Selecting viable sperm for successful fertilisation and optimal developmental competence is an unmet challenge in reproductive medicine. There are limitations of methods based solely on motility and morphology criteria and this has prompted innovative approaches. Advances in understanding of the immunobiology of female reproductive tract sperm selection shows the role of immune cells in sperm selection. Knowledge of the relevant biomolecules provides novel avenues to develop surface chemistry-based approaches to emulate the molecular interactions between suboptimal sperm and immune cells. In pursuit of this objective, we have investigated plasma polymerisation approaches to recapitulate immune-mediated sequestration of suboptimal sperm on functionalised glass surfaces. We applied a plasma polymerised polyoxazoline (PPOx) film to glass using 2-methyl-2-oxazoline monomer. The PPOx film enabled the covalent binding of antibodies reacting with suboptimal sperm. Samples of human donor sperm (n=3-5) prepared by a standard ‘swim-up’ technique, or neat semen, were introduced to the activated surface and the rate of sperm attachment was measured. The characteristics of recovered unattached sperm were then assessed by flow cytometry to measure proportions of viable sperm (propidium iodide), apoptotic sperm (Annexin V expression), as well as levels of reactive oxygen species (CellROX Green), and sperm DNA fragmentation (HALO-sperm). We then applied similar surface functionalisation to antibody-coated glass channel-slides and showed that sperm recovered after introduction of neat semen exhibit superior functional characteristics compared with sperm prepared by standard swim-up or uncoated channelled slides. Sperm recovered from functionalised channel-slides exhibited a significantly lower Annexin V+ sperm subpopulation (6.3%±1.1%) compared with standard swim-up or uncoated channel slides (19%±4.1% and 15%±3.1%, respectively). The proportion of sperm exhibiting DNA fragmentation was also significantly decreased around (5.7%±1.2%) compared with the swim-up (11.5%±3.1%) samples.
In conclusion, our findings demonstrate the efficacy of surface functionalised antibody-coated channel-slides and potential in preparing high-quality sperm for applications in ART.