Proliferation assay for fixed solid tumors
AssigneesVentana Medical Systems, Inc., The Francis Crick Institute Limited, The Royal Marsden NHS Foundation Trust
InventorsNelson ALEXANDER, Lisa GALLEGOS, Brian Hanley, Samra Turajlic
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Abstract
Disclosed herein is a method of analyzing flow cytometry data for cells derived from homogenized whole tumor samples. In some embodiments, the present disclosure is directed to a cytometric assay for distinguishing between tumor cells expressing a cell proliferation marker and normal cells expressing the cell proliferation marker. In some embodiments, the present disclosure is also directed to quantifying a percentage of normal cells expressing a cell proliferation marker and a percentage of tumor cells expressing the cell proliferation marker.
Claims20
A method of quantifying a percentage of normal cells and a percentage of tumor cells expressing a cell proliferation marker comprising:
- obtaining a sample;
- staining cells within a first aliquot derived from the sample for the presence of at least one cell proliferation marker;
- staining cells within a second aliquot derived from the sample for the presence of at least one tumor marker;
- optionally counterstaining the cells within the first and second aliquots for the presence of DNA;
- generating scatter plots of fluorescence versus side scatter for the stained cells within each of the first and second aliquots;
- based on the obtained scatter plots, gating the stained cells into at least one of a cell proliferation marker positive tumor cell population and a cell proliferation marker positive normal cell population;
- and
- quantifying the percentage of the normal cells and the percentage of the tumor cells in each of the cell proliferation marker positive tumor cell and cell proliferation marker positive normal cell populations.
A method of assessing a percentage of cell proliferation marker positive normal cells and a percentage of cell proliferation marker positive tumor cells comprising:
- (i) obtaining at least two aliquots of a sample, wherein cells within a first aliquot of the at least two aliquots of the sample are fluorescently stained for the presence of a cell proliferation marker, and wherein cells within a second aliquot of the at least two aliquots of the sample are fluorescently stained for the presence of a tumor marker;
- (ii) generating a first scatter plot of fluorescence versus side scatter for the fluorescently stained cells within the first aliquot of the sample;
- (iii) generating a second scatter plot of fluorescence versus side scatter for the fluorescently stained cells within the second aliquot of the sample;
- and
- (iv) performing at least two gating operations using at least the first and second generated scatter plots to assess the percentage of cell proliferation positive normal cells and the percentage of cell proliferation positive tumor cells.
Dependencies are inferred from the claim text.