Cancer, a prominent cause of death, presents treatment challenges, including high dosage requirements, drug resistance, poor tumour penetration and systemic toxicity in traditional chemotherapy. Photodynamic therapy, using photosensitizers like rose bengal (RB) with a green laser, shows promise against breast cancer cells in vitro. However, the hydrophilic RB struggles to efficiently penetrate the tumour site due to the unique clinical microenvironment, aggregating around rather than entering cancer cells. In this study, we have synthesized and characterized RB-encapsulated chitosan nanoparticles with a peak particle size of ~200 nm. These nanoparticles are readily internalized by cells and, in combination with a green laser (λ = 532 nm) killed 94–98% of cultured human breast cancer cells (MCF-7) and prostate cancer cells (PC3) at a low dosage (25 μg/mL RB-nanoparticles, fluence ~126 J/cm2, and irradiance ~0.21 W/cm2). Furthermore, these nanoparticles are not toxic to cultured human normal breast cells (MCF10A), which opens an avenue for translational applications.