A compact integrated platform for real-time live-cell imaging and photodynamic therapy: design and experimental demonstration

Real-time integration of live-cell imaging and photodynamic therapy (PDT) addresses a critical experimental gap between treatment delivery and longitudinal observation of cellular responses. We present a compact, incubator-compatible platform that integrates inverted bright-field microscopy, automated X–Y scanning, and a uniform 630-nm PDT illumination module within a single experimental workflow, allowing the same cell culture to undergo repeated imaging and PDT without transfer between separate incubation, treatment, and imaging setups. The optical module provides a 1.5 mm² field of view at 2.2 μm resolution. The integrated mechanical and electronic architecture enables automated scanning and repeatable acquisition of predefined imaging regions. To experimentally demonstrate the dual functionality of the platform, A375 melanoma cells treated with 5-aminolevulinic acid (5-ALA, 10–500 µM) underwent fractionated PDT and longitudinal imaging over 72 h. Complementary MTT assays confirmed a time- and dose-dependent decline in viability, while longitudinal image analysis quantified treatment-associated changes in cell confluency and individual-cell morphology over time. In addition, ROI overlap analysis demonstrated substantially improved preservation of imaging regions compared with a conventional workflow involving repeated manual sample transfer. By integrating treatment delivery and long-term imaging within the incubator, the platform provides an experimental infrastructure for continuous observation of the same cell populations during prolonged PDT protocols and other time-dependent cell-based studies.