Abstract
Organotypic and microphysiological culture of primary human tissues and cancers has emerged as a powerful set of technologies that allow to faithfully mimic cellular metabolism and functions ex vivo. The predominant 3D culture methods include spheroids and microfluidic chips. These cultures use low cell numbers and culture volumes, which, however, poses important limitations for the available amounts of sample for downstream analyses. Here, we describe a detailed method for the measurement of glucose consumption dynamics in organotypic culture using a bienzymatic colorimetric assay that accurately quantifies glucose levels using nanoliter input volumes. As an example we utilize spheroids consisting of primary human hepatocytes. The assay has been carefully optimized and benchmarked and is compatible with both longitudinal and high-throughput screening in both static and perfused conditions. The method is straightforward and only requires a microplate reader capable of running absorbance kinetic measurements.
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Acknowledgments
The work in the laboratory is funded by the Swedish Research Council [grant agreement numbers: 2016-01153, 2016-01154, and 2019-01837] and by the Swedish Strategic Research Programmes in Diabetes (SFO Diabetes) and Stem Cells and Regenerative Medicine (SFO StratRegen).
Competing Interests
VML is CEO and shareholder of HepaPredict AB, cofounder and shareholder of PersoMedix AB and discloses consultancy work for Enginzyme AB. AMK does not have competing interests to disclose.
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Kemas, A.M., Lauschke, V.M. (2022). Measuring Glucose Consumption and Gluconeogenesis in 3D Human Tissue Cultures with Nanoliter Input Volumes. In: Norberg, H., Norberg, E. (eds) Autophagy and Cancer. Methods in Molecular Biology, vol 2445. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2071-7_21
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DOI: https://doi.org/10.1007/978-1-0716-2071-7_21
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