{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,4,20]],"date-time":"2026-04-20T12:20:56Z","timestamp":1776687656112,"version":"3.51.2"},"reference-count":53,"publisher":"MDPI AG","issue":"4","license":[{"start":{"date-parts":[[2015,4,15]],"date-time":"2015-04-15T00:00:00Z","timestamp":1429056000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100000038","name":"Natural Sciences and Engineering Research Council of Canada","doi-asserted-by":"publisher","award":["400953"],"award-info":[{"award-number":["400953"]}],"id":[{"id":"10.13039\/501100000038","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100015668","name":"Ontario Ministry of Research and Innovation","doi-asserted-by":"publisher","award":["500327"],"award-info":[{"award-number":["500327"]}],"id":[{"id":"10.13039\/501100015668","id-type":"DOI","asserted-by":"publisher"}]},{"name":"Turkey Farmers of Ontario","award":["323"],"award-info":[{"award-number":["323"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Sensors"],"abstract":"<jats:p>Influenza pandemics cause millions of deaths worldwide. Effective surveillance is required to prevent their spread and facilitate the development of appropriate vaccines. In this study, we report the fabrication of a homogenous fluorescence-quenching-based assay for specific and sensitive detection of influenza virus surface antigen hemagglutinins (HAs). The core of the assay is composed of two nanoprobes namely the glycan-conjugated highly luminescent quantum dots (Gly-QDs), and the HA-specific antibody-modified gold nanoparticle (Ab-Au NPs). When exposed to strain-specific HA, a binding event between the HA and the two nanoprobes takes place, resulting in the formation of a sandwich complex which subsequently brings the two nanoprobes closer together. This causes a decrease in QDs fluorescence intensity due to a non-radiative energy transfer from QDs to Au NPs.  A resulting correlation between the targets HA concentrations and fluorescence changes can be observed. Furthermore, by utilizing the specific interaction between HA and glycan with sialic acid residues, the assay is able to distinguish HAs originated from viral subtypes H1 (human) and H5 (avian). The detection limits in solution are found to be low nanomolar and picomolar level for sensing H1-HA and H5-HA, respectively. Slight increase in assay sensitivity was found in terms of detection limit while exposing the assay in the HA spiked in human sera solution. We believe that the developed assay could serve as a feasible and sensitive diagnostic tool for influenza virus detection and discrimination, with further improvement on the architectures.<\/jats:p>","DOI":"10.3390\/s150408852","type":"journal-article","created":{"date-parts":[[2015,4,15]],"date-time":"2015-04-15T10:51:24Z","timestamp":1429095084000},"page":"8852-8865","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":19,"title":["A Homogenous Fluorescence Quenching Based Assay for Specific and Sensitive Detection of Influenza Virus A Hemagglutinin Antigen"],"prefix":"10.3390","volume":"15","author":[{"given":"Longyan","family":"Chen","sequence":"first","affiliation":[{"name":"BioNano Laboratory, School of Engineering, University of Guelph, Guelph, ON N1G 2W1, Canada"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-0990-0235","authenticated-orcid":false,"given":"Suresh","family":"Neethirajan","sequence":"additional","affiliation":[{"name":"BioNano Laboratory, School of Engineering, University of Guelph, Guelph, ON N1G 2W1, Canada"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2015,4,15]]},"reference":[{"key":"ref_1","unstructured":"WHO (2014). Available online: http:\/\/www.who.int\/mediacentre\/factsheets\/fs211\/en\/."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"243","DOI":"10.1128\/CMR.00037-06","article-title":"Avian influenza virus (H5N1): A threat to human health","volume":"20","author":"Peiris","year":"2007","journal-title":"Clin. Microbiol. Rev."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"531","DOI":"10.1146\/annurev.biochem.69.1.531","article-title":"Receptor binding and membrane fusion in virus entry: The influenza hemagglutinin","volume":"69","author":"Skehel","year":"2000","journal-title":"Annu. Rev. Biochem."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"81","DOI":"10.1016\/S1286-4579(00)01351-4","article-title":"The 1918 Spanish influenza: Integrating history and biology","volume":"3","author":"Reid","year":"2001","journal-title":"Microb. Infect."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"1143","DOI":"10.1016\/j.jmb.2005.11.002","article-title":"Glycan microarray analysis of the hemagglutinins from modern and pandemic influenza viruses reveals different receptor specificities","volume":"355","author":"Stevens","year":"2006","journal-title":"J. Mol. Biol."