A Zwitterionic-Enabled One-Pot Electrochemical Strategy for Stable End-Point Detection of Isothermal Nucleic Acid Amplification

Abstract

Indicator-based electrochemical loop-mediated isothermal amplification (EC-LAMP) is highly attractive for portable nucleic acid testing, yet its practical deployment is fundamentally constrained by severe signal instability caused by amplification-induced interfacial fouling. Here, we report a zwitterionic interfacial stabilization strategy that enables robust, one-pot, label-free electrochemical end-point detection of isothermal nucleic acid amplification. By grafting poly(carboxybetaine methacrylate) (PCBMA) onto screen-printed carbon electrodes, nonspecific adsorption of methylene blue and amplification components is effectively suppressed, resulting in minimal background drift and highly reproducible signal-off readout under sealed reaction conditions. Using Staphylococcus aureus as a model target, the stabilized EC-LAMP platform achieves detection limits of 12 CFU mL⁻¹ in pure culture and 64 CFU mL⁻¹ in spiked milk samples within 30 min. More importantly, this work establishes interfacial engineering as a generalizable design principle for stabilizing indicator-based EC-LAMP systems, providing a practical pathway toward deployable electrochemical nucleic acid testing in resource-limited settings.