An Fe3O4-Au heterodimer nanoparticle-based lateral flow assay for rapid and simultaneous detection of multiple influenza virus nucleic acids

Sensitive, convenient and rapid detection and subtyping of influenza viruses are crucial for timely treatment and management of infected people. Compared with antigen detection, nucleic acid detection has higher specificity and can shorten the detection window. Hence, in this work, we improved the lateral flow assay (LFA, one of the most promising user-friendly and on-site methods) to achieve detection and subtyping of H1N1, H3N2 and H9N2 influenza virus nucleic acids. Firstly, the antigen-antibody recognition mode was transformed into a nucleic acid hybridization reaction. Secondly, Fe3O4-Au heterodimer nanoparticles were prepared to replace frequently used Au nanoparticles to obtain better coloration. Thirdly, four lines were arranged on the LFA strip, which were three test (T) lines and one control (C) line. Three T lines were respectively sprayed by the DNA sequences complementary to one end of H1N1, H3N2 and H9N2 influenza virus nucleic acids, while Fe3O4-Au nanoparticles were respectively coupled with the DNA sequences complementary to the other end of H1N1, H3N2 and H9N2 nucleic acids to construct three kinds of probes. The C line was sprayed by the complementary sequences to the DNAs on all three kinds of probes. In the detection, by hybridization reaction, the probes were combined with their target nucleic acids which were captured by the corresponding T lines to form color bands. Finally, according to the position of the color bands and their grey intensity, simultaneous qualitative and semi-quantitative detection of the three influenza virus nucleic acids was realized. The detection results showed that this multi-channel LFA had good specificity, and there was no significant cross reactivity among the three subtypes of influenza viruses. The simultaneous detection achieved comparable detection limits with individual detections. Therefore, this multi-channel LFA had good application potential for sensitive and rapid detection and subtyping of influenza viruses.