Abstract
This chapter describes three methods for measuring the bactericidal activity of neutrophils. All utilize colony counting techniques to quantify viable bacteria. A simple “one-step” protocol provides a composite measure of phagocytosis and killing, while a “two-step” protocol that separates extracellular and intracellular bacteria allows calculation of rate constants for both of these processes. We also present a method for selectively monitoring the long-term survival of bacteria within the phagosome. This may have application in identifying resistant strains and searching for compounds that sensitize pathogens to destruction.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Winterbourn CC, Kettle AJ, Hampton MB (2016) Reactive oxygen species and neutrophil function. Annu Rev Biochem 85:765–792
Gresham HD, Lowrance JH, Caver TE et al (2000) Survival of Staphylococcus aureus inside neutrophils contributes to infection. J Immunol 164:3713–3722
Shapiro HM (2008) Flow cytometry of bacterial membrane potential and permeability. Methods Mol Med 142:175–186
Parker HA, Magon NJ, Green JN et al (2014) Analysis of neutrophil bactericidal activity. Methods Mol Biol 1124:291–306
Nauseef WM (2014) Isolation of human neutrophils from venous blood. Methods Mol Biol 1124:13–18
Tiden AK, Sjogren T, Svensson M et al (2011) 2-thioxanthines are mechanism-based inactivators of myeloperoxidase that block oxidative stress during inflammation. J Biol Chem 286:37578–37589
Hampton MB, Vissers MC, Winterbourn CC (1994) A single assay for measuring the rates of phagocytosis and bacterial killing by neutrophils. J Leukoc Biol 55:147–152
Decleva E, Menegazzi R, Busetto S et al (2006) Common methodology is inadequate for studies on the microbicidal activity of neutrophils. J Leukoc Biol 79:87–94
Gargan RA, Brumfitt W, Hamilton-Miller JM (1989) Failure of water to lyse polymorphonuclear neutrophils completely. Role of pH and implications for assessment of bacterial killing. J Immunol Methods 124:289–291
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
1 Electronic Supplementary Material
Data 1
: (XLSX 88 kb)
Rights and permissions
Copyright information
© 2020 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Magon, N.J., Parker, H.A., Ashby, L.V., Springer, R.J., Hampton, M.B. (2020). Analysis of Neutrophil Bactericidal Activity. In: Quinn, M., DeLeo, F. (eds) Neutrophil. Methods in Molecular Biology, vol 2087. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0154-9_12
Download citation
DOI: https://doi.org/10.1007/978-1-0716-0154-9_12
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-0153-2
Online ISBN: 978-1-0716-0154-9
eBook Packages: Springer Protocols