SALSA: a novel flow cytometry assay to detect ascorbate at the single-cell level.

ABSTRACT: Ascorbate (AA) is an essential antioxidant and enzymatic cofactor with emerging roles in epigenetic regulation, redox biology, and immune function. However, single-cell quantification of intracellular AA has remained technically challenging. Here, we present SALSA (Single-cell Ascorbate Level Sensing Assay), a novel flow cytometry-based method that enables sensitive, specific detection of intracellular AA at the single-cell level. Inspired by the mechanism of the in vitro AA assay, we identified 4,5-diaminofluorescein (DAF-2), a common nitric oxide (NO) probe, as a selective AA reporter. We showed that the chemical oxidation of AA into dehydroascorbic acid (DHA) facilitated its reaction with DAF-2 to form a highly fluorescent product. Surprisingly, the DAF-2-DHA adduct exhibits a red-shift

SUBMITTER: Lee B 

PROVIDER: S-EPMC12398858 | biostudies-literature | 2025 Oct

REPOSITORIES: biostudies-literature