Single-cell mapping of metabolic dependences using ATP as a biomarker

Mapping metabolic dependences and capacities using ATP as a biomarker | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Article Mapping metabolic dependences and capacities using ATP as a biomarker Peter McGuire, Jose Marin, Amanda Fuchs, Tatiana Tarasenko, Emily Warren, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-4836421/v2 This work is licensed under a CC BY 4.0 License Status: Posted Version 2 posted You are reading this latest preprint version Show more versions Abstract Metabolic dependences highlight a cell's reliance on specific pathways to meet its bioenergetic needs, with these pathways being interrogated using chemical inhibitors to assess their significance. While surrogate markers of bioenergetics (e.g., oxygen consumption) have yielded important insights, we asked whether metabolic dependences could be defined using ATP as a biomarker. To address this gap, we developed Mitochondrial/Energy Flow Cytometry (MitE-Flo), a method that evaluates the contributions of glycolysis, fatty acid oxidation (FAO), and oxidative phosphorylation (OXPHOS) to cellular ATP content. In models of mitochondrial disease due to complex I or complex IV deficiency, we identified impaired OXPHOS with a compensatory shift to glycolysis. To define the utility of ATP monitoring in immunometabolism research, we analyzed previously inaccessible cell populations: light zone (LZ) and dark zone (DZ) germinal center (GC) B cells. Highly proliferative DZ B cells exhibited elevated ATP levels and a preference for FAO and OXPHOS over glycolysis, with uniform increased activity across ETC complexes. In contrast, less proliferative LZ B cells showed lower ATP levels and an equal reliance on glycolysis and OXPHOS. Using ATP as a biomarker to define metabolic dependences provides valuable insights into disease states and elusive immune cell subtypes, thereby enhancing the metabolism research toolkit. Biological sciences/Biochemistry Biological sciences/Immunology Biological sciences/Physiology Bioenergetics MitE-Flo (Mitochondrial/Energy Flow Cytometry) ATP immunometabolism influenza Full Text Additional Declarations The authors declare no competing interests. Supplementary Files SupplementaryMaterials.docx Cite Share Download PDF Status: Posted Version 2 posted You are reading this latest preprint version Show more versions Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-4836421","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Article","associatedPublications":[],"authors":[{"id":338882364,"identity":"4df0cfe7-0da7-4a39-8101-88352b2d68f4","order_by":0,"name":"Peter 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