Isolation and Characterization of Adipose Tissue Macrophages
This chapter describes a technique that can be used to isolate adipose tissue macrophages (ATMs) from the visceral white adipose tissue. Nevertheless, this technique can also be used to isolate ATMs from subcutaneous white adipose tissue and brown adipose tissue from mouse, human subcutaneous fat depot, and also from the fat body of the toad Xenopus. We detail the flow-cytometric gating strategy that has been developed to identify ATM population, and we describe the isolation of RNA from this population and its use for gene expression profiling. Finally, we describe in vitro culture of ATMs for downstream applications.
Key wordsMacrophage Adipose tissue Flow cytometry (FACS) Immunity Ribonucleic acid (RNA)
The work in the authors’ laboratory is supported by a German Research Fund (DFG) grant and by the European Foundation for the Study of Diabetes (EFSD). Scopus author identifiers of the authors are 56764443800 (G.A.) and 7801656864 (T.R.).
- 15.Union OJotE Directive 2010/63/EU of the European Parliament and of the Council of 22 September 2010 on the protection of animals used for scientific purposes Text with EEA relevance. 20.10.2010 [cited 2018; 29.04.2018]; Available from http://eur-lex.europa.eu/eli/dir/2010/63/oj
- 16.Scientific T Convert between times gravity (×g) and centrifuge rotor speed (RPM) 2009 [cited 2018; 29.04.2018]; Available from https://tools.thermofisher.com/content/sfs/brochures/TR0040-Centrifuge-speed.pdf