1. USER GUIDE FOR COLOCALIZATION
The concept of colocalization refers to the presence of different molecules at the same physical location within a given resolution. Among a variety of different techniques that have been developed to detect colocalization, fluorescence related techniques are increasingly attractive since they permit to localize cellular structures with a high degree of specificity and sensitivity.
Within the context of multi-channel (confocal) fluorescence microscopy and digital image processing, two fluorescent molecules colocalize when the emitted photons are detected inside the same pixel (2 dimensions) or voxel (3 dimensions). So far, no standard method has been established in science which would characterize colocalization unequivocally for the huge diversity of existing biological structures. For this reason, we implemented a series of graphic-statistical and mathematical tools into SCIAN which facilitate the application of the most frequently used qualitative and quantitative characterizations methods. The degree of colocalization can be determined by:
In contrast to all preceding methods which calculate colocalization coefficients for an image set with rather simple, global mathematical coefficients (based on the theories of Manders or Pearson), our approach presents a consequent
bottom-up algorithm based on the calculation of
local scaling functions which we term Colocalization Scaling Index Methods (C-SIM). These local C-SIM are defined within a given radius r for every position in the image matrices.
Finally, SCIAN contains a number of ‘randomization’ procedures which simulate case specific random situations on the basis of the explicit experimental data. The following pages summarize and outline the most important features for colocalization in SCIAN. You can download our User Guide for Colocalization here:
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