Immunofluorescence-based Determination of Centrosome Number in Tissue Samples.

Centrosome numerical abnormalities have been reported in a variety of tumors. Centrosome numbers in cancer cells display both inter-tumor and intra-tumor heterogeneity. The over production of centrosomes (centrosome amplification) is unique in cancer cells and is a promising target for therapy. Thus, a method to quantify centrosome numbers on a single cell level is needed. Here, we describe a protocol to quantify centrosome numbers in formalin fixed paraffin embedded (FFPE) tissue samples by multiplexing antibodies to define bona fide centrosomes and cell borders. Centrosomes in single cells are identified using high resolution immunofluorescent microscopy with Z-sectioning. This protocol is easy to perform and has been used to quantify centrosome numbers on a single cell level in a variety of human tissue samples.