Comparative image and flow cytometric TUNEL analysis of fine needle samples of breast carcinoma

Abstract

The assessment of apoptosis in solid tumors is of interest because of its biological role in tumor evolution and response to therapy. A commonly used method for apoptosis measurement is the TUNEL 3′ end-labeling technique, which has shown wide variations in results when applied to solid tumors. Thirty-one tine needle breast carcinoma samples were analyzed by fluorescent TUNEL assay and DNA content using image analysis and flow cytometry. TUNEL positivity, seen both in cells with apoptotic morphology and in a subset of morphologically normal cells, was categorized into five staining patterns and quantitated. Values for patterns of TUNEL-positive cells were compared with TUNEL positivity measured by flow cytometry. Flow cytometric quantitation showed a mean of 24.3% positive cells, which correlated (P > 0.02) with total positive cells (all patterns) measured by image (22.4%). Image analysis quantitation of morphologically apoptotic cells (4.2%) did not correlate with flow cytometric TUNEL positivity and the majority of TUNEL-stained cells were morphologically normal (17%). Image analysis allows discrimination of TUNEL-positive morphologically apoptotic and nonapoptotic cells, which are included in the total number of TUNEL-positive events measured by flow cytometry. © 2001 Wiley-Liss, Inc.