Intensity calibration and shading correction for fluorescence microscopes.
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 dye solution 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 that make it inexpensive, reproducible, and straightforward. Descriptions are given for both wide-field and confocal scanning fluorescence microscopy.
Model
Michael A
MA
Kent State University, Kent, Ohio, USA.
D002138:Calibration; D019793:Fluorescein; D007089:Image Enhancement / Q000379:methods; D007090:Image Interpretation, Computer-Assisted / Q000379:methods / Q000592:standards; D007091:Image Processing, Computer-Assisted / Q000379:methods; D018613:Microscopy, Confocal / Q000379:methods* / Q000592:standards; D008856:Microscopy, Fluorescence / Q000379:methods* / Q000592:standards; D012680:Sensitivity and Specificity
D019793:Fluorescein