Glycan arrays for high throughput screening of viruses

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.

We claim: 
     
         1 - 49 . (canceled) 
     
     
         50 . An array, comprising:
 a) 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) one or more viruses comprising a specific influenza serotype bound to the corresponding glycan capture probe for the one or more viruses comprising a specific influenza serotype;   c) at least one type of nanoparticle probe conjugated to a detector moiety, wherein the detector moiety is bound to the specific influenza serotype;   wherein the at least one type of glycan capture probe, the one or more viruses comprising a specific influenza serotype, and the nanoparticle probe form a complex at the discrete location on the substrate, and   wherein the nanoparticle comprises a noble metal.   
     
     
         51 . The array of  claim 50 , 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. 
     
     
         52 . The array of  claim 50 , wherein the substrate comprises an array of a plurality of glycan capture probes that bind specifically to a plurality of different influenza serotypes. 
     
     
         53 . The array of  claim 52 , 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. 
     
     
         54 . The array of  claim 53 , wherein the capture probes are able to differentially bind to influenza subtypes Cal/09 H1N1, Brisbane H1N1, Brisbane H3N2, and RG14 H5N1. 
     
     
         55 . The array of  claim 50 , wherein the detector moiety comprises an antibody fragment that binds to the specific influenza serotype. 
     
     
         56 . The array of  claim 50 , wherein the detector moiety comprises an antibody fragment from a phage particle from a phage display, and the phage particle comprises a plurality of nanoparticles. 
     
     
         57 . The array of  claim 56 , wherein the antibody fragment recognizes a hemagglutinin (HA) characteristic of an influenza subtype. 
     
     
         58 . The array of  claim 50 , wherein the detector moiety comprises a molecule that binds specifically to one or more influenza serotypes. 
     
     
         59 . The array of  claim 55 , wherein the antibody fragment recognizes a neuraminidase (NA) characteristic of an influenza subtype. 
     
     
         60 . The array of  claim 58 , wherein the molecule is a neuraminidase binding agent selected from Oseltamivir (Tamiflu), Zanamivir (Relenza), Laninamivir (Inavir), or Peramivir. 
     
     
         61 . The array of  claim 60 , wherein the neuraminidase binding agent detects co-infection by at least two different strains of influenza viruses. 
     
     
         62 . The array of  claim 50 , wherein the number of viral particles is less than 10 4  viral particles. 
     
     
         63 . The array of  claim 50 , wherein the nanoparticles are made of gold or silver. 
     
     
         64 . The array of  claim 50 , wherein the substrate is a magnetic bead. 
     
     
         65 . The array of  claim 50 , wherein the substrate is made of glass, quartz, ceramic, or plastic. 
     
     
         66 . The array of  claim 50 , wherein the substrate is addressable. 
     
     
         67 . The array of  claim 50 , wherein a plurality of glycan capture probes, each of which can recognize a different target influenza serotype, are attached to the substrate in an array of discrete spots. 
     
     
         68 . The array of  claim 67 , wherein the plurality of glycan capture probes comprises a glycan structure of at least one molecule selected from: Neu5Ac(α2-3)Galβ, Neu5Ac(α2-3)Gal(β1-4)Glcβ, Neu5Ac(α2-3)Gal(β1-4)GlcNAcβ, Neu5Ac(α2-3)Gal(β1-4)(6S)GlcNAcβ, Neu5Ac(α2-3)Gal(β1-4)[Fuc(α1-3)]GlcNAcβ, Neu5Ac(α2-3)Gal(β1-3)GlcNAcβ, Neu5Ac(α2-3)Gal(β1-3)(6S)GlcNAcβ, Neu5Ac(α2-3)Gal(β1-3)GalNAcα, Neu5Ac(α2-3)Gal(β1-6)Manα, Neu5Ac(α2-3)Gal(β1-3)GalNAc(β1-3)Galα, Neu5Ac(α2-3)Gal(β1-3)GalNAc(β1-3)Galβ, Neu5Ac(α2-3)Gal(β1-3)GalNAc(β1-3)Gal(α1-4)Gal(β1-4)Glcβ, Neu5Ac(α2-3)Gal(β1-6)[Neu5Ac(α2-3)Gal(β1-3)]GlcNAcβ, Neu5Ac(α2-3)Gal(β1-6)[Neu5Ac(α2-3)Gal(β1-3)]Manα, Neu5Ac(α2-3)Gal(β1-3)[Neu5Ac(α1-6)]GlcNAcβNeu5Ac(α2-3)[GalNAc(β1-4)]Gal(β1-4)Glc(3, Gal(β1-3)GalNAc(β1-4)[Neu5Ac(α1-3)]Gal(β1-4)Glc(3, Fuc(α1-2)Gal(β1-3)GalNAc(β1-4) [Neu5Ac(α1-3)]Gal(β1-4)Glcβ, Neu5Ac(α2-3)Fuc(α1-4)Gal(β1-4)Glcβ, Neu5Acα, 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)GalNAcα, Neu5Ac(α2-6)Gal(β1-3)GlcNAcβ, Neu5Ac(α2-6)Gal(β1-4)(6S)GlcNAcβ, