Quantitative Analysis of Cellular Ferroptosis Using Dual-Channel Colorimetry Based on Gold Nanoparticle Catalytic Activity.

Abstract

This study constructed a dual-substrate colorimetry based on the catalytic activity single-stranded DNA (ssDNA), i.e., 5'-GGT TGG TGT GGT TGG-3', functionalized gold nanoparticles (AuNPs-ssDNA). By utilizing the peroxidase-mimicking activity of AuNPs and generating differential colorimetric response fingerprints through dual substrates o-phenylenediamine (OPD) and 3-amino-9-ethylcarbazole (AEC), simultaneous discrimination and quantitative detection of Fe²⁺ and Fe³⁺ were achieved. Characterization results showed that the synthesized AuNPs had an average size of about 12 nm. After functionalization, the average diameter of the formed AuNPs-ssDNA complexes increased to 13 nm, and the absorption peak position remained unchanged, indicating that ssDNA modification did not affect their dispersity and morphology. This method, combined with multivariate calibration models, can extract quantitative information from complex response patterns, with a linear detection range for both Fe²⁺ and Fe³⁺ from 0.01 to 100 μM. In practical application validation, the method was successfully used to detect iron ion concentrations in A549 cells treated with the ferroptosis inducer GPX4-IN-7. The results showed high consistency (P ≤ 0.001) with those obtained using a commercial FerroOrange detection kit, demonstrating the potential of this method for practical application in complex biological systems.