Analytical chemistryDec 19, 2025

Highly Sensitive Detection of Multiple Food Poisoning Germs Using CRISPR and a Hand-Operated Microfluidic Device with a Built-in Pump

CRISPR Gene Editing Weekly Brief ↗PubMed ↗DOI ↗

Updated Jan 3, 2026

Abstract

A limit of detection (LOD) of 1.01 CFU/mL for Listeria monocytogenes was achieved using a novel detection method.

The method utilizes a recombinase polymerase amplification (RPA)-CRISPR-Cas12a approach for detecting foodborne pathogens.
Target-specific CRISPR RNAs (crRNAs) are employed in conjunction with a reconfigurable microfluidic device.
The microfluidic device includes a modular pressurizing pump (MoPP) that enhances reusability and reduces cross-contamination risks.
Testing confirmed the method's effectiveness with genomic DNA extracted from pathogen-spiked milk samples.
The detection system is designed for user-defined fluidic routing and multiplexed assay workflows.

Simplified

Foodborne pathogens pose a serious threat to global health and the economy, causing gastrointestinal illnesses and potentially leading to fatalities. Here, we present a recombinase polymerase amplification (RPA)-CRISPR-Cas12a-based method for the detection of foodborne pathogens using target-specific CRISPR RNAs (crRNAs) on a reusable, reconfigurable finger-actuated microfluidic device. Unlike previous finger-actuated pushbutton-based microfluidic devices, the device incorporates a modular pressurizing pump (MoPP), a standalone, reconfigurable actuation module that not only enhances reusability and reduces cross-contamination risks but also provides a flexible interface that allows user-defined fluidic routing and multiplexed assay workflows. Using a MoPP-integrated finger-actuated microfluidic device, the RPA-CRISPR-Cas12a-based detection of three foodborne pathogens was validated with an optimized crRNA and RPA primer sequence. Genomic DNA (gDNA) extracted from pathogen-spiked milk samples further demonstrated real-world applicability, achieving a limit of detection (LOD) of 1.62, 1.84, and 1.01 CFU/mL forO157:H7,spp., and, respectively. The developed microfluidic RPA-CRISPR-Cas12a-based detection platform is expected to be a reconfigurable, user-friendly, and highly sensitive point-of-care testing system for monitoring foodborne pathogens throughout the food supply chain. Escherichia coli Salmonella Listeria monocytogenes

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Highly Sensitive Detection of Multiple Food Poisoning Germs Using CRISPR and a Hand-Operated Microfluidic Device with a Built-in Pump

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