Abstract
Histamine produced by bacteria through decarboxylation of histidine in spoiled foods such as fish is known to cause food poisoning. Therefore, accurate and facile measurement of histamine is of practical importance. Using the recently discovered RNA aptamer that specifically recognizes histamine (A1-949 aptamer), we developed an aptasensor based on the structure-switching mechanism. Specifically, the aptamer A1-949 was fluorescently labeled at the 5′ end and hybridized with a short quencher DNA strand that is partially complementary to the aptamer. The quencher strand was modified with a fluorescence quencher at its 3′ terminus. Displacement of the quencher strand upon histamine binding results in an increased fluorescence. After optimizing the assay condition, the enantiomeric version of the aptasensor (L-RNA and L-DNA) was synthesized which could detect the achiral analyte with identical sensitivity and improved biochemical stability. The aptasensor performance was validated by measuring fish samples spiked with known concentrations of histamine. Finally, histamine content in spoiled fish samples was measured, and the results were compared with the measurements using a commercial enzymatic assay kit.
Highlights
Histamine produced by bacteria through decarboxylation of histidine in spoiled foods such as fish is known to cause food poisoning
We successfully selected an RNA aptamer (A1-949 aptamer) (Fig. 1a) that can bind histamine with high affinity and specificity from a pool of 3 × 1014 random sequences through systematic evolution of ligands by exponential enrichment (SELEX) aided by deep sequencing[18]
The recently discovered histamine aptamer A1-949 (Fig. 1a) binds histamine with a dissociation constant (Kd) of 370 nM while associating with L-histidine with Kd > 20 μM based on isothermal titration calorimetry (ITC)[18]
Summary
Histamine produced by bacteria through decarboxylation of histidine in spoiled foods such as fish is known to cause food poisoning. We employed the A1-949 aptamer to engineer a fluorescence-based aptasensor that can detect histamine concentration as low as 1 μM. The sensor performance was further evaluated in the presence of www.nature.com/scientificreports varying concentrations of histamine and histidine stereoisomers (Fig. 4a,b).
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