Virus removal from semen with a pinched flow fractionation microfluidic chip.

Nowadays pigs are bred with artificial insemination to reduce costs and transportation. To prevent the spread of diseases, it is important to test semen samples for viruses. Screening techniques applied are enzyme-linked immunosorbent assays and/or polymerase chain reaction, which are labor-intensive and expensive methods. In contrast to the current used screening techniques, it is possible to remove viruses physically from semen. However, existing methods for virus removal techniques have a low yield of spermatozoa. Therefore, we have developed a microfluidic chip that performs size-based separation of viruses and spermatozoa in boar semen samples, thereby having the potential to reduce the risk of disease spreading in the context of artificial insemination in the veterinary industry. As the head of a spermatozoon is at least twenty times larger than a virus particle, the particle size can be used to achieve separation, resulting in a semen sample with lower viral load and of higher quality. To achieve the size separation, our microfluidic device is based on pinched-flow fractionation. A model virus, cowpea chlorotic mottle virus, was used and spiked to porcine semen samples. With the proposed microfluidic chip and the optimized flow parameters, at least 84 ± 4% of the model viruses were removed from the semen. The remaining virus contamination is caused by the model virus adhering to spermatozoa instead of the separation technique. The spermatozoa recovery was 86 ± 6%, which is an enormous improvement in yield compared to existing virus removal techniques.

Microfluidic single sperm entrapment and analysis.

Selection of healthy spermatozoa is of crucial importance for the success rates of assisted reproduction technologies (ART) such as in vitro fertilization and intra-cytoplasmic sperm injection. Although sperm selection for ART procedures is predominantly based on sperm motility, successful fertilization is not predicted by good motility alone. For example, sperm characteristics such as the acrosome state and DNA integrity have shown significant impact on ART outcome. Although fertilization can be achieved with a single spermatozoon of high quality, current quality assessments are population-based and do not allow investigation of multiple sperm characteristics on a single spermatozoon simultaneously. In order to study sperm cells on the single cell level, we designed and characterized a PDMS microfluidic platform that allows single sperm entrapment. After spatially confining individual sperm cells within microfluidic cell traps, the cell viability, chromosomal content and acrosome state were studied. This platform is suitable for the analysis of individual sperm cells, which could be exploited for (non-invasive) sperm analysis and selection by impedance or Raman spectroscopy.