Engineered Affinity Proteins Enable Sensitive Label-Free Electrochemical Detection

The increasing need for sensitive and rapid diagnostic tests for disease biomarkers has become a critical focus in medical research and development. This study evaluates the efficacy of an engineered affinity protein, the hyperthermostable binding scaffold rcSso7d, produced using the Rapid Affinity Pair Identification via Directed Selection (RAPIDS) method, in comparison to traditional antibodies for label-free electrochemical detection. Our research establishes that the binding scaffold rcSso7d can be used as a bioreceptor for electrochemical detection, using magnetic beads to immobilize the affinity protein on screen-printed electrodes. We demonstrate that this engineered affinity protein exhibits superior sensitivity compared to a high-quality traditional antibody in detecting the SARS-CoV-2 nucleocapsid protein (N-protein, NP), a key biomarker for COVID-19. The developed electrochemical sensor, based on the rcSso7d protein, achieves a detection limit of 10 pg/mL. These findings suggest that the rcSso7d protein holds significant potential as an affinity agent in electrochemical diagnostic assays and highlights the importance of exploring alternative molecular scaffolds in electrochemical biosensing.