Microfluidics-free hydrogel-based digital PCR for detection of Salmonella in food.

Salmonella is a major foodborne pathogen, underscoring the need for rapid, sensitive, and accurate quantitative detection methods for food safety monitoring. Most existing digital PCR technologies rely on microfluidic platforms, specialized instruments, and high operating costs. To address these limitations, a one-step and microfluidics-free digital PCR method based on polyacrylamide hydrogel (PAM-dPCR) was developed for the detection and absolute quantification of Salmonella in food samples. Without the need for microfluidic chips, this method relies on the three-dimensional polyacrylamide hydrogel network to provide a spatially confined reaction matrix, within which acrydite-modified primers are covalently incorporated and immobilized. This hydrogel-based confinement localized the amplification of individual target molecules to their original locations, generating discrete in situ fluorescent amplification spots and enabling absolute quantification of the target bacterium. The results showed that PAM-dPCR achieved a detection limit as low as 1 copy/μL, with the entire assay completed within 1.5 h, demonstrating high sensitivity and ease of operation. In artificially contaminated food samples, the recovery rates of this method ranged from 86.17% to 99.56%, indicating good accuracy and applicability. In conclusion, PAM-dPCR enables rapid and highly sensitive detection of Salmonella in food samples and demonstrates specificity under the tested conditions, highlighting its potential for food safety monitoring.