Label‐free multiphoton microscopic imaging as a novel real‐time approach for discriminating colorectal lesions: A preliminary study

Background and Aim Although various endoscopic technologies have been developed to increase the visual diagnostic accuracy of colorectal precancerosis and early carcinoma, the biopsy‐dependent pathology still remains the golden standard. During routine endoscopy, real‐time optical histological diagnosis is desired. Without fluorescent label, multiphoton microscopy (MPM) imaging directly reveals live cellular morphology and tissue microenvironment based on intrinsic two‐photon excited fluorescence and second harmonic generation signals. Its high‐imaging resolution and performance are comparable with the histopathology. We thus aimed to initially investigate the original features of colorectal diseases under MPM and evaluate its potential for real‐time diagnosis. Methods Experimental and diagnostic cohorts were designed. Multiphoton images of 40 ex vivo fresh tissues confirmed pathologically of colorectal normal tissues, hyperplastic polyps, adenomas, and adenocarcinomas were collected. Features were recorded to establish diagnostic standards with MPM. For the second cohort with 92 fresh tissues, we distinguished the various colorectal diseases with conclusive MPM features. Results Through the investigation, the colorectal diseases were presented differences in the crypt opening, gland structure, epithelial cells, and collagen fibers. With the typical features, we preliminarily tested the diagnostic efficiency and found that its sensitivity for distinguishing normal, hyperplastic polyps, adenoma and adenocarcinoma was 88.89%, 76.47%, 83.33%, and 97.92%, while the specificity was 99.32%, 94.00%, 94.92%, and 94.12%, respectively. Conclusion The real‐time multiphoton microscopic imaging can be effective to identify the colorectal lesions with high resolution. Via integrating with the endoscopes in the future, it could promote precise optical diagnosis in clinics.