25 April 2008 Quantitative phase restoration in differential interference contrast (DIC) microscopy
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Abstract
Phase contrast imaging is a specific technique in optical microscopy that is able to capture the minute structures of unlabeled biological sample from contrast generated in the variations of the object's refractive index. It is especially suitable for living cells and organisms that are hardly visible under conventional light microscopy as they barely alter the intensity and only introduce phase shifts in transmitted light. Optical phase imaging provides sensitivity to measure optical path length (OPL) differences down to nanometers, which has great potential in biomedical applications from examining both topological and three-dimensional biophysical properties of cells and organisms. Conventional DIC microscopy with partially coherent light source is a very powerful technique for phase imaging, and is able to yield higher lateral resolution compared to other interferometric phase imaging methods. However, it is inherently qualitative and the information obtained is a phase-gradient image rather than a true linear mapping of OPL differences. This hinders its further application as it is difficult to infer the results directly. Work has been done previously to obtain the quantitative phase information out of DIC. However, some of these methods not only involves costly hardware modification but also complicated computation. Here we investigate another approach that combines the correlation of light intensity and phase with polarization-modulated differential interference contrast (DIC) microscopy. The required hardware modification is simple, and numerically solving the relationship of light propagation in a series of through-focus DIC images allows phase information to be restored from phase gradients in two-dimensional planes. Quantitative phase information in three-dimensional space can then be reconstructed from 3D rendering of the calculated phase images. Initial results of application in biological cells are also demonstrated.
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Shan Shan Kou, Shan Shan Kou, Colin J. R. Sheppard, Colin J. R. Sheppard, } "Quantitative phase restoration in differential interference contrast (DIC) microscopy", Proc. SPIE 7000, Optical and Digital Image Processing, 700005 (25 April 2008); doi: 10.1117/12.780912; https://doi.org/10.1117/12.780912
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