},{"key":"ref_6","first-page":"1","article-title":"Evaluating the control of HPAIV H5N1 in Vietnam: Virus transmission within infected flocks reported before and after vaccination","volume":"6","author":"Pfeiffer","year":"2010","journal-title":"BMC Vet. Res."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"156","DOI":"10.1007\/s00216-004-2927-0","article-title":"Detection of influenza virus: Traditional approaches and development of biosensors","volume":"381","author":"Amano","year":"2005","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"375","DOI":"10.1002\/rmv.370","article-title":"Molecular diagnosis of influenza","volume":"12","author":"Ellis","year":"2002","journal-title":"Rev. Med. Virol."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"341","DOI":"10.1016\/j.cimid.2008.01.009","article-title":"Conventional and future diagnostics for avian influenza","volume":"32","author":"Charlton","year":"2009","journal-title":"Comp. Immunol. Microbiol. Infect. Dis."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"517","DOI":"10.2307\/1592678","article-title":"Evaluation of a competitive ELISA for detection of antibodies against avian influenza virus nucleoprotein","volume":"42","author":"Zhou","year":"1998","journal-title":"Avian Dis."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"937","DOI":"10.1128\/JCM.37.4.937-943.1999","article-title":"Detection of antibody to avian influenza a (H5N1) virus in human serum by using a combination of serologic assays","volume":"37","author":"Rowe","year":"1999","journal-title":"J. Clin. Microbiol."},{"key":"ref_12","unstructured":"WHO Global Influenza Surveillance Network (2011). Manual for the Laboratory Diagnosis and Virological Surveillance of Influenza, WHO Press."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"361","DOI":"10.1016\/0042-6822(83)90150-2","article-title":"Receptor determinants of human and animal influenza virus isolates: Differences in receptor specificity of the H3 hemagglutinin based on species of origin","volume":"127","author":"Rogers","year":"1983","journal-title":"Virology"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"155","DOI":"10.1016\/0168-1702(93)90056-S","article-title":"Influenza virus strains selectively recognize sialyloligosaccharides on human respiratory epithelium; the role of the host cell in selection of hemagglutinin receptor specificity","volume":"29","author":"Couceiro","year":"1993","journal-title":"Virus Res."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"317","DOI":"10.1016\/0042-6822(89)90249-3","article-title":"Receptor binding properties of human and animal H1 influenza virus isolates","volume":"173","author":"Rogers","year":"1989","journal-title":"Virology"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"2773","DOI":"10.1016\/j.bmc.2004.02.042","article-title":"Binding of an influenza A virus to a neomembrane measured by surface plasmon resonance","volume":"12","author":"Critchley","year":"2004","journal-title":"Bioorg. Med. Chem."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"66","DOI":"10.1016\/j.aca.2008.06.005","article-title":"Monitoring of influenza virus hemagglutinin in process samples using weak affinity ligands and surface plasmon resonance","volume":"623","author":"Mandenius","year":"2008","journal-title":"Anal. Chim. Acta"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"5641","DOI":"10.1021\/ac401085c","article-title":"Attomolar detection of influenza a virus hemagglutinin human h1 and avian H5 using glycan-blotted field effect transistor biosensor","volume":"85","author":"Hideshima","year":"2013","journal-title":"Anal. Chem."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"5080","DOI":"10.1016\/j.actbio.2012.09.027","article-title":"Neu5Ac\u03b12,6Gal and Neu5Ac\u03b12,3Gal receptor specificities on influenza viruses determined by a waveguide-mode sensor","volume":"9","author":"Gopinath","year":"2013","journal-title":"Acta Biomater."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"236","DOI":"10.1016\/j.bios.2012.10.067","article-title":"Colorimetric viral detection based on sialic acid stabilized gold nanoparticles","volume":"42","author":"Lee","year":"2013","journal-title":"Biosens. Bioelectron."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"109","DOI":"10.1007\/s00216-008-2435-8","article-title":"Nanobioanalytical luminescence: F\u00f6rster-type energy transfer methods","volume":"393","author":"Roda","year":"2009","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"16660","DOI":"10.3390\/s121216660","article-title":"Nucleic acid sandwich hybridization assay with quantum dot-induced fluorescence resonance energy transfer for pathogen detection","volume":"12","author":"Chou","year":"2012","journal-title":"Sensors"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"83","DOI":"10.1016\/j.aca.2012.02.030","article-title":"Detection of influenza A virus based on fluorescence resonance energy transfer from quantum dots to carbon nanotubes","volume":"723","author":"Tian","year":"2012","journal-title":"Anal. Chim. Acta"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"806","DOI":"10.1002\/cphc.200800504","article-title":"Positively charged compact quantum dot\u2013DNA complexes for detection of nucleic acids","volume":"10","author":"Lee","year":"2009","journal-title":"Chem. Phys. Chem."