Quantitative phase and stain concentration imaging using single-shot color differential interference contrast microscopy for photonic histopathology

Abstract

Differential interference contrast (DIC) microscopy is widely used in biomedical studies for imaging transparent samples due to its excellent lateral resolution and axial discrimination. Aided by computation, quantitative DIC, which inherits the advantages of the DIC approach, has emerged as one attractive quantitative phase imaging modality. For the routine Hematoxylin-Eosin (H and E) stained histopathology specimens, the concentration maps of Hematoxylin and Eosin carry additional diagnostic information because of the negatively charged affinity of acidic eosin for cytoplasmic proteins and the positively charged affinity of basic hematoxylin for nuclear structures. It will be desirable to quantify the stain concentration maps in addition to the quantitative phase for H and E stained specimens. Here we present Quantitative phase and stain concentration imaging using Single-shot color differential interference contrast (C-DIC) microscopy for photonic histopathology for hematoxylin and eosin-stained tissue sections. The recorded image is first normalized by the corresponding image for blank slides under identical imaging conditions. The absorbance contributions from the H and E stains and the contributions from the phase gradient of the H and E stained specimen are unmixed from the recorded color-DIC image. The concentrations of the H and E stains are computed based on their respective extinction coefficients at the R: G: B: channels of the color camera. The quantitative phase map of the specimen is obtained from the results of the phase gradient. We apply C-DIC to image a cohort of routine prostate cancer specimens. The accuracy of diagnosis of prostate cancer improves with the addition of stain concentration maps compared to the quantitative phase alone.