Universität zu Köln

Gutierrez, Saray, Wolke, Martina, Plum, Georg and Robinson, Nirmal ORCID: 0000-0002-7361-9491 (2017). Isolation of Salmonella typhimurium-containing Phagosomes from Macrophages. J. Vis. Exp. (128). CAMBRIDGE: JOURNAL OF VISUALIZED EXPERIMENTS. ISSN 1940-087X

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Abstract

Salmonella typhimurium is a facultative intracellular bacterium that causes gastroenteritis in humans. After invasion of the lamina propria, S. typhimurium bacteria are quickly detected and phagocytized by macrophages, and contained in vesicles known as phagosomes in order to be degraded. Isolation of S. typhimurium-containing phagosomes have been widely used to study how S. typhimurium infection alters the process of phagosome maturation to prevent bacterial degradation. Classically, the isolation of bacteria-containing phagosomes has been performed by sucrose gradient centrifugation. However, this process is time-consuming, and requires specialized equipment and a certain degree of dexterity. Described here is a simple and quick method for the isolation of S. typhimurium-containing phagosomes from macrophages by coating the bacteria with biotin-streptavidin-conjugated magnetic beads. Phagosomes obtained by this method can be suspended in any buffer of choice, allowing the utilization of isolated phagosomes for a broad range of assays, such as protein, metabolite, and lipid analysis. In summary, this method for the isolation of S. typhimurium-containing phagosomes is specific, efficient, rapid, requires minimum equipment, and is more versatile than the classical method of isolation by sucrose gradient-ultracentrifugation.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Gutierrez, Saray UNSPECIFIED UNSPECIFIED UNSPECIFIED Wolke, Martina UNSPECIFIED UNSPECIFIED UNSPECIFIED Plum, Georg UNSPECIFIED UNSPECIFIED UNSPECIFIED Robinson, Nirmal UNSPECIFIED orcid.org/0000-0002-7361-9491 UNSPECIFIED
URN:	urn:nbn:de:hbz:38-215012
DOI:	10.3791/56514
Journal or Publication Title:	J. Vis. Exp.
Number:	128
Date:	2017
Publisher:	JOURNAL OF VISUALIZED EXPERIMENTS
Place of Publication:	CAMBRIDGE
ISSN:	1940-087X
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language MATURATION; PATHOPHYSIOLOGY; MYCOBACTERIUM; TRAFFICKING; PHYSIOLOGY; PROTEOME; BRUCELLA; SYSTEM; ROS Multiple languages Multidisciplinary Sciences Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/21501