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"124","DOI":"10.1002\/cyto.10121","article-title":"Long wavelength fluorophores and cell-by-cell correction for autofluorescence significantly improves the accuracy of flow cytometric energy transfer measurements on a dual-laser benchtop flow cytometer","volume":"48","author":"Nagy","year":"2002","journal-title":"Cytometry"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"630","DOI":"10.1038\/nmat961","article-title":"Self-assembled nanoscale biosensors based on quantum dot FRET donors","volume":"2","author":"Medintz","year":"2003","journal-title":"Nat. Mater."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"301","DOI":"10.1021\/ja037088b","article-title":"Fluorescence resonance energy transfer between quantum dot donors and dye-labeled protein acceptors","volume":"126","author":"Clapp","year":"2003","journal-title":"J. Am. Chem. Soc."},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1609","DOI":"10.1007\/s00216-007-1703-3","article-title":"Quantum dots as donors in fluorescence resonance energy transfer for the bioanalysis of nucleic acids, proteins, and other biological molecules","volume":"391","author":"Algar","year":"2008","journal-title":"Anal. Bioanal. Chem."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"4562","DOI":"10.1002\/anie.200503873","article-title":"Materials for fluorescence resonance energy transfer analysis: beyond traditional donor\u2013acceptor combinations","volume":"45","author":"Sapsford","year":"2006","journal-title":"Angew. Chem. Int. Ed."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"18","DOI":"10.1016\/S1748-0132(07)70016-6","article-title":"Au nanoparticles target cancer","volume":"2","author":"Jain","year":"2007","journal-title":"Nano Today"},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"3637","DOI":"10.1002\/chem.200701871","article-title":"A new nanobiosensor for glucose with high sensitivity and selectivity in serum based on fluorescence resonance energy transfer (FRET) between CdTe quantum dots and Au nanoparticles","volume":"14","author":"Tang","year":"2008","journal-title":"Chem. Eur. J."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"168","DOI":"10.1016\/j.bios.2012.04.007","article-title":"A new nanoprobe based on FRET between functional quantum dots and gold nanoparticles for fluoride anion and its applications for biological imaging","volume":"36","author":"Xue","year":"2012","journal-title":"Biosens. Bioelectron."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"43","DOI":"10.1016\/j.aca.2012.12.011","article-title":"A high-throughput homogeneous immunoassay based on F\u00f6rster resonance energy transfer between quantum dots and gold nanoparticles","volume":"763","author":"Qian","year":"2013","journal-title":"Anal. Chim. Acta"},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"385","DOI":"10.1016\/j.talanta.2014.03.033","article-title":"Efficient fluorescence resonance energy transfer between oppositely charged CdTe quantum dots and gold nanoparticles for turn-on fluorescence detection of glyphosate","volume":"125","author":"Guo","year":"2014","journal-title":"Talanta"},{"key":"ref_35","first-page":"20","article-title":"Controlled nucleation for the regulation of the particle size in monodisperse gold suspensions","volume":"241","author":"Frens","year":"1973","journal-title":"Nature"},{"key":"ref_36","doi-asserted-by":"crossref","unstructured":"Hermanson, G.T., and Hermanson, G.T. (2007). Chapter 24\u2014Preparation of Colloidal Gold-Labeled Proteins, Bioconjugate Techniques, Academic Press. [2nd ed.].","DOI":"10.1016\/B978-0-12-370501-3.00024-2"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"5052","DOI":"10.1021\/nl501709s","article-title":"Fluorescence quenching of quantum dots by gold nanoparticles: a potential long range spectroscopic ruler","volume":"14","author":"Samanta","year":"2014","journal-title":"Nano Lett."},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"5291","DOI":"10.1016\/S0040-4020(02)00489-1","article-title":"A detailed examination of boronic acid-diol complexation","volume":"58","author":"Springsteen","year":"2002","journal-title":"Tetrahedron"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Ferrier, R.J., and Prasad, D. (1965). 