Viewing Cancer in a New Light: Multimodal Label-Free Specific Chemical Imaging to Distinguish Healthy Cell Tissue from Invasive Carcinoma

Abstract

Microscopy has always been an important tool when determining whether a biopsy contains invasive carcinomas. For this purpose, light microscopy is often used due to its simplicity in construction and operation. The downside to this is the time lost in preparing biopsies before imaging. Typically, the process of obtaining a biopsy, molding, sectioning, mounting and staining of the biopsy on microscopy glass slides and finally imaging the biopsy takes between 6-10 days. If the preparatory steps could be eliminated, valuable time would be freed. Coherent anti-Stokes Raman Scattering (CARS) microscopy offers a possible solution to this obstacle. CARS is a non-linear optical microscopy technique that allows label-free specific imaging of chemical moieties present in a sample. Specifically, lipids (rich in CH2-moieties) and protein (rich in CH-moieties), as well as water, are easily imaged using CARS. A system capable of performing CARS microscopy will generally also be capable of doing Second-Harmonic Imaging Microscopy (SHIM). This technique uses second-harmonic generation (SHG) to achieve frequency doubling of the incident light. SHIM enables imaging of biological structures, which possess the characteristics of being both highly polarizable and ordered on medium to long length scales. Specifically collagen, which is present especially in the basement membrane, is very capable of producing a second-harmonic signal. By combining CARS, SHIM and multi-photon excitation of auto-fluorescent molecules, it is possible to acquire multimodal micrographs of epithelial tissue samples. The CARS measurements gives a unique insight into cell organization in tissue by clearly visualizing the cell membrane bilayer and lipid distribution, while the SHIM reveals the location of the basement membrane and underlying connecting tissue. Our results show that it is possible, in an unlabeled sample, to differentiate between healthy tissue samples and samples that display varying degrees of dysplasia or invasive carcinomas. This opens up for the possibility for future fast bedside cancer diagnostics using CARS microscopy.