Autofluorescence-Raman Spectroscopy for Ex Vivo Mapping Colorectal Liver Metastases and Liver Tissue

Identifying colorectal liver metastases (CRLM) during liver resection could assist in achieving clear surgical margins, which is an important prognostic variable for both disease-free and overall survival. The aim of this study was to investigate the effect of auto-fluorescence (AF) and Raman spectroscopy for ex vivo label-free discrimination of CRLMs from normal liver tissue. Secondary aims include exploring options for multimodal AF-Raman integration with respect to diagnosis accuracy and imaging speed on human liver tissue and CRLM. Liver samples were obtained from patients undergoing liver surgery for CRLM who provided informed consent (15 patients were recruited). AF and Raman spectroscopy was performed on CRLM and normal liver tissue samples and then compared to histology. AF emission spectra demonstrated that the 671 nm and 775/785 nm excitation wavelengths provided the highest contrast, as normal liver tissue elicited on average around eight-fold higher AF intensity compared to CRLM. The use of the 785 nm wavelength had the advantage of enabling Raman spectroscopy measurements from CRLM regions, allowing discrimination of CRLM from regions of normal liver tissue eliciting unusual low AF intensity, preventing misclassification. Proof-of-concept experiments using small pieces of CRLM samples covered by large normal liver tissue demonstrated the feasibility of a dual-modality AF-Raman for detection of positive margins within few minutes. AF imaging and Raman spectroscopy can discriminate CRLM from normal liver tissue in an ex vivo setting. These results suggest the potential for developing integrated multimodal AF-Raman imaging techniques for intraoperative assessment of surgical margins.