Ultra-efficient label-free cell death detection based on multifunctional colorimetric fluorescent in-situ silver...

In this study an efficient label-free multifunctional probe based on in situ hemoglobin-stabilized silver nanocluster/graphene nanohybridzymes (Hb-AgNCs/graphene NHZs) was designed and employed for dual-modal naked-eye colorimetric and fluorometric detection of cell death biomarkers involved in ferroptosis, cuproptosis, and apoptosis. The interesting feature of developed NHZs is the colorimetric speciation of "turn-on" Fe²⁺, Cu²⁺, and Cyt c and "turn-off" cysteine (Cys) and glutathione (GSH), and its speciation ability of Fe²⁺ from Fe³⁺as well. The presence of analytes causes a colorimetric limit of detection of 10.8-191 nM and fluorometric detection limit from 0.25 to 121 nM in the appropriate linear ranges, with fast times ranging 5-20 min. The designed NHZs were successfully employed for fluorescence imaging of HT29 human colon cancer and also quantitatively validated in just 2 μL of cell lysates, environmental, and biological samples. Compared to the semi-quantitative conventional Western blot and other cell death assays, the novelty of presented probe is low cost, simple preparation, high sensitivity, multifunctionallity, and dual-modality with the speciation and quantitative ability without using any targeting agent, which makes it appropriate for dual-modal tracking cell death processes and drug evaluation in a very short time.