An Integrated Microfluidic Chip Array Biosensor for Simultaneous ERA-Based Multiplex Meat Adulteration Identification Powered by MnO2@PDANPs@AuNPs Nanocomposites

Abstract

Conventional strategies for food species authentication are severely limited by intensive laboratory procedures and poor quantitative reliability in complex multispecies adulteration scenarios. Herein, a standardized, fully integrated, and lab-free PMMA-based centrifugal microfluidic chip biosensor platform was developed by combining micromachining technologies of UV lithography and microdevice bonding, enabling quantitative multispecies meat adulteration detection. Conserved nuclear genes of chicken, duck, pork, beef, mutton, and horseflesh were systematically selected and applied to enzymatic recombinase amplification (ERA), realizing stable copy number enrichment and accurate meat content quantification. An innovative multifunctional MnO2@PDANPs@AuNPs nanocomposite exhibiting enhanced peroxidase-like catalytic activity and improved FRET capability was synthesized and combined with ERA amplification for colorimetric and fluorescence detection, with detection limits as low as 0.031% for colorimetric mode and 0.005% for fluorescence mode, respectively. The entire analytical workflow, including on-chip gene extraction, amplification, and nanomaterial-assisted signal readout, was seamlessly integrated within closed microfluidic chambers, allowing standardized multichannel and multitarget analysis without manual intervention. This work established an integrated, portable, and automation-enabled solution for standardized multispecies meat authentication, offering a universal approach for on-site quantitative food adulteration monitoring and gene diagnostics through appropriate conserved gene selection and primer design.