Real-Time Metabolic Detection in Living Cells Using Hyperpolarized 13C NMR.

The protocol presents a method to integrate an in-house-built bioreactor system with hyperpolarized ¹³C nuclear magnetic resonance (NMR) spectroscopy to measure pyruvate metabolism in live cells in real time. The protocol outlines the construction of a bioreactor system that allows precise timing of hyperpolarized substrate injection and NMR signal acquisition while maintaining a stable cell culture environment. The method reliably detects the metabolic conversion of pyruvate to lactate within a 60 s measurement window while maintaining a sufficient signal-to-noise ratio throughout the process. The in situ mixing capability of the system accelerates NMR signal detection and reduces T1 relaxation losses. The protocol also includes detailed instructions for the preparation of alginate gels to encapsulate cells and place them within the bioreactor. The proposed method exhibits improved reproducibility and kinetic accuracy compared to conventional methods, indicating potential applications in cancer metabolism studies and drug screening. Further refinements of the system, such as the adoption of alternative gel matrices, may improve its versatility in various biological studies.