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Clinical and Diagnostic Laboratory Immunology, 09 1995, 598-603, Vol 2, No. 5
Copyright © 1995 by the American Society for Microbiology. All rights reserved.

In vitro detection of apoptotic stimuli by use of the HL-60 myeloid leukemic cell line

JL Shellhaas and SH Zuckerman
Division of Cardiovascular Research, Lilly Research Labs, Indianapolis, Indiana 46285, USA.

The human histiocytic lymphoma line HL-60 has served as a model of myeloid cell differentiation and can be induced to differentiate along the neutrophil or monocytic lineage, depending on the external stimulus. The nondifferentiated cell line retains a premyeloid leukemic phenotype and is capable of anchorage-independent growth and proliferation. The role of apoptosis in the regulation of immunologic and inflammatory events associated with homeostasis and disease has been most intensively studied in lymphocytes. In the present study, nondifferentiated HL-60 has served as a model for studying myeloid cell apoptosis by investigating apoptotic changes induced by camptothecin, a DNA topoisomerase inhibitor, as well as physiologic stimuli, including ceramide analogs and a monoclonal antibody against the Fas antigen. Multiparameter flow cytometry was used to evaluate apoptosis by measuring changes in both side scatter and propidium iodide staining. The appearance of apoptotic cells was confirmed biochemically by measuring DNA endonuclease activity by both enzyme-linked immunosorbent assay quantitation and DNA ladder formation on agarose gels and morphologically with the detection of micronuclei by confocal laser microscopy. These studies demonstrate that HL-60 can serve as an in vitro model for the detection of physiologic and pharmacologic apoptotic stimuli and for understanding the early and late cellular changes associated with induction of the apoptotic program.

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