We present for the first time one-to-one correspondence between standard hematoxylin/eosin (H&E) stained tissue sections and stimulated Raman histology (SRH)-a label-free technique in which stimulated Raman scattering (SRS) and second harmonic generation (SHG) are combined to generate virtual H&E images. Experiments were performed on both human thin cryogenic slides from the gastrointestinal tract (GI) and thick freshly excised biopsies from endoscopic surgery. Results on cryogenic slides evidenced an excellent agreement between SRH and H&E images while the ones on biopsies established the relevance of SRH for rapid intraoperative histology to assist in surgical decision making.

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.

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