Simultaneous Dual-Sensing Platform Based on Aptamer-Functionalized DNA Hydrogels for Visual and Fluorescence Detection of Chloramphenicol and Aflatoxin M1

In this study, a chloramphenicol and aflatoxin M1 aptamer-functionalized DNA hydrogel was designed for the simultaneous detection of chloramphenicol and aflatoxin M1 for the first time. The acrydite-modified chloramphenicol aptamer sequence was used to synthesize the DNA hydrogel and for visual detection of chloramphenicol depending on the gel-to-sol transition of the target-responsive DNA hydrogel. The DNA hydrogel formulation was set as follows: 60% of each linear polyacrylamide-DNA conjugate and 40% of acrylamide and chloramphenicol aptamer/DNA strand-1 at a molar ratio of 1:1, and the lowest concentration of chloramphenicol leading to gel dissociation was 1.0 nM at 25 °C. Furthermore, the formalized aptamer-functionalized DNA hydrogel was used to detect aflatoxin M1 by measuring the recovery of the fluorescence signal that was quenched when the FAM-labeled aflatoxin M1 aptamer and BHQ1-labeled DNA strand-2 were hybridized to form a double-stranded DNA in the network of hydrogel. The detection platform was successfully applied to the detection of chloramphenicol and aflatoxin M1, both in aqueous solution and in milk. The aptamer-functionalized DNA hydrogel had detection (LOD) and quantification limits (LOQ) for aflatoxin M1 as 1.7 and 5.2 nM, respectively. Using two aptamer sequences with high affinity and specificity, the dual-sensing platform based on the DNA hydrogel achieved higher selectivity for chloramphenicol and aflatoxin M1, which demonstrated its potential as a reliable simultaneous detection platform against two different targets for monitoring food safety.