Immuno-phenotype of cells in apoptosis and/or necrosis.

What is it about?

To date, the methods used to detect apoptosis and/or necrosis of whole cells were incompatible with the procedures required to detect intracytoplasmic antigens or to inactivate intracellular or extracellular pathogens. Thus, it was not possible to measure by flow cytometry intracellular antigen expression, such as cytokines, in apoptotic or necrotic cells. Moreover, only laboratories equipped with a flow cytometer in BSL3 had the chance to measure the phenotypic characteristics of apoptotic/necrotic cells following infection with certain pathogens.

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Why is it important?

There are several causes inducing cell apoptosis and/or necrosis, including infections with certain pathogens. Flow cytometry is a useful tool to detect apoptotic and necrotic cells in association with surface exposed antigens. However, the surface expression of molecules in early apoptotic cells only could be considered reliable, since the increased permeability of late apoptotic and necrotic cell membrane may cause the passive acquisition of the fluorochrome-labelled probes used to detect membrane phenotype, causing false positivity. The proposed method circumvents this limitation permitting to detect both surface exposed and intracellular markers of cells in apoptosis. Moreover, the possibility to fix stained cells, that is permitted by the proposed method, gives the opportunity to inactivate pathogens and to study the phenotype of cells induced to programmed cell death or necrosis by infectious agents.

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