An Ultra-High Throughput Method for ¹³C6-Fructose-6-Phosphate in Cells Using Acoustic Ejection Mass Spectrometry (AEMS)

As an endogenous compound, F6P is subject to endogenous interference. Isotopic-labeled compound 13C6-F6P is used to reduce endogenous interference and enhance specificity. The traditional LC-MS/MS method for 13C6-F6P is complex and time-consuming, often requiring derivatization, ion-pair reagents, pH adjustments of complex mobile phases, or other lengthy procedures, making them unsuitable for high-throughput screening (HTS). This study describes an ultra-high throughput method for detecting 13C6-F6P in cell matrices using AEMS technology. This method is rapid, high-throughput, sensitive, and reliable, aiding F6P-targeted PFKP research.Glucose-based aerobic glycolysis, also known as the Warburg effect, supplies energy for rapid tumor growth. The phosphorylation of fructose-6-phosphate (F6P) by phosphofructokinase-1 (PFK1) is a key rate-limiting step in the Warburg effect and a target for cancer therapy.