Quantification of hepatic steatosis in chronic liver disease using novel automated method of second harmonic generation and two-photon excited fluorescence

The presence of hepatic steatosis (HS) is an important histological feature in a variety of liver disease. It is critical to assess HS accurately, particularly where it plays an integral part in defining the disease. Conventional methods of quantifying HS remain semi-quantitative, with potential limitations in precision, accuracy and subjectivity. Second Harmonic Generation (SHG) microscopy is a novel technology using multiphoton imaging techniques with applicability in histological tissue assessment. Using an automated algorithm based on signature SHG parameters, we explored the utility and application of SHG for the diagnosis and quantification of HS. SHG microscopy analysis using GENESIS (HistoIndex, Singapore) was applied on 86 archived liver biopsy samples. Reliability was correlated with 3 liver histopathologists. Data analysis was performed using SPSS. There was minimal inter-observer variability between the 3 liver histopathologists, with an intraclass correlation of 0.92 (95% CI 0.89–0.95; p < 0.001). Good correlation was observed between the histopathologists and automated SHG microscopy assessment of HS with Pearson correlation of 0.93: p < 0.001. SHG microscopy provides a valuable tool for objective, more precise measure of HS using an automated approach. Our study reflects proof of concept evidence for potential future refinement to current conventional histological assessment.