Electrospun poly(lactic acid) biopolymer fibers functionalized with Y-sperm-specific scFv enable immunoaffinity-driven bovine sperm sexing

Assisted reproductive technologies increasingly demand efficient approaches for sperm sexing in livestock, ideally compatible with scalable manufacturing and reduced resource use. This study serves as a proof-of-concept for a biocompatible electrospun poly (lactic acid) (PLA) biomaterial platform functionalized with a recombinant single-chain variable fragment antibody specific to bovine Y-sperm (BY-scFv), enabling selective separation of bovine Y-sperm from X-sperm populations. Electrospun mats were prepared by selection PLA concentration (6–10% w/w) and deposited solution volume (10–20 mL), and were evaluated based on electrospinning performance, fiber morphology, thickness, water-accessible porosity and water uptake capacity. A mat fabricated at 8% PLA with a 15 mL deposited volume was selected for subsequent functionalization and separation experiments. The presence and retention of immobilized BY-scFv after EDC/NHS coupling were supported by ELISA, FT‑IR, SEM, and fluorescence labeling. Using a syringe‑packed format, the functionalized mats enabled selective sperm capture and produced sex sperm ratio in different fractions, with maximum values of 71.4% X‑sperm in the supernatant fraction and 76.1% Y‑sperm in the eluted fraction. These results indicate that antibody‑functionalized electrospun PLA scaffolds can serve as a biopolymer-based platform for sperm immunoaffinity processing, with potential for further optimization and translational evaluation.