Label-free ex vivo pathological studying on breast cancer tissue by using nonlinear multiphoton and micro-Raman spectral imaging

Abstract

Raman spectral and multiphoton microscopy (MPM) were utilized in this study to understand the compositional and structural characteristics of the breast tumor microenvironment. Combining these techniques allowed us to address the pathological features of ex vivo healthy and invasive ductal carcinoma (IDC) breast tissue sections from different perspectives. MPM-based two-photon excited fluorescence and second-harmonic generation offer significant advantages for large-area structural tissue imaging. Raman spectral imaging identified the compositional information from a spectroscopic perspective and visualized the specific distribution patterns of biomedical components using a multivariate imaging method (K-mean cluster analysis, KCA). The MPM results revealed the loss of collagen bundles, malignant cell invasion, basement membrane damage, and blood vessel growth in IDC. Additionally, variations in tryptophan, nucleic acid, carotenoid, protein, and lipid content were found in different tissues. The reconstructed Raman images illustrate the morphological alterations of these contents and the edge contours of the cancerous tissue. The acquired multimodal optical results, which extracted complementary information of a sample based on different contrast mechanisms, provided new label-free pathological interpretations of the tumor microenvironment and offered the experimental foundations for developing multimode optical modalities as an immediate intraoperative diagnosis method.