Patent ID: 8036833

Claim:
A computer implemented method for localizing and quantitating a particular biomarker within a first marker defined subcellular compartment relative to a second marker defined subcellular compartment present in individual cells of interest contained in a tissue sample comprising: a) incubating the tissue sample with a first stain that specifically labels the first marker defined subcellular compartment, second stain that specifically labels the second marker defined subcellular compartment, and a third stain that specifically labels the biomarker; b) obtaining a high resolution image of each of the first, the second, and the third stain in the tissue sample using an upright or inverted optical microscope so as to obtain: i) a first image of the first marker defined subcellular compartment; ii) a second image of the second marker defined subcellular compartment; and iii) a third image of the biomarker; wherein each image comprises 1024×1024pixel locations; c) reiteratively analyzing each pixel location in the first and the second image so as to assign each such pixel location to the first, the second or neither subcellular compartment based upon an intensity value of the first stain relative to the second stain at that pixel location; d) analyzing in the third image the pixel locations assigned to the first or the second subcellular compartment in step (c) so as to identify those pixel locations having an intensity value indicative of the third stain, and determining the total intensity value of the third stain at the pixel locations in each of the first and the second subcellular compartment; so as to thereby localize and quantitate the biomarker in the first subcellular compartment relative to the second subcellular compartment.