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INTRODUCTION: Flow cytometry has become an important technique for rapid monitoring of cellular receptor expression and proliferation in hematological tumors. In contrast flow cytometry has been of limited use for the analysis of human solid tumors. Except for determination of DNA content and proliferation, monitoring of other cellular parameters by flow cytometry in breast tumors has been difficult. Most of these difficulties are posed by architectural characteristics of breast tumors where the tumor cells are embedded in stromal components containing infiltrating lymphoid cells and other non-cellular elements such as collagen. As it is comparatively easy to isolate nuclei from human breast tumors, we have worked on the hypothesis that one can use flow cytometry for monitoring the expression of nuclear markers of clinical significance. This could include nuclear hormone receptors (e.g., estrogen, progesterone, vitamin D) in combination with measurement of nuclear volume and DNA content. As the house keeping oncogenes, p53 and the proliferation marker, Ki-67 have important prognostic significance and are also expressed in nuclei, our objective was to develop multiparametric methods for rapid analysis of nuclear markers by flow cytometry.