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International Test Results for Objective Lens Quality, Resolution, Spectral Accuracy and Spectral Separation for Confocal Laser Scanning Microscopes

Published online by Cambridge University Press:  08 October 2013

Richard W. Cole ,
Marc Thibault ,
Carol J. Bayles ,
Brady Eason ,
Anne-Marie Girard ,
Tushare Jinadasa ,
Cynthia Opansky ,
Katherine Schulz  and
Claire M. Brown
Richard W. Cole
Affiliation:
New York State Department of Health, Wadsworth Center, P.O. Box 509, Albany, NY 12201, USA
Marc Thibault
Affiliation:
Montreal General Hospital, C9, 1650 Cedar, Montreal H3G 1A4, Canada
Carol J. Bayles
Affiliation:
Life Sciences Core Laboratory Center, Cornell University, Weill Hall, Ithaca, NY 14853, USA
Brady Eason
Affiliation:
Life Sciences Complex Advanced BioImaging Facility (ABIF), McGill University, 3649 Prom, Sir William Osler, Bellini Building, Room 137, Montreal, QC H3G 0B1, Canada
Anne-Marie Girard
Affiliation:
Center for Genome Research and Biocomputing, Oregon State University, 3021 Agriculture and Life Sciences Building, Corvallis, OR 97331, USA
Tushare Jinadasa
Affiliation:
Life Sciences Complex Advanced BioImaging Facility (ABIF), McGill University, 3649 Prom, Sir William Osler, Bellini Building, Room 137, Montreal, QC H3G 0B1, Canada
Cynthia Opansky
Affiliation:
Blood Center of Wisconsin, Blood Research Institute, 8733 Watertown Plank Road, Milwaukee, WI 53226, USA
Katherine Schulz
Affiliation:
Blood Center of Wisconsin, Blood Research Institute, 8733 Watertown Plank Road, Milwaukee, WI 53226, USA
Claire M. Brown*
Affiliation:
Life Sciences Complex Advanced BioImaging Facility (ABIF), McGill University, 3649 Prom, Sir William Osler, Bellini Building, Room 137, Montreal, QC H3G 0B1, Canada
*
*Corresponding author.claire.brown@mcgill.ca
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Abstract

As part of an ongoing effort to increase image reproducibility and fidelity in addition to improving cross-instrument consistency, we have proposed using four separate instrument quality tests to augment the ones we have previously reported. These four tests assessed the following areas: (1) objective lens quality, (2) resolution, (3) accuracy of the wavelength information from spectral detectors, and (4) the accuracy and quality of spectral separation algorithms. Data were received from 55 laboratories located in 18 countries. The largest source of errors across all tests was user error which could be subdivided between failure to follow provided protocols and improper use of the microscope. This truly emphasizes the importance of proper rigorous training and diligence in performing confocal microscopy experiments and equipment evaluations. It should be noted that there was no discernible difference in quality between confocal microscope manufactures. These tests, as well as others previously reported, will help assess the quality of confocal microscopy equipment and will provide a means to track equipment performance over time. From 62 to 97% of the data sets sent in passed the various tests demonstrating the usefulness and appropriateness of these tests as part of a larger performance testing regiment.

Keywords

laser scanning confocalresolutionquality assuranceobjective lenspoint spread function (PSF)spectral imagingspectral unmixingspectral accuracy
Type
Techniques and Instrumentation Development
Information
Microscopy and Microanalysis , Volume 19 , Issue 6 , December 2013 , pp. 1653 - 1668
Copyright
Copyright © Microscopy Society of America 2013 

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