Detection of avian influenza virus by fluorescent DNA barcode-based immunoassay with sensitivity comparable to PCR.
In this paper, a coupling of fluorophore-DNA barcode and bead-based immunoassay for detecting avian influenza virus (AIV) with PCR-like sensitivity is reported. The assay is based on the use of sandwich immunoassay and fluorophore-tagged oligonucleotides as representative barcodes. The detection involves the sandwiching of the target AIV between magnetic immunoprobes and barcode-carrying immunoprobes. Because each barcode-carrying immunoprobe is functionalized with a multitude of fluorophore-DNA barcode strands, many DNA barcodes are released for each positive binding event resulting in amplification of the signal. Using an inactivated H16N3 AIV as a model, a linear response over five orders of magnitude was obtained, and the sensitivity of the detection was comparable to conventional RT-PCR. Moreover, the entire detection required less than 2 hr. The results indicate that the method has great potential as an alternative for surveillance of epidemic outbreaks caused by AIV, other viruses and microorganisms.
Cao
Cuong
C
DTU-Nanotech, Department of Micro and Nanotechnology, Technical University of Denmark, DTU Building 345 East, DK-2800 Kongens Lyngby, Denmark.
Dhumpa
Raghuram
R
Bang
Dang Duong
DD
Ghavifekr
Zohreh
Z
Høgberg
Jonas
J
Wolff
Anders
A
D000818:Animals; D015374:Biosensing Techniques; D002645:Chickens; D007118:Immunoassay; D009980:Influenza A virus / Q000235:genetics* / Q000472:pathogenicity; D005585:Influenza in Birds / Q000175:diagnosis* / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*; D013050:Spectrometry, Fluorescence