UNIT 10.14 Calibration and Shading Correction for Fluorescence Microscopes

  1. Michael A. Model

Published Online: 1 AUG 2002

DOI: 10.1002/0471142956.cy1014s21

Current Protocols in Cytometry

Current Protocols in Cytometry

How to Cite

Model, M. A. 2002. Calibration and Shading Correction for Fluorescence Microscopes. Current Protocols in Cytometry. 10:10.14.

Author Information

  1. Cleveland Clinic Foundation, Cleveland, Ohio

Publication History

  1. Published Online: 1 AUG 2002
  2. Published Print: JUL 2002

This is not the most recent version of the article. View current version (1 APR 2014)


Standardization in image cytometry involves intensity calibration and shading correction. This unit presents a method using concentrated solutions of fluorophores. A drop of highly concentrated fluorescein or rhodamine placed between a slide and a coverslip produces a spatially uniform fluorescent sample with reproducible quantum yield and resistance to photobleaching. The technique has a number of practical features making it inexpensive, reproducible, and straightforward.