1360. Boric acid derivatives as reagents in carbohydrate chemistry. Part VI. Phenylboronic acid as a protecting group in disaccharide synthesis. J. Chem. Soc., 7429\u20137432.","DOI":"10.1039\/jr9650007429"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"5512","DOI":"10.1021\/la035037m","article-title":"Infrared Detection of a phenylboronic acid terminated alkane thiol monolayer on gold surface","volume":"20","author":"Brewer","year":"2004","journal-title":"Langmuir"},{"key":"ref_41","doi-asserted-by":"crossref","unstructured":"Dulkeith, E., Morteani, A.C., Niedereichholz, T., Klar, T.A., Feldmann, J., Levi, S.A., van Veggel, F.C.J.M., Reinhoudt, D.N., M\u00f6ller, M., and Gittins, D.I. (2002). Fluorescence quenching of dye molecules near gold nanoparticles: radiative and nonradiative effects. Phys. Rev. Lett., 89.","DOI":"10.1103\/PhysRevLett.89.203002"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"709","DOI":"10.1007\/s10719-013-9477-7","article-title":"Binding kinetics of sulfatide with influenza A virus hemagglutinin","volume":"30","author":"Takahashi","year":"2013","journal-title":"Glycoconjugate J."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"195","DOI":"10.1016\/j.bios.2011.12.003","article-title":"Monitoring influenza hemagglutinin and glycan interactions using surface plasmon resonance","volume":"32","author":"Suenaga","year":"2012","journal-title":"Biosens. Bioelectron."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"1880","DOI":"10.1039\/c0ay00491j","article-title":"Detection of influenza viruses by a waveguide-mode sensor","volume":"2","author":"Gopinath","year":"2010","journal-title":"Anal. Methods"},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"362","DOI":"10.1016\/j.jviromet.2013.03.006","article-title":"Selection and characterization of DNA aptamers for use in detection of avian influenza virus H5N1","volume":"189","author":"Wang","year":"2013","journal-title":"J. Virol. Methods"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"4020","DOI":"10.1021\/ac702625d","article-title":"Attomole detection of hemagglutinin molecule of influenza virus by combining an electrochemiluminescence sensor with an immunoliposome that encapsulates a ru complex","volume":"80","author":"Egashira","year":"2008","journal-title":"Anal. Chem."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"312","DOI":"10.1016\/j.ab.2005.04.013","article-title":"Selection of RNA aptamers against human influenza virus hemagglutinin using surface plasmon resonance","volume":"342","author":"Misono","year":"2005","journal-title":"Anal. Biochem."},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"1885","DOI":"10.1021\/nn800285p","article-title":"Monitoring biomolecular interactions on a digital versatile disk: A bioDVD platform technology","volume":"2","author":"Gopinath","year":"2008","journal-title":"ACS Nano"},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"8932","DOI":"10.1016\/j.actbio.2013.06.016","article-title":"Aptamers that bind to the hemagglutinin of the recent pandemic influenza virus H1N1 and efficiently inhibit agglutination","volume":"9","author":"Gopinath","year":"2013","journal-title":"Acta Biomater."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"146","DOI":"10.1016\/j.electacta.2013.07.113","article-title":"Effect of the size of receptor in allergy detection using field effect transistor biosensor","volume":"110","author":"Hideshima","year":"2013","journal-title":"Electrochimi. Acta"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"173","DOI":"10.1016\/0022-1759(85)90074-2","article-title":"The importance of antibody affinity in the performance of immunoassays for antibody","volume":"78","author":"Steward","year":"1985","journal-title":"J. Immunol. Methods"},{"key":"ref_52","doi-asserted-by":"crossref","first-page":"798","DOI":"10.1038\/nn.3418","article-title":"Immunogold cytochemistry in neuroscience","volume":"16","author":"Ottersen","year":"2013","journal-title":"Nat. Neurosci."},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"261","DOI":"10.1146\/annurev.bioeng.4.112601.125916","article-title":"Physics and applications of microfluidics in biology","volume":"4","author":"Beebe","year":"2002","journal-title":"Annu. Rev. Biomed. Eng."}],"container-title":["Sensors"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1424-8220\/15\/4\/8852\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T20:44:47Z","timestamp":1760215487000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1424-8220\/15\/4\/8852"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2015,4,15]]},"references-count":53,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2015,4]]}},"alternative-id":["s150408852"],"URL":"https:\/\/doi.org\/10.3390\/s150408852","relation":{},"ISSN":["1424-8220"],"issn-type":[{"value":"1424-8220","type":"electronic"}],"subject":[],"published":{"date-parts":[[2015,4,15]]}}}