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Research Article
Quantization of widefield fluorescence images using structured illumination and image analysis software
Article first published online: 27 NOV 2006
DOI: 10.1002/jemt.20389
Copyright © 2006 Wiley-Liss, Inc.
Additional Information
How to Cite
Barlow, A. L. and Guerin, C. J. (2007), Quantization of widefield fluorescence images using structured illumination and image analysis software. Microscopy Research and Technique, 70: 76–84. doi: 10.1002/jemt.20389
Publication History
- Issue published online: 14 DEC 2006
- Article first published online: 27 NOV 2006
- Manuscript Accepted: 16 SEP 2006
- Manuscript Received: 7 APR 2006
- Abstract
- References
- Cited By
Keywords:
- quantitative studies;
- measurements;
- transferrin;
- internalization;
- intensity
Abstract
It is difficult to obtain precise quantitative measurements from fluorescent images captured from widefield microscopes. We wished to ascertain if reliable quantitative measurements of both biological and nonbiological specimens were possible using a widefield microscope equipped with a structured illumination system and image analysis software. In a nonbiological specimen, images were obtained from fluorescent beads of known intensity. For a biologically relevant model we used the uptake of fluorescently labeled transferrin by HeLa cells in culture. In the bead sample, the mean intensity of beads acquired with the structured illumination system showed relative intensities near to the predicted values without any further manipulation of data. In the transferrin uptake experiments, uptake and subsequent recycling of the labeled transferrin could be accurately and reproducibly measured. We conclude that using a combination of structured illumination and image processing algorithms accurate quantization of fluorescent images can be achieved in widefield microscopyMicrosc. Res. Tech., 2007. © 2006 Wiley-Liss, Inc.