INTRODUCTION:
UCLA researchers in the Department of Molecular and Medical Pharmacology reported a novel single cell assay for profiling lactate production via the use of coupled enzyme reactions on surface-grafted resazurin molecules.
BACKGROUND:
The upregulation of aerobic glycolysis, known as the Warburg effect, is a hallmark of many cancers. Recent studies revealed that aerobic glycolysis is also capable of modulating cell-signaling networks. Understanding the interplay between Warburg effect and the oncogenic signaling will provide new insights into potential therapeutics. However, efforts in deciphering such interactions can be confounded by the intra- and inter-tumoral heterogeneity—a common feature of all cancers. In order to untangle the mechanistic links in heterogenous tumors, it is necessary to perform simultaneous measurements on a panel of oncogenic signaling phosphoproteins, glucose influx and lactate production at the single cell level. Dr Wei’s group at UCLA has previously developed integrated proteomic and metabolic single cell assays on the single cell barcode chip (SCBC) platform, where glycose influx and signaling protein activities were simultaneously quantified from single cells with fluorescence readouts.
Currently, quantification of lactate at the single cell level remains challenging. A major obstacle is the secretion nature of lactate, which renders those well-established uptake-based detection schemes incompatible. Another challenge is the lack of lactate-specific antibodies, due to their extremely small size and endogenous existence. Therefore, it is impractical to develop immuno-based lactate detection methods
INNOVATION:
UCLA researchers led by Prof. Wei Wei developed a novel single cell assay for quantifying lactate production from single cells using a surface-immobilized redox-labile fluorescent probe and coupled enzyme reactions. The immobilization of the redox-labile probes was achieved through chemical modification on resazurin, followed by bio-orthogonal click reactions. The lactate detection scheme was demonstrated to be sensitive and specific. The method was incorporated into the SCBC platform for simultaneous quantification of aerobic glycolysis activities and oncogenic signaling phosphoproteins in cancer.
POTENTIAL APPLICATIONS:
• Comprehensive analysis of cellular redox-active metabolites
ADVANTAGES:
• Single cell assay
• Simultaneously quantify aerobic glycolysis, oncogenic signaling phosphoproteins, and lactate production
• The immobilization methods retain the detection sensitivity and exhibit excellent stability
DEVELOPMENT-TO-DATE:
The researchers tested the platform on a glioblastoma cell line.