The pixel values in the channels are R i and G i, respectively with i the pixel index. In the definitions of the coefficients below we follow the naming convention for the two compared channels: R for the first channel, G for the second channel. See Colocalization Basics for illustrations of the experimental difficulties that affect colocalization. If the image is affected by chromatic aberration, it is advised to correct the image with Huygens Chromatic Aberration Corrector. In case the coefficients need to be computed from raw data it is possible to have this function compute the image background on a frame by frame basis. Deconvolution has proven to sensibly enhance colocalization analysis ( 1, 2 ), see Blur And Noise Affect Colocalization. For these reasons we strongly recommend to compute colocalization coefficients only on deconvolved images. Generally the colocalization coefficients depend much on correct estimation of the image background and resolution. Still, it should be remarked at this point that the specific properties of the coefficients, especially properties related to the image background, make cross-study comparison problematic. Therefore SVI's Colocalization Analyzer focusses on these established coefficients. Several of these coefficients are widely used in literature and lend themselves in principle for comparison of results obtained in different studies. The purpose of a Colocalization coefficient is to characterize the degree of overlap between two channels in a microscopy image. Microscopy colocalization theoretical background
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