White light phase shifting interferometric microscopy with whole slide imaging for quantitative analysis of biological samples

Abstract

In this paper, we demonstrate the white light phase shifting interferometer employed as whole slide scanner and phase profiler for determining qualitative and quantitative information over large field-of-view (FOV). Experiments were performed on human erythrocytes and MG63 Osteosarcoma cells. Here, we have recorded microscopic images and phase shifted white light interferograms simultaneously in a stepped manner. Sample slide is translated in transverse direction such that there exists a correlation between the adjacent frames, and they were stitched together using correlation functions. Final stitched image has a FOV of 0.24 Â 1.14 mm with high resolution ~0.8 μm. Circular Hough transform algorithm is implemented to the resulting image for cell counting and five-step phase shifting algorithm is utilised to retrieve the phase profiles over a large FOV. Further, this technique is utilised to study the difference between normal and anaemic erythrocytes. Significant changes are observed in anaemic cells as compared to normal cells.

White blood cells segmentation and classification using a random forest and residual networks implementation

“…14,[22][23][24] After classification, the resulting binary image was submitted to a morphological opening (an erosion followed by dilation) by a (5, 5) kernel of ellipsoidal format, followed by a connected components analysis (CCA) with statistics, both functions available on OpenCV library. The opening kernel was an ellipsoidal one, with a size of (5,5), and the parameters of connectivity and ltype, for CCA, as being to 8 and CV 32S, respectively.…”

“…2 Other interesting instruments have been developed in recent years to perform cell counting in stained slides of blood smears, to overcome the highly time-consuming task of cell count in a standard brightfield microscope. [3][4][5] These systems are routinely called whole slide imaging systems. 6 With the development of such interesting systems, the need for intelligent software for cell count has significantly increased.…”

Quantitative Phase Imaging: Recent Advances and Expanding Potential in Biomedicine

“…20 These dual path interferometers were followed by common-path interferom- ). The number of visible interference fringes generated depends on the optical setup and coherence of the light source, with low coherence light sources (e.g., white light) 24 producing fewer visible fringes than highly coherent lasers. 25 An in-focus interferogram is then used to generate the phase image, and, in phase shifting interferometry, the reference and sample path lengths are adjusted in steps, e.g., with a piezo, to shift the fringes by a fraction of a wavelength.…”

Multispectral polarization microscopy of different stages of human oral tissue: A polarization study

“…The recent advancements in cancer diagnosis and treatments are exceptionally remarkable but the increment in the survival rate again is not very promising in last 40-50 years [3,4]. Early diagnosis is the primary requirement of healthcare amenities [5]. From past few decades number of optical techniques has been explored in medical diagnostics imaging but some of the labelling optical techniques like fluorescence microscopy requires staining of the sample with a specific dye which can hinder the cell functionality and the sample preparation is a little bit more time consuming as compared to the label-free approaches [6,7].…”

“…The detailed study of polarimetric changes in oral tissue with multiple wavelengths from a single white light source by separating its respective multiple channels digitally is performed. A set of six intensity images has been recorded by rotating the plane of polarization of different polarizing optical components and these white light images are then digitally separated out into their respective RGB components [5]. These RGB intensity images are further processed for stokes mapping and polarization imaging like degree of polarization, fast axis orientation, retardance, linear birefringence is achieved at these wavelengths to have further insight into the characteristics of light and their interaction with the samples.…”