Protocol for measuring cellular energetics through noncanonical amino acid tagging in human peripheral blood and murine tissue immune cells
Cellular metabolism dictates immune cell function, yet we lack tools to functionally profile immunometabolism in low-yield, complex samples. We present a flow cytometry-based protocol for measuring cellular energetics through noncanonical amino acid tagging (CENCAT) in human peripheral blood and murine tissue immune cells. We describe steps for sample preparation, metabolic inhibition, protein synthesis analysis using click chemistry, immunophenotyping, and calculation of metabolic dependencies. For complete details on the use and execution of this protocol, please refer to Vrieling et al.<sup>1</sup>.
Authors
- Rinke Stienstra (ORCID: https://orcid.org/0000-0001-6729-7193)
- Hendrik J.P. van der Zande (ORCID: https://orcid.org/0000-0003-1479-1934)
- Frank Vrieling (ORCID: https://orcid.org/0000-0002-8064-672X)
- Marline Dumont
Institutions
- Radboud University Nijmegen (NL)
- University Medical Center (US)
- Radboud University Medical Center (NL)
- Wageningen University & Research (NL)
Publication Details
- Journal
- STAR Protocols
- Published
- 2025-10-09
- DOI
- https://doi.org/10.1016/j.xpro.2025.104134
- Primary Topic
- RNA and protein synthesis mechanisms
- Type
- article
- Field-Weighted Citation Impact
- 0.00
Funders
- Health~Holland
- Universiteit Leiden
- ZonMw
- Diabetes Fonds
- European Society for Clinical Nutrition and Metabolism
- Canadian Institutes of Health Research