Patent ID: 8116551

Claim:
A method for the analysis of an image of a biological sample comprising one or more candidate objects of interest comprising the steps of: (a) obtaining a digitised image of the biological sample; (b) separating the image of the biological sample into a First Image and a Second Image, the separating step comprising; (i) defining a first threshold for hue saturation intensity and three colour stains (RGB colour space) properties of a first optical enhancer and using the first threshold properties in the digitised image to produce the First Image comprising the background and the three colour stains (RGB colour space) information of the first optical enhancer and (ii) defining a second threshold for hue saturation intensity and three colour stains (RGB colour space) properties of a second optical enhancer and using the second threshold properties in the digitised image to produce the Second Image comprising the background and the three colour stains (RGB colour space) information of the second optical enhancer; (c) assimilating the morphological features or morphological pattern of the one or more candidate objects of interest in the First Image, the assimilating step comprising processing the First Image by conducing colour space transformation to convert the three colour stains (RGB colour space) information to a perceptually uniform colour space followed by a segmentation step comprising firstly conducting colour demarcation and secondly conducting watershed segmentation to create a 3D view of the candidate objects of interest from which the morphological features or morphological pattern are measured; (d) applying to the Second Image the morphological features or morphological pattern assimilated from the First Image by matching the morphological features or morphological pattern assimilated from the First Image with morphological features or morphological pattern in the Second Image; (e) eliminating from the Second Image objects with non-matching morphological features or non-matching morphological pattern with the one or more candidate objects of interest in the First Image; and (f) quantitating the level of expression of the one or more candidate objects of interest in the First Image using the matching morphological features or morphological pattern of the one or more candidate objects in the First Image and the Second Image.