Gullo_2025_Biosens.Bioelectron_282_117471

Terrorist attacks using chemical warfare agents have been unfortunately well documented in history, and among chemical warfare agents, nerve agents are the preferred ones due to their ease of synthesis and transport. A huge issue is the contamination of drinking water by adding nerve agents into aqueducts, making their on-line monitoring an urgent need to ensure the safety of drinking water. A compact and portable shamrock-like electrochemical inhibitive biosensor was fabricated by 3D printing the entire electrochemical cell and the flow cell for on-line nerve agent measurements. Paraoxon was selected as the nerve agent simulant and detected by harnessing its capability to inhibit irreversibly the butyrylcholinesterase enzyme. This enzyme was chemically immobilized onto the 3D printed electrode previously modified by drop casting with Carbon Black-Prussian Blue nanoparticles to ensure the detection of enzymatic by-product thiocholine at low applied potential (+300 mV vs. Ag/AgCl), with enhanced selectivity and sensitivity. This 3D printed analytical tool demonstrated a detection limit as low as 0.9 ppb and 1.6 ppb with a linear range of up to 20 ppb in standard solution and untreated tap water, respectively. The accuracy was evaluated by the recovery study, obtaining recovery values comprised in the range of 99-105 %. These results demonstrated the effectiveness of the combination of 3D printing technology, nanomaterials, and electrochemical sensing to deliver a customized on-line device able to work as a smart warning system for environmental surveillance and public health protection.