Independent fluorescent labeling of two different components is a commonly used method to determine their co-localization. This website contains an ImageJ plugin for automatic quantification and visualization of co-localized fluorescent signals. The plugin is based on the method originally described by Jaskolski et. al. (2005). This algorithm computes correlation of intensities between pairs of individual pixels in two different channels. Results are presented as a co-localization color map where hot colors represent positive correlation (co-localization), whereas colors that are cold represent negative correlation (exclusion).
Spatial representation of colocalization
Correlation between pair of pixels (described by x,y coordinates in the image) is calculated according to the formula below:
Distribution of nMDP (normalized mean deviation product) values (ranging from -1 to 1) is visualized with a color scale. Negative indexes are represented by cold colors (exclusion). Indexes above 0 are represented by hot colors (co-localization).
Automatic co-localization calculation using Colocalization Colormap plugin.
(A) Neuronal body(asterisks) and dendritic processes (arrows)
stained with antibody against MAP2 protein.
(B) Immunoreactivity of CD44 adhesion molecule in the brain.
(C-D) Color map presenting co-localization between adhesion molecule and MAP2 neuronal marker.