Abstract: The invention relates generally to a process of analyzing and visualizing the expression of biomarkers in an individual cell wherein the cell is examined to develop patterns of expression by using a grouping algorithm, and a system to perform and display the analysis.

Abstract: Novel methods of probing multiple targets in a biological sample are provide whereby the targets are DNA, RNA and protein. The method comprises subjecting the sample to an in situ hybridization reaction using a labeled nucleic acid probe that binds an RNA target, observing a signal, and optionally removing the signal. The method further comprises an antigen retrieval protocol, observing a signal, removing the signal, and optionally applying a protease treatment to access the sample's DNA targets by subjecting the sample to an in situ hybridization reaction using a labeled nucleic acid probe, observing a signal from the labeled DNA targets, and optionally removing the signal.

Abstract: Automated methods and devices that facilitate iterative staining of biological samples from imaging applications are provided. The methods include the steps of providing a small volume flow cell containing a biological sample, applying a stain to the biological sample, combining at least two precursor reagents to form an activated destaining agent and wherein the activated destaining agent decomposition rate is greater than or similar to the destaining reaction rate, flowing the destaining agent over the biological sample at a flow rate that is greater than the decomposition rate of the activated destaining agent, and releasing the sample from the flow cell wherein the integrity of the sample is intact. The process of staining, combining and flowing may be iteratively repeated.

Abstract: The present application discloses a technique for obtaining and storing data on expression of multiple biomarkers in individual cells or the compartments of individual cells in a tissue specimen and methods of utilizing that data to create groups, the members of which share some degree of similarity greater than the general population from which the data is drawn, by an analysis of digital images of a portion of the tissue specimen which has been iteratively stained to generate optical signals, typically fluorescent, reflective of the amount of each of the biomarkers examined.