Analysis and isolation of adipocytes by flow cytometry.

Majka SM, Miller HL, Helm KM, Acosta AS, Childs CR, Kong R, Klemm DJ
Methods Enzymol. 2014 537: 281-96

PMID: 24480352 · PMCID: PMC4143162 · DOI:10.1016/B978-0-12-411619-1.00015-X

Analysis and isolation of adipocytes via flow cytometry is particularly useful to study their biology. However, the adoption of this technology has often been hampered by the presence of stromal/vascular cells in adipocyte fractions prepared from collagenase-digested adipose tissue. Here, we describe a multistep staining method and gating strategy that effectively excludes stromal contaminants. Initially, we set a gate optimized to the size and internal complexity of adipocytes. Exclusion of cell aggregates is then performed based on fluorescence of a nuclear stain followed by positive selection to collect only those cell events containing lipid droplets. Lastly, negative selection of cells expressing stromal or vascular lineage markers removes any remaining stromal contaminants. These procedures are applicable to simple analysis of adipocytes and their subcellular constituents by flow cytometry as well as isolation of adipocytes by flow sorting.

© 2014 Elsevier Inc. All rights reserved.

MeSH Terms (8)

Adipocytes Adipose Tissue Biomarkers Cell Differentiation Cell Lineage Cell Separation Flow Cytometry Humans

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