Nano TiO2 and direct black marking system for precise localization in intraoperative pathological diagnosis of small tumors

Concordance between intraoperative frozen section (FS) diagnosis and permanent section diagnosis is essential for quality assurance in pathology. However, in cases involving small lesions (≤ 1 cm), tissue dehydration following FS processing often reduces visibility and hinders localization, leading to diagnostic discrepancies between FS and permanent sections. This study introduces nano-titanium dioxide (nano-TiO2) and Direct Black (DB) for intraoperative tissue marking to enhance tumor identification. Surgical specimens from lung, thyroid, and breast cancers (January 2024-March 2025) with tumors ≤ 1 cm were included. Following FS diagnosis, tissues were marked using methylene blue, carbon nanoparticle suspension, nano-TiO2, and DB. Hematoxylin and eosin (H&E) staining, immunohistochemistry, and reverse transcription polymerase chain reaction were applied to evaluate visibility, stability, and diagnostic impact. Nano-TiO2 and DB exhibited excellent visibility and tissue adherence across all three tumor types without interfering with H&E, immunomarkers (PD-L1, TTF-1, and HER2), or molecular pathology assays (EGFR and BRAF). Compared with unmarked specimens, marked tissues demonstrated higher concordance between FS and permanent diagnosis: Lung cancer (95% versus 84.38%), thyroid cancer (100% versus 81.25%), and breast cancer (100% versus 85.71%) (all p < 0.05). These findings indicate that nano-TiO2 and DB are safe, effective, and significantly improve intraoperative localization and diagnostic accuracy of small lesions. The method offers potential for workflow standardization and broader use in pathological practice.