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2014 ; 1124
(ä): 251-68
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Immunofluorescence and confocal microscopy of neutrophils
#MMPMID24504957
Allen LA
Methods Mol Biol
2014[]; 1124
(ä): 251-68
PMID24504957
show ga
Rapid recruitment of neutrophils to sites of infection and their ability to
phagocytose and kill microbes is an important aspect of the innate immune
response. Challenges associated with imaging of these cells include their short
lifespan and small size and the fact that unstimulated cells are nonadherent. In
addition, although cytoplasmic granules are plentiful, the abundance of many
other organelles is diminished. Here we reprise methods for analysis of resting
and activated cells using immunofluorescence and confocal microscopy, including
kinetic analysis of phagosome maturation and degranulation, and detection of
intraphagosomal superoxide accumulation. We describe approaches for rapid cell
fixation and permeabilization that maximize antigen detection and discuss other
variables that also affect data interpretation and image quality (such as cell
spreading, degranulation, and phagocytosis). Finally, we show that these methods
are also applicable to studies of neutrophil interactions with the extracellular
matrix.