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Analytical Cellular Pathology
Volume 36, Issue 5-6, Pages 163-172
http://dx.doi.org/10.3233/ACP-140089

Simultaneous Dual-Color Fluorescence Microscope: A Characterization Study

Zheng Li,1 Xiaodong Chen,2 Liqiang Ren,1 Jie Song,1,3 Yuhua Li,1 Bin Zheng,1 and Hong Liu1

1School of Electrical and Computer Engineering, University of Oklahoma, Norman, OK, USA
2College of Precision Instruments and Opto-electronics Engineering, Tianjin University, Tianjin, China
3Department of Biology, Mudanjiang Medical University, Mudanjiang City, China

Received 8 January 2014; Accepted 11 May 2014

Copyright © 2013 Hindawi Publishing Corporation and the authors. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Background: High spatial resolution and geometric accuracy is crucial for chromosomal analysis of clinical cytogenetic applications. High resolution and rapid simultaneous acquisition of multiple fluorescent wavelengths can be achieved by utilizing concurrent imaging with multiple detectors. However, such class of microscopic systems functions differently from traditional fluorescence microscopes.

Objective: To develop a practical characterization framework to assess and optimize the performance of a high resolution and dual-color fluorescence microscope designed for clinical chromosomal analysis.

Methods: A dual-band microscopic imaging system utilizes a dichroic mirror, two sets of specially selected optical filters, and two detectors to simultaneously acquire two fluorescent wavelengths. The system’s geometric distortion, linearity, the modulation transfer function, and the dual detectors’ alignment were characterized.

Results: Experiment results show that the geometric distortion at lens periphery is less than 1%. Both fluorescent channels show linear signal responses, but there exists discrepancy between the two due to the detectors’ non-uniform response ratio to different wavelengths. In terms of the spatial resolution, the two contrast transfer function curves trend agreeably with the spatial frequency. The alignment measurement allows quantitatively assessing the cameras' alignment. A result image of adjusted alignment is demonstrated to show the reduced discrepancy by using the alignment measurement method.

Conclusions: In this paper, we present a system characterization study and its methods for a specially designed imaging system for clinical cytogenetic applications. The presented characterization methods are not only unique to this dual-color imaging system but also applicable to evaluation and optimization of other similar multi-color microscopic image systems for improving their clinical utilities for future cytogenetic applications.