High-throughput analysis of horse sperms' 3D swimming patterns using computational on-chip imaging.

Using a high-throughput optical tracking technique that is based on partially-coherent digital in-line holography, here we report a detailed analysis of the statistical behavior of horse sperms' three-dimensional (3D) swimming dynamics. This dual-color and dual-angle lensfree imaging platform enables us to track individual 3D trajectories of ∼1000 horse sperms at sub-micron level within a sample volume of ∼9μL at a frame rate of 143 frames per second (FPS) and collect thousands of sperm trajectories within a few hours for statistical analysis of their 3D dynamics. Using this high-throughput imaging platform, we recorded >17,000 horse sperm trajectories that can be grouped into six major categories: irregular, linear, planar, helical, ribbon, and hyperactivated, where the hyperactivated swimming patterns can be further divided into four sub-categories, namely hyper-progressive, hyper-planar, hyper-ribbon, and star-spin. The large spatio-temporal statistics that we collected with this 3D tracking platform revealed that irregular, planar, and ribbon trajectories are the dominant 3D swimming patterns observed in horse sperms, which altogether account for >97% of the trajectories that we imaged in plasma-free semen extender medium. Through our experiments we also found out that horse seminal plasma in general increases sperms' straightness in their 3D trajectories, enhancing the relative percentage of linear swimming patterns and suppressing planar swimming patterns, while barely affecting the overall percentage of ribbon patterns.