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Lectin‐based three‐color flow cytometric approach for studying cell surface glycosylation changes that occur during apoptosis
Author(s) -
Batisse Chantal,
Marquet Jeanine,
Greffard Anne,
FleuryFeith Jocelyne,
Jaurand MarieClaude,
Pilatte Yannick
Publication year - 2004
Publication title -
cytometry part a
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.316
H-Index - 90
eISSN - 1552-4930
pISSN - 1552-4922
DOI - 10.1002/cyto.a.20094
Subject(s) - glycosylation , propidium iodide , agglutinin , apoptosis , lectin , flow cytometry , sialic acid , cell , annexin , biology , microbiology and biotechnology , biochemistry , chemistry , programmed cell death
Background Changes in cell surface glycosylation that accompany apoptosis are thought to be involved in the recognition and removal of apoptotic cells by phagocytes, but in most instances these changes are ill defined. To improve our understanding of this phenomenon, we designed a trivariate flow cytometry procedure that allows direct comparison of cell surface glycosylation in apoptotic and viable cells Methods The annexin V/propidium iodide assay has been adapted for cell surface glycosylation analysis by combining the use of these two reagents with biotinylated lectins, and this has been used to investigate camptothecin‐induced apoptosis in U‐937 cells. Results Although numerous lectins are potent inducers of apoptosis, we found that it is possible to determine lectin concentrations that produce interpretable data without inducing significant cytotoxicity even when using apoptogenic lectins. That apoptosis is associated with a marked decrease in cell surface sialylation was confirmed by using the sialic acid–specific lectins Maackia amurensis agglutinin and Sambucus nigra agglutinin. These observations were corroborated by lectin blotting analysis with the same lectins. Conclusions Species‐ and cell‐dependent altered glycosylation patterns are likely to be associated with different modes of apoptosis. The easy and versatile method described in this report should be useful for exploring this field. © 2004 Wiley‐Liss, Inc.