Biosensors typically comprise of two key components: the biological recognition element, that binds to an analyte of interest, and a transducer, that measures the binding event. Two different sections are used to attach a biological recognition element to the transducer: the anchoring group, which keeps the structure bound to the sensing surface, and a linker, which links the anchoring group and the recognition element together. Poly-histidine tags (His-tags) are commonly used linkers for biosensors because they selectively coordinate with divalent metal ions, enabling oriented immobilization of recognition elements. Despite their widespread use, the electrochemical and physiological stability of His-tags remain unexplored. In this work, we evaluated the electrochemical stability and degradation profile of His-tags on a modified glassy carbon surface in phosphate-buffered saline and biological media using square wave voltammetry (SWV). Glassy carbon electrodes were functionalized with a benzoic acid layer, then activated using EDC/NHS coupling to attach amino-nitriloacetic acid (NTA), which serves as a chelating ligand. The NTA modified surface was then loaded with Ni
2+ to enable His-tag binding. The His-tag has been chemically modified with a Ferrocene redox probe, allowing monitoring via SWV at a frequency of 250Hz over a potential range from 500mV to 50mV vs Ag/AgCl. Initial measurements confirm the presence of the His-tag on the surface. However, signal intensity from the His-tag declined within hours, suggesting lack of stability. These findings underscore the limitations of His-tags for long-term, continuous biosensing applications and highlight the need for more robust linker chemistries.
Support was provided by a National Science Foundation EPSCoR award (#2119237).
