Signal Amplification and Near-Infrared Translation of Enzymatic Reactions by Nanosensors

Enzymatic reactions are used to detect analytes in a range of biochemical methods. To measure the presence of an analyte, the enzyme is conjugated to a recognition unit and converts a substrate into a (colored) product that is detectable by visible (VIS) light. Thus, the lowest enzymatic turnover that can be detected sets a limit on sensitivity. Here, we report that substrates and products of horseradish peroxidase (HRP) and beta-galactosidase change the near-infrared (NIR) fluorescence of (bio)polymer modified single-walled carbon nanotubes (SWCNTs). They translate a VIS signal into a beneficial NIR signal. Moreover, the affinity of the nanosensors leads to a higher effective local concentration of the reactants. This causes a non-linear sensor-based signal amplification and translation (SENSAT). We find signal enhancement up to approximate to 120x for the HRP substrate p-phenylenediamine (PPD), which means that reactions below the limit of detection in the VIS can be followed in the NIR (approximate to 1000 nm). The approach is also applicable to other substrates such as 3,3 '-5,5 '-tetramethylbenzidine (TMB). An adsorption-based theoretical model fits the observed signals and corroborates the sensor-based enhancement mechanism. This approach can be used to amplify signals, translate them into the NIR and increase sensitivity of biochemical assays. ELISA substrates and products modulate the near-infrared fluorescence of single-walled carbon nanotubes (SWCNTs) modified with specific (bio)polymers. The affinity of these nanosensors for the analyte increases the effective local concentration compared to normal absorption measurements. This causes a signal enhancement, which improves the readout of enzymatic reactions.**image