Glycan arrays for high throughput screening of viruses

Patent metadata

Field	Value
Publication number	US-10338069-B2
Application number	US-201114376837-A
Country	US
Kind code	B2
Filing date	Apr 12, 2011
Priority date	Apr 12, 2010
Publication date	Jul 2, 2019
Grant date	Jul 2, 2019

Abstract

Glycan arrays that can detect and distinguish between various sub-types and strains of influenza virus are provided. Methods for using the glycan arrays with assays using nanoparticle amplification technique are disclosed. Sandwich assays using gold nanoparticles conjugated to phage particles comprising influenza virus-specific antibodies for detecting multiple serotypes using a single reaction are provided. Plurality of glycans directed to specific target HA of influenza virus comprises the array. Detector molecules comprising noble metals conjugated to (a) phage display particles expressing antibodies against hemagglutinin and (b) neuraminidase binding agents are disclosed.

First claim

What is claimed is:

1. A method for detecting at least one influenza serotype in a sample suspected of or comprising one or more viruses, the method comprising the steps of:
a) providing a substrate having at least one type of glycan capture probe bound at a discrete location on the substrate, wherein the capture probes can bind to a specific influenza serotype target;
b) providing at least one type of nanoparticle probe conjugated to a detector moiety, wherein the detector moiety binds to the specific influenza serotype;
c) contacting the substrate with the sample suspected of comprising one or more viruses and the nanoparticle probe under conditions suitable for the binding of the glycan capture probes to the specific influenza serotype and the binding of the nanoparticle probe to the specific influenza serotype to form a complex at the discrete location on the substrate; and
d) detecting the presence or absence of the complex wherein the presence or absence of the complex is indicative of the presence or absence of the specific influenza serotype in the sample;
wherein the nanoparticle consists essentially of a noble metal, and
wherein the limit of detection of at least one influenza serotype in said sample suspected of or comprising one or more viruses is less than 10 4 viral particles.

2. The method of claim 1 , wherein the influenza serotype is selected from the group consisting of influenza A, influenza B, influenza A serotypes H1N1, H2N2, H3N1, H3N2, H5N1, H7N7, H1N2, H9N2, H7N2, H7N3, and H10N7.

3. The method of claim 1 , wherein the substrate comprises an array of a plurality of glycan capture probes that bind specifically to a plurality of different influenza serotypes.

4. The method of claim 3 , wherein the plurality of capture probes comprises one or more of sialosides which is selected from the group consisting of: Neu5Ac(α2-3)Gal(β1-3)GalNAcα, Neu5Ac(α2-3)Gal(β1-6)Manα, Neu5Ac(α2-3)Fuc(α1-4)Gal(β1-4)Glcβ, Neu5Ac(α2-6)Galβ, Neu5Ac(α2-6)GalNAcβ, Neu5Ac(α2-6)Gal(β1-4)GlcNAcβ, Neu5Ac(α2-6)Gal(β1-4)Glcβ, Neu5Ac(α2-6)Gal(β1-4)GlcNAcβ, and Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-3) Gal(β1-4)GlcNAc(β1-3)Gal(β1-4)GlcNAcβ.

5. The method of claim 4 , wherein the capture probes are able to differentially bind to influenza subtypes Cal/09 H1N1, Brisbane H1N1, Brisbane H3N2, and RG14 H5N1.

6. The method of claim 1 , wherein the detector moiety comprises an antibody fragment that binds to the specific influenza serotype.

7. The method of claim 6 , wherein the antibody fragment comprises a phage particle from a phage display, and the phage particle comprises a plurality of nanoparticles.

8. The method of claim 7 , wherein the antibody fragment recognizes a hemagglutinin (HA) characteristic of an influenza subtype.

9. The method of claim 1 , wherein the detector moiety comprises a molecule that binds specifically to one or more influenza serotypes.

10. The method of claim 6 , wherein the antibody fragment recognizes a neuraminidase (NA) characteristic of an influenza subtype.

11. The method of claim 9 , wherein the molecule is a neuraminidase binding agent.

12. The method of claim 11 , wherein the molecule is selected from Oseltamivir (Tamiflu), Zanamivir (Relenza), Laninamivir (Inavir), or Peramivir.

