Hydrogen peroxide sensitive hemoglobin-capped gold nanoclusters as a fluorescence enhancing sensor for the label-free detection of glucose

Abstract

In this paper, a novel label-free fluorescent detection system based on blue-emitting gold nanoclusters capped by hemoglobin (Hb-AuNCs) was designed as a fluorescence enhancing–quenching (on–off) sensor for the direct detection of low levels of glucose in serum samples. In the presence of glucose as a substrate and glucose oxidase (GOx) as catalyst, H₂O₂ is produced that can intensively enhance the fluorescence intensity of Hb-AuNPs. In the presence of hydrogen peroxide, an intramolecular electron transfer to Hb-AuNCs takes place under heme degradation and/or iron release from Hb which is accompanied by the fluorescence enhancement of the nanoclusters. The DLS measurement, X-ray photoelectron spectroscopy patterns, and fluorescence and UV-visible spectra well supported this process. Upon the addition of H₂O₂, the fluorescence intensity enhanced linearly over the range of 0.5 μM to 700 μM with a limit of detection of 0.21 μM. The specific interaction of H₂O₂ with the sensing probe, allows it to develop a sensitive and selective glucose detection system by using glucose oxidase (GOx) with no need of complicated enzyme immobilization or any modification of the Hb-AuNCs. With this approach, the fluorescence quenching results in two linear dynamic ranges of 5 to 100 μM, and 100 to 1000 μM with a limit of detection of 1.65 μM for glucose detection. The designed fluorescent sensing system was then applied to the determination of H₂O₂ in two rain water samples and to five human serum samples for glucose detection. The Hb-AuNCs could be an effective candidate for new types of luminescence biosensors in the future, due to their fascinating features such as good water solubility, biocompatibility, excellent stability and potential applicability, and also to other H₂O₂-producing enzymatic reactions. In addition, the effective electron transfer between AuNCs and the heme group of Hb in the presence of H₂O₂, makes the system promising for electrochemical applications.