were marked with #. Data were presented as average ± standard error (SEM) (N=22–109).

Figure 4

Mouse sperm sorted using space-constrained microfluidic sorting (SCMS) system with 15 mm long channel was compared to those using swim-up technique and non-sorted sperm. The comparisons were performed for (A) Curvilinear velocity, (B) Straight-line velocity, (C) Linearity, and (D) percentage of motile sperm for 30 min of incubation. SCMS system with 15 mm length and 30 min of incubation resulted in sperm with higher motility and percentage of motile sperm compared to swim-up technique and non-sorted sperm. Data were presented as average ± standard error (SEM) (N=4–7). (E) The sperm curvilinear velocity, straight-line velocity, and collectable sperm percentage were compared after sorting using space-constrained microfluidic sorting (SCMS) system with different channel lengths (7 mm, 10 mm, 15 mm and 20 mm) for 30 minutes of incubation. Data were presented as average ± standard error (SEM) (N=3).

Figure 5

Comparison of experimental and simulated human sperm distributions within 20 mm long channel of space-constrained microfluidic sorting (SCMS) microchips after varying incubation times. (A) Distribution of sperm within the microchannel after an incubation period of 5 min, 15 min, 30 min and 1 h. Experimental results are compared with simulation results from Persistent Random Walk (PRW) model with no exhaustion time and 25% initially dead sperm population for the corresponding incubation time. Data were presented as average ± standard error. (B & C) The effective sorting of the microchannels with varying channel lengths was illustrated through (B) Curvilinear velocity (VCL), and (C) Straight-line velocity (VSL) at the inlet and the outlet of each channel for 30 min and 1 h of incubation. The statistical significance between channel lengths were marked with *, and between inlets and outlets were marked with #. Data were presented as average ± standard error (SEM) (N=27–81).