Selection and application of highly specific Salmonella typhimurium aptamers against matrix interference

In practical applications, the structure and performance of aptamers can be influenced by the presence of sample matrices, which interferes with the specific binding between the aptamer and its target. In this work, to obtain aptamer chains resistant to matrix interference, four typical food matrices were introduced as negative selection targets and selection environments in the process of selecting aptamers for Salmonella typhimurium using the systematic evolution of ligands by exponential enrichment (SELEX) technology. As a result, some highly specific candidate aptamers for Salmonella typhimurium (BB-34, BB-37, ROU-8, ROU-9, ROU-14, ROU-24, DAN-3, NAI-12, and NAI-21) were successfully obtained. Based on the characterization results of secondary structure, affinity, and specificity of these candidate aptamers, ROU-24 selected in the pork matrix and BB-34 selected in the binding buffer were chosen to develop label-free fluorescence aptasensors for the sensitive and rapid detection of the Salmonella typhimurium and verify the performance against matrix interference. The ROU-24-based aptasensor demonstrated a larger linear range and better specificity compared to the BB-34-based aptasensor. Meanwhile, the recovery rate of the ROU-24-based aptasensor in real sample detection (ranging from 94.2% to 110.7%) was significantly higher than that of the BB-34-based aptasensor. These results illustrated that the negative selection of food matrices induced in SELEX could enhance specific binding between the aptamer and its target and the performance against matrix interference. Overall, the label-free fluorescence aptasensors were developed and successfully validated in different foodstuffs, demonstrating a theoretical and practical basis for the study of aptamers against matrix interference.