Advanced

Quantitative assessment of neutrophil phagocytosis using flow cytometry

Nordenfelt, Pontus LU (2014) In Neutrophil Methods and Protocols 1124. p.89-279
Abstract

Neutrophils have an incredible ability to find and eradicate intruders such as bacteria and fungi. They do this largely through the process of phagocytosis, where the target is internalized into a phagosome, and eventually destroyed by the hostile phagosomal environment. It is important to study phagocytosis in order to understand how neutrophils interact with various pathogens and how they respond to different stimuli. Here, I describe a method to study neutrophil phagocytosis of bacteria using flow cytometry. The bacteria are fluorescently labeled before being introduced to neutrophils. After phagocytosis, both any remaining extracellular bacteria and neutrophils are labeled using one-step staining before three-color analysis. To... (More)

Neutrophils have an incredible ability to find and eradicate intruders such as bacteria and fungi. They do this largely through the process of phagocytosis, where the target is internalized into a phagosome, and eventually destroyed by the hostile phagosomal environment. It is important to study phagocytosis in order to understand how neutrophils interact with various pathogens and how they respond to different stimuli. Here, I describe a method to study neutrophil phagocytosis of bacteria using flow cytometry. The bacteria are fluorescently labeled before being introduced to neutrophils. After phagocytosis, both any remaining extracellular bacteria and neutrophils are labeled using one-step staining before three-color analysis. To assess phagocytosis, first the average time it takes for the neutrophils to internalize all bound bacteria is determined. Experiments are then performed using that time point while varying the bacteria-to-neutrophil ratio for full control of the analysis. Due to the ease with which multiple samples can be analyzed, and the quantitative nature of flow cytometry, this approach is both reproducible and sensitive.

(Less)
Please use this url to cite or link to this publication:
author
publishing date
type
Chapter in Book/Report/Conference proceeding
publication status
published
keywords
Flow Cytometry, Humans, Neutrophils, Phagocytosis, Journal Article, Research Support, Non-U.S. Gov't
in
Neutrophil Methods and Protocols
editor
Quinn, Mark T.; DeLeo, Frank R.; and
volume
1124
pages
11 pages
publisher
Humana Press
external identifiers
  • scopus:84899000167
ISSN
1064-3745
ISBN
978-1-62703-844-7
978-1-62703-845-4
DOI
10.1007/978-1-62703-845-4_18
language
English
LU publication?
no
id
5b8fee79-0626-451c-9d42-0273c618b6cc
date added to LUP
2017-02-16 08:03:58
date last changed
2017-02-19 04:45:25
@inbook{5b8fee79-0626-451c-9d42-0273c618b6cc,
  abstract     = {<p>Neutrophils have an incredible ability to find and eradicate intruders such as bacteria and fungi. They do this largely through the process of phagocytosis, where the target is internalized into a phagosome, and eventually destroyed by the hostile phagosomal environment. It is important to study phagocytosis in order to understand how neutrophils interact with various pathogens and how they respond to different stimuli. Here, I describe a method to study neutrophil phagocytosis of bacteria using flow cytometry. The bacteria are fluorescently labeled before being introduced to neutrophils. After phagocytosis, both any remaining extracellular bacteria and neutrophils are labeled using one-step staining before three-color analysis. To assess phagocytosis, first the average time it takes for the neutrophils to internalize all bound bacteria is determined. Experiments are then performed using that time point while varying the bacteria-to-neutrophil ratio for full control of the analysis. Due to the ease with which multiple samples can be analyzed, and the quantitative nature of flow cytometry, this approach is both reproducible and sensitive.</p>},
  author       = {Nordenfelt, Pontus},
  editor       = {Quinn, Mark T. and DeLeo, Frank R.},
  isbn         = {978-1-62703-844-7},
  issn         = {1064-3745},
  keyword      = {Flow Cytometry,Humans,Neutrophils,Phagocytosis,Journal Article,Research Support, Non-U.S. Gov't},
  language     = {eng},
  pages        = {89--279},
  publisher    = {Humana Press},
  series       = {Neutrophil Methods and Protocols},
  title        = {Quantitative assessment of neutrophil phagocytosis using flow cytometry},
  url          = {http://dx.doi.org/10.1007/978-1-62703-845-4_18},
  volume       = {1124},
  year         = {2014},
}