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Quantization of widefield fluorescence images using structured illumination and image analysis software
Author(s) -
Barlow Andrew L.,
Guerin Christopher J.
Publication year - 2007
Publication title -
microscopy research and technique
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 118
eISSN - 1097-0029
pISSN - 1059-910X
DOI - 10.1002/jemt.20389
Subject(s) - fluorescence , microscope , fluorescence microscope , microscopy , software , optics , image processing , quantization (signal processing) , computer science , artificial intelligence , computer vision , biological system , biomedical engineering , materials science , physics , image (mathematics) , biology , medicine , programming language
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.