Hourglass-mimicking biosensor based on disposable centrifugal tube for bacterial detection in large-volume sample

An hourglass-mimicking biosensor was developed to detect target bacteria in 15 mL centrifugal tube using immune magnetic nanobeads to isolate the target from large-volume sample, gold@platinum nanozymes to label the target for amplification of biological signals, and a microplate reader/colorimetric card for determination of the target. First, a centrifugal tube with an iron ball framework was first coaxially assembled in the center of a Halbach ring magnet. After immune magnetic nanobeads, gold@platinum nanozymes and bacterial sample were mixed by repeated bottom-up of the tube using a stepper motor, nanobead-bacteria-nanozyme complexes were formed. Then, colorless H₂O₂-TMB was catalyzed by the nanozymes to produce blue TMBox. The color change was finally analyzed using the microplate reader or colorimetric card to determine bacterial concentration. This hourglass-mimicking biosensor could separate ∼95% targets from 10 mL bacteria sample and detect targets from 1.6 × 10¹ to 1.6 × 10⁶ CFU/mL in 1.0 h with low detection limit of 16 CFU/mL.