Macropinosome quantitation assay

Wang, Jack T. H., Teasdale, Rohan D. and Liebl, David (2014) Macropinosome quantitation assay. MethodsX, 1 1: 36-41. doi:10.1016/j.mex.2014.05.002


Author Wang, Jack T. H.
Teasdale, Rohan D.
Liebl, David
Title Macropinosome quantitation assay
Journal name MethodsX   Check publisher's open access policy
ISSN 2215-0161
Publication date 2014
Year available 2014
Sub-type Article (original research)
DOI 10.1016/j.mex.2014.05.002
Open Access Status DOI
Volume 1
Issue 1
Start page 36
End page 41
Total pages 6
Place of publication Amsterdam, Netherlands
Publisher Elsevier
Collection year 2015
Language eng
Formatted abstract
In contrast to phagocytosis, macropinocytosis is not directly initiated by interactions between cell surface receptors and cargo ligands, but is a result of constitutive membrane ruffling driven by dynamic remodelling of cortical actin cytoskeleton in response to stimulation of growth factor receptors. Wang et al. (2010) [13] developed a reliable assay that allows quantitative assessment of the efficiency and kinetics of macropinosome biogenesis and/or maturation in cells where the function of a targeted protein has been perturbed by pharmacological inhibitors or by knock-down or knock-out approaches. In this manuscript we describe a modified quantitative protocol to measure the rate and volume of fluid phase uptake in adherent cells. This assay:

• uses fluorescent dextran, microscopy and semi-automated image analysis;

• allows quantitation of macropinosomes within large numbers of individual cells;

• can be applied also to non-homogenous cell populations including transiently transfected cell monolayers. We present the background necessary to consider when customising this protocol for application to new cell types or experimental variations.
Keyword Macropinocytosis
Macropinosome
Fluid-phase endocytosis
Dextran
Fluorescence microscopy
Quantitation of macropinocytosis
Amiloride
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Official 2015 Collection
School of Chemistry and Molecular Biosciences
Institute for Molecular Bioscience - Publications
 
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