Purpose: Neurotrophins such as brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT3) exhibit pro-survival and homeostatic properties, but their clinical translation is limited by protein instability and rapid clearance. We evaluated a PEGylated human serum albumin (HSA) nanoparticle system for BDNF/NT3 co-delivery, focusing on physicochemical stability, ocular biodistribution in the rabbit eye, intracellular protein delivery, and protection against oxidative stress–associated cellular damage in human cells. Materials and Methods: PEGylated HSA–BDNF–NT3 nanoparticles with nominal neurotrophin concentrations of 5 µg/mL (NeO5) or 10 µg/mL (NeO10) were generated by spontaneous self-assembly and characterized using multiangle dynamic light scattering, electrophoretic light scattering, and atomic force microscopy. In vivo performance was assessed after intravitreal injection in rabbits by enzyme-linked immunosorbent assay (ELISA)–based protein quantification and exploratory reverse transcription quantitative polymerase chain reaction (RT-qPCR) profiling of survival-, proliferation-, and apoptosis-related genes. Functional delivery was examined in sodium iodate-stressed ARPE-19 and 6-hydroxydopamine–stressed retinoic acid-differentiated SH-SY5Y cells using ELISA assays, JC-1 analysis, Annexin V/ propidium iodide flow cytometry, high-performance liquid chromatography for malondialdehyde quantification, and RT-qPCR. Results: Both formulations formed stable, spherical nanoparticles (5.9– 54.2 nm) with low polydispersity index (≈ 0.18) and preserved colloidal integrity over 28 days. In vivo, BDNF was detectable in ocular tissues up to 72 h and RT qPCR did not reveal a coordinated pro-apoptotic response under the tested conditions. In vitro, nanoparticle treatment significantly increased intracellular BDNF and NT3 levels, improved viability, reduced apoptotic cell fractions, and markedly decreased lipid peroxidation, particularly for NeO10. Increased tropomyosin receptor kinase B TRKB and cAMP response element-binding protein CREB expression provided supportive molecular evidence consistent with neurotrophin-related cellular responses. Conclusion: PEGylated HSA nanoparticles enable stable neurotrophin loading, efficient intracellular delivery, and attenuation of oxidative stress–induced cytotoxicity. These findings support further development of albumin-based nanocarriers for translational nanomedicine applications.