Quality assessment of confocal microscopy slide based systems: Performance

Authors

  • Robert M. Zucker

    Corresponding author
    1. Reproductive Toxicology Division, National Health and Environmental Effects Research Laboratory, Office of Researchand Development U.S. Environmental Protection Agency, Research Triangle Park, North Carolina 27711
    • Reproductive Toxicology Division, National Health and Environmental Effects Research Laboratory, Office of Researchand Development U.S. Environmental Protection Agency, Research Triangle Park, North Carolina 27711
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  • The research described in this article has been reviewed and approved for publication as an EPA document. Approval does not necessarily signify that the contents reflects the views and policies of the Agency, or does mention of trade names or commercial products constitute endorsement or recommendation for use.

Abstract

Background:

All fluorescence slide-based cytometry detections systems basically include the following components: (1) an excitation light source, (2) intermediate optics, and (3) a detection device consisting of a CCD camera or a PMT. The optical principles employed is slide-based systems are similar to those of confocal microscopes (CLSM).

Methods:

The following tests evaluated confocal equipment performance: dichroic reflectivity, field illumination, lens performance, laser power output, spectral registration, axial resolution, PMT reliability, and system noise.

Results:

Quality assurance tests provide a basis to determine if the equipment is operating correctly. Laser power, PMTs function, dichroic reflection, spectral registration, axial registration, system noise and sensitivity, lens performance and laser stability were tested colocalization of UV and visible peaks of a bead should be less than 210 nm. Interference contrast optics decrease fluorescence resolution.

Conclusions:

QA tests that assess CLSM system performance are also applicable to other slide-based systems. By utilization this type of testing approach, the subjective nature of assessing the CLSM may be eliminated. These tests serve as guidelines for other investigators to ensure that their machines are providing data that is accurate with the necessary resolution, sensitivity and precision. © 2006 International Society for Analytical Cytology

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