13. The method of claim 11 , wherein the neuraminidase binding agent detects co-infection by at least two different strains of influenza viruses.

14. A method for distinguishing between a first influenza virus serotype and a second influenza virus serotype in a sample comprising a first influenza virus serotype and a second influenza virus serotype, the method comprising the steps of:
a) providing a substrate having at least two different types of glycan capture probes bound at discrete locations on the substrate, wherein a first capture probe can bind to the first influenza virus serotype but not the second influenza virus serotype, and a second capture probe can bind to the second influenza virus serotype but not the first influenza virus serotype;
b) providing at least one type of nanoparticle probe comprising detector moieties, wherein the detector moieties on each type of nanoparticle probe has a configuration that can bind to both the first influenza virus serotype and the second influenza serotype;
c) contacting the substrate with the sample under conditions suitable for binding of the first influenza virus serotype and the second influenza virus serotype in the sample to the glycan capture probes, and under suitable conditions for the binding of the nanoparticle probe to the first influenza serotype or second influenza serotype;
d) contacting the first influenza virus serotype and the second influenza virus serotype immobilized on the substrate with the nanoparticle probe under conditions that are effective for the binding of the detector moieties to the first influenza virus serotype and the second influenza virus serotype; and
e) detecting whether the nanoparticle probe binds to the first influenza virus serotype and/or the second influenza virus serotype at the discrete locations on the substrate where the first capture probe is located and where the second capture probe is located, wherein the presence or absence of the nanoparticle-first influenza virus serotype complex at the discrete location on the substrate where the first capture probe is located is indicative of the presence or absence of the specific first influenza virus serotype in the sample, and the presence or absence of the nanoparticle-second influenza virus serotype complex at the discrete location on the substrate where the second capture probe is located is indicative of the presence or absence of the specific second influenza virus serotype in the sample;
wherein the nanoparticle consists essentially of a noble metal.

15. The method of claim 1 or 14 , wherein the sample is blood, serum, anti-serum, monoclonal antibody preparation, lymph, plasma, saliva, urine, semen, breast milk, ascites fluid, tissue extract, cell lysate, cell suspension, viral suspension, nasopharyngeal aspirate, or a combination thereof.

16. The method of claim 14 , wherein the detector moiety comprises an antibody fragment that binds to the first influenza serotype or the second influenza serotype.

17. The method of claim 16 , wherein the antibody fragment thereof comprises a phage particle from a phage display, and the phage particle comprises a plurality of nanoparticles.

18. The method of claim 14 , wherein the captured nanoparticle-first influenza virus serotype or nanoparticle-second influenza virus serotype complex is detected by photonic, electronic, acoustic, opto-acoustic, gravity, electro-chemical, electro-optic, mass-spectrometric, enzymatic, chemical, biochemical, or physical means.

19. The method of claim 1 or 14 , wherein the nanoparticles are made of gold or silver.

20. The method of claim 19 , wherein the nanoparticles are made of gold.

21. The method of claim 1 or 14 , wherein the substrate is a magnetic bead.

22. The method of claim 1 or 14 , wherein the substrate has a planar surface.

23. The method of claim 1 or 14 , wherein the substrate is made of glass, quartz, ceramic, or plastic.

24. The method of claim 1 or 14 , wherein the detecting comprises contacting the substrate with specific metal stain.

25. The method of claim 1 or 14 , wherein the detecting comprises detecting light scattered by the nanoparticle.

26. The metho

Assignees

• Wong Chi Huey
• Wu Chung Yi
• Liang Chi Hui
• Yang An Suei
• Academia Sinica

Inventors

• Wong Chi-Huey
• Wu Chung-Yi
• Liang Chi-Hui
• Yang An-Suei

Classifications

• G01N33/56983 Primary

  Viruses · CPC title

• G01N33/53

  Immunoassay; Biospecific binding assay; Materials therefor · CPC title

• G01N2400/00

  Assays, e.g. immunoassays or enzyme assays, involving carbohydrates · CPC title

• G01N2400/38

  Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence, e.g. gluco- or galactomannans, Konjac gum, Locust bean gum or Guar gum · CPC title

• G01N33/54346

  Nanoparticles · CPC title