Development of an optical sandwich ELONA using a pair of DNA aptamers for yellow fever virus NS1

Here, we proposed an enzyme-linked oligonucleotide assay (ELONA) for yellow fever (YF) diagnosis that uses a pair of aptamers, YFns1-4 and YFns1-31. The aptamers were selected to specifically bind to nonstructural protein 1 (NS1), which is secreted at a high concentration after YF infection. We applied the aptamers which did not interfere with each other on binding to the NS1 in a sandwich ELONA. In the assay, the best detection sensitivity was obtained when the combination of YFns1-31 as a capture aptamer and YFns1-4 as a detect aptamer was used. The sensitivity could be attributed to the results of the direct ELONA with each YFns1-4 and YFns1-31; a great absorbance intensity and a broad detectable range of NS1, respectively. The sandwich ELONA achieved a low detection limit of 0.85 nM in buffer and was highly specific to the YFV-NS1 as its detection signals were significantly distinct from those of other flavivirus-derived NS1. In addition, the assay showed a desirable sensitivity in serum-spiked condition. Our developed sandwich ELONA can be a new practical and applicable serological diagnostics in YF endemic regions where other flaviviruses coexist and facilities for complex diagnostic tests are lacking.