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High-fidelity color image reconstruction for holographic microscopy

6 May | By Biophotonics.World
High-fidelity color image reconstruction for holographic microscopy
Figure 1. Schematics of the absorbance spectrum estimation-based high-fidelity colorization technique for holographic microscopy.

Digital holographic microscopy (DHM) has been widely applied to biomedical imaging applications due to its digital refocusing capability, simple optical configuration, and reconstruction of both the amplitude and phase information of the sample. However, holographic microscopy systems in general present some challenges for high fidelity color reproduction due to the use of narrow-band and coherent illumination. This is especially important for pathology applications, where the colorimetric contrast introduced by various histological stains is ubiquitous in making reliable and consistent diagnoses. 

Researchers at UCLA have developed a spectrum estimation-based colorization technique, which enables holographic microscopy to reconstruct high fidelity color images of samples with simultaneous illumination at three different wavelengths, optimized for a given tissue and stain combination. This technique, termed as absorbance spectrum estimation-based colorization (ASEC), makes it possible to image the specimen with substantially reduced number of measurements, in comparison to previous holographic color imaging approaches. In this approach (see Figure 1), the raw hologram is initially demultiplexed into three color channels, representing the absorbance of the specimen at three illumination wavelengths. Then, the absorbance spectrum of the sample in the visible band is estimated and finally, a high-fidelity color image of the sample is reconstructed by projecting the estimated spectrum onto the RGB color space using the standard color matching functions. Tested with 25 pathology slides using different tissue and stain combinations, this technique showed a significant reduction on color aberrations compared to directly using the demultiplexed holographic images into the RGB color channels of the reconstruction.

This research was published in the Journal of Biophotonics, an open access journal of the Wiley Online Library. The research was led by Dr. Aydogan Ozcan, the Chancellor's Professor of electrical and computer engineering at the UCLA Henry Samueli School of Engineering and Applied Science, and an associate director of the California NanoSystems Institute (CNSI), along with Yibo Zhang, a graduate student, Tairan Liu, a graduate student, and Dr. Yair Rivenson, an assistant adjunct Professor of electrical and computer engineering at the UCLA. 

“The absorbance spectrum estimation-based colorization technique can significantly improve the color accuracy of holographic microscopy by building up a library of the statistical color information for each type of tissue and stain combination. This one-time effort eliminates the need to densely sample the visible band with a hyperspectral imaging technique.” said Dr. Ozcan. This study provides a practical guide and a database to achieve accurate-color holographic imaging of pathology slides, spanning different dyes and tissue types.

The research of Ozcan Lab is supported by the Koc Group, NSF and HHMI.  

Author: Aydogan Ozcan


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