动力磷酸化和糖酵解是哺乳动物新陈代谢中主要的产ATP途径。这两个途径之间的平衡常常会因为促进细胞活化、增殖或分化的刺激而发生变化，以执行细胞特定功能。然而，对这些能量代谢转换的测量一直以来主要是定性的，很难区分健康的、生理变化所致的能量转导或代谢功能失调引起的代偿反应。因此，我们提出了一种广泛适用的方法，通过使用SeaHorse XF分析仪的数据和经验转化因子，计算氧化磷酸化和糖酵解产生的ATP产生率。我们量化了巨噬细胞极化和癌细胞适应体外培养条件中观察到的生物能学变化。此外，我们还检测到在神经元去极化和T细胞受体活化过程中，ATP利用发生了实质性的变化，这些变化在稳态ATP测量中是不明显的。这种方法生成了一个单一的读数，可以直接比较活细胞中氧化磷酸化和糖酵解产生的ATP。此外，本文还提供了一个框架，以适应特定细胞系统或实验条件的计算。© 2023 作者公开发表遵循CC BY 4.0许可协议。

Oxidative phosphorylation and glycolysis are the dominant ATP-generating pathways in mammalian metabolism. The balance between these two pathways is often shifted to execute cell-specific functions in response to stimuli that promote activation, proliferation, or differentiation. However, measurement of these metabolic switches has remained mostly qualitative, making it difficult to discriminate between healthy, physiological changes in energy transduction or compensatory responses due to metabolic dysfunction. We therefore present a broadly applicable method to calculate ATP production rates from oxidative phosphorylation and glycolysis using Seahorse XF Analyzer data and empirical conversion factors. We quantify the bioenergetic changes observed during macrophage polarization as well as cancer cell adaptation to in vitro culture conditions. Additionally, we detect substantive changes in ATP utilization upon neuronal depolarization and T cell receptor activation that are not evident from steady-state ATP measurements. This method generates a single readout that allows the direct comparison of ATP produced from oxidative phosphorylation and glycolysis in live cells. Additionally, the manuscript provides a framework for tailoring the calculations to specific cell systems or experimental conditions.© 2023 The Authors. Published under the terms of the CC BY 4.0 license.