Fast stimulated Raman and second harmonic generation imaging for intraoperative gastro-intestinal cancer detection

Abstract

Conventional haematoxylin, eosin and saffron (HES) histopathology, currently the ‘gold-standard’ for pathological diagnosis of cancer, requires extensive sample preparations that are achieved within time scales that are not compatible with intra-operative situations where quick decisions must be taken. Providing to pathologists a close to real-time technology revealing tissue structures at the cellular level with HES histologic quality would provide an invaluable tool for surgery guidance with evident clinical benefit. Here, we specifically develop a stimulated Raman imaging based framework that demonstrates gastro-intestinal (GI) cancer detection of unprocessed human surgical specimens. The generated stimulated Raman histology (SRH) images combine chemical and collagen information to mimic conventional HES histopathology staining. We report excellent agreements between SRH and HES images acquire on the same patients for healthy, pre-cancerous and cancerous colon and pancreas tissue sections. We also develop a novel fast SRH imaging modality that captures at the pixel level all the information necessary to provide instantaneous SRH images. These developments pave the way for instantaneous label free GI histology in an intra-operative context.

Protein and Lipid Mass Concentration Measurement in Tissues by Stimulated Raman Scattering Microscopy

“…For example, NoRI measurements of protein density may provide a label-free way to visualize condensation of proteins from liquid-liquid phase separation. The concentration of protein and lipid can serve as a histological marker 20,21 .…”

“…For example, NoRI measurements of protein density may provide a label-free way to visualize condensation of proteins from liquid-liquid phase separation. The concentration of protein and lipid can serve as a histological marker 20,21 . Unlike conventional histological images that may have different staining intensities from batch to batch, NoRI histology takes advantage of the protein and lipid concentrations, which are intrinsic properties of tissues that can be directly compared between different samples.…”

Stimulated Raman histology: one to one comparison with standard hematoxylin and eosin staining

“…Recently, stimulated Raman histology (SRH) [1] has emerged as a label-free microscopy technique, using stimulated Raman scattering (SRS) [2], that produces virtually stained images that imitate standard eosin and hematoxylin (H&E) staining. SRH was first demonstrated on brain cryogenic sections [3] and later on applied to the gastrointestinal tract (GI) [4]. In the GI, SRS had to be combined with second harmonic generation (SHG) to concurrently extract the collagen distribution whose role is crucial in both cancer development and progression [5,6].…”

“…This information is sufficient to generate H&E like images [3]. Adding the collagen fibrils organisation using SHG allows to generate HES (hematoxylin, eosin, saffron) images where both cell bodies, cell nuclei and fibrotic surrounding tissue are revealed for cancer stage and aggressiveness assessments [4]. Contrary to standard histology, SRH does not require any labelling and can perform virtual HES sections from unprepared fresh bulk biopsy samples opening the road for fast intra-operative histology to assist in surgical and decision making.…”

“…Contrary to standard histology, SRH does not require any labelling and can perform virtual HES sections from unprepared fresh bulk biopsy samples opening the road for fast intra-operative histology to assist in surgical and decision making. So far earlier SRH reports [3,4] evaluated the agreement between SRH and H&E images on the same tissue but at different locations or on adjacent sections, but the one-to-one comparison on identical samples is still missing. In this paper we demonstrate for the first time the quasi-perfect matching between SRH and H&E images on identical cryogenic slides for both normal and cancerous tissues.…”