Multiplex nanozymatic biosensing of Salmonella on a finger-actuated microfluidic chip

A colorimetric biosensor was elaboratively designed for fast, sensitive and multiplex bacterial detection on a single microfluidic chip using immune magnetic nanobeads for specific bacterial separation, immune gold@platinum palladium nanoparticles for specific bacterial labeling, a finger-actuated mixer for efficient immunoreaction and two coaxial rotatable magnetic fields for magnetic nanobead capture (outer one) and magnet-actuated valve control (inner one). First, preloaded bacteria, nanobeads and nanozymes were mixed through a finger actuator to form nanobead-bacteria-nanozyme conjugates, which were captured by the outer magnetic field. After the inner magnetic field was rotated to successively wash the conjugates and push the H₂O₂-TMB substrate for resuspending these conjugates, colorless TMB was catalyzed into blue TMB_ox products, followed by color analysis using ImageJ software for bacterial determination. This simple biosensor enabled multiplex Salmonella detection as low as 9 CFU per sample in 45 min.