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. 1993 Mar;169(3):375-382.
doi: 10.1111/j.1365-2818.1993.tb03313.x.

Measurement of co-localization of objects in dual-colour confocal images

E M M Manders  1 , F J Verbeek  2   3 , J A Aten  1
Affiliations

Measurement of co-localization of objects in dual-colour confocal images

E M M Manders et al. J Microsc. 1993 Mar.

Abstract

A method to measure the degree of co-localization of objects in confocal dual-colour images has been developed. This image analysis produced two coefficients that represent the fraction of co-localizing objects in each component of a dual-channel image. The generation of test objects with a Gaussian intensity distribution, at well-defined positions in both components of dual-channel images, allowed an accurate investigation of the reliability of the procedure. To do that, the co-localization coefficients were determined before degrading the image with background, cross-talk and Poisson noise. These synthesized sources of image deterioration represent sources of deterioration that must be dealt with in practical confocal imaging, namely dark current, non-specific binding and cross-reactivity of fluorescent probes, optical cross-talk and photon noise. The degraded images were restored by filtering and cross-talk correction. The co-localization coefficients of the restored images were not significantly different from those of the original undegraded images. Finally, we tested the procedure on images of real biological specimens. The results of these tests correspond with data found in the literature. We conclude that the co-localization coefficients can provide relevant quantitative information about the positional relation between biological objects or processes.

Keywords: Co-localization; confocal microscopy; correlation; double labelling; image analysis; image reconstruction.

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References

    1. Akner, G., Mossberg, K., Wilkström, A., Sundqvist, K. & Gustafsson, J. (1991) Evidence for colocalization of glucocorticoid receptor with cytoplasmic microtubiles in human gingival fibroblasts, using two different monoclonal anti-gr antibodies, confocal microscopy and image analysis. J. Steroid Biochem. Molec. Biol. 39, 419-432.
    1. Akner, G., Sundqvist, K., Denis, M., Wilkström, A. & Gustafsson, J. (1990) Immunocytochemical localization of glucocorticoid receptor in human gingival fibroblasts and evidence for colocalization of glucocorticoid receptor with cytoplasmic microtubiles. Eur. J. Cell Biol. 53, 390-401.
    1. Arndt-Jovin, D.J., Robert-Nicoud, M. & Jovin, T.M. (1990) Probing DNA structures and function with multi-wavelength fluorescence confocal laser microscope. J. Microsc. 157, 61-72.
    1. Aten, J.A., Bakker, P.J.M., Stap, J., Boschman, G.A. & Veenhof, C.H.N. (1992) DNA double labeling with IdUrd and CldUrd for spatial and temporal analysis of cell proliferation and DNA replication. Histochem. J. 24, 251-259.
    1. Carlsson, K. (1990) Scanning and detection techniques used in a confocal scanning laser microscope. J. Microsc. 157, 21-27.

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