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Phenotypic and functional consequences of different isolation protocols on skin mononuclear phagocytes

Lookup NU author(s): James Fletcher, Professor Muzlifah Haniffa

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This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).


Abstract

© The Author(s). Mononuclear phagocytes are present in skin and mucosa and represent one of the first lines of defense against invading pathogens, which they detect via an array of pathogen-binding receptors expressed on their surface. However, their extraction from tissue is difficult, and the isolation technique used has functional consequences on the cells obtained. Here, we compare mononuclear phagocytes isolated from human skin using either enzymatic digestion or spontaneous migration. Cells isolated via enzymatic digestion are in an immature state, and all subsets are easily defined. However, cells isolated by spontaneous migration are in a mature state, and CD141 cross-presenting DCs (cDC1) are more difficult to define. Different pathogen-binding receptors are susceptible to cleavage by blends of collagenase, demonstrating that great care must be taken in choosing the correct enzyme blend to digest tissue if carrying out pathogen-interaction assays. Finally, we have optimized mononuclear phagocyte culture conditions to enhance their survival after liberation from the tissue.


Publication metadata

Author(s): Botting RA, Bertram KM, Baharlou H, Sandgren KJ, Fletcher J, Rhodes JW, Rana H, Plasto TM, Wang XM, Lim JJK, Barnouti L, Kohout MP, Papadopoulos T, Merten S, Olbourne N, Cunningham AL, Haniffa M, Harman AN

Publication type: Article

Publication status: Published

Journal: Journal of Leukocyte Biology

Year: 2017

Volume: 101

Issue: 6

Pages: 1393-1403

Print publication date: 01/06/2017

Online publication date: 07/03/2017

Acceptance date: 14/02/2017

Date deposited: 21/06/2017

ISSN (print): 0741-5400

ISSN (electronic): 1938-3673

Publisher: Federation of American Societies for Experimental Biology

URL: https://doi.org/10.1189/jlb.4A1116-496R

DOI: 10.1189/jlb.4A1116-496R


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