Refractive Index–Correlated Pseudocoloring for Adaptive Color Fusion in Holotomographic Cytology

Conventional bright-field (BF) cytology of thyroid fine-needle aspiration biopsy (FNAB) suffers from staining variability and limited subcellular contrast. Here, we present a refractive index-correlated pseudocoloring (RICP) framework that integrates quantitative refractive index (RI) maps obtained by holotomography (HT) with color BF images to enhance subcellular visualization and image interpretability. A digital micromirror device (DMD)-based HT system combined with an RGB LED illumination module enabled sequential acquisition of RI tomograms and BF images from Papanicolaou (PAP)-stained thyroid samples over the same field of view. The RICP algorithm adaptively embeds RI-derived structural information into a complementary hue channel while preserving the overall BF appearance. Applied to benign and malignant thyroid clusters, RICP highlighted subcellular features such as nucleoli, lipid droplets, and nuclear irregularities. Hue-saturation analysis revealed differences between benign and malignant clusters, while complementary image-fusion metrics and delentropy analysis both indicated systematic changes in structural image content after fusion. An ablation analysis further showed that the RI-derived input contributes structural content beyond that obtained when the RI map is replaced by an inverted Value channel of the BF image in HSV space within the same pseudocoloring framework. These results indicate that RICP provides a perceptually grounded approach for correlative multimodal visualization that bridges conventional color cytology and quantitative optical imaging.