<p>Background: Traditional pathological techniques for diagnosing liver cancer are limited by the need for examination of multiple points. Preoperative treatment can complicate the evaluation by causing tumor necrosis. The evolving treatment landscape requires precise, integrated, and comprehensive management to address the limitations, specifically in localization, sampling, and speed. This study aims to innovatively explore the use of indocyanine green (ICG) fluorescence imaging to guide accurate and rapid localization of residual cancer foci, facilitating rapid pathological sampling and evaluation, termed as rapid pathological technology. Patients and Methods: In this prospective and observational study, patients undergoing surgical resection for HCC were enrolled. Traditional evaluation involved specimen fixation, sampling, sectioning, and microscopic observation, while rapid evaluation entailed fluorescence-guided rapid pathological sampling and evaluation post-dissection. Results: A total of 37 HCC patients were included, and 103 samples of varying fluorescence intensity were collected. Analysis revealed a significant negative correlation between fluorescence intensity and tumor necrosis ratio (ρ = −0.89, p &lt; 0.001). Furthermore, logistic regression modeling yielded an AUC of 0.947, with a specificity of 0.900 and sensitivity of 0.930, indicating strong discriminatory power for tumor necrosis status. An optimal cut-off fluorescence intensity of ≥ 20.5 was identified for predicting partial tumor necrosis. The Youden index is 0.830. Decision curve analysis demonstrated the clinical utility of the prediction model for guiding treatment decisions regarding tumor necrosis. Conclusions: Rapid pathological evaluation using ICG fluorescence-guided resection specimens in liver cancer is feasible, enabling effective localization of residual cancer foci and enhancing the accuracy and efficiency of pathological sampling.</p><p>Trial registration: ChiCTR2400082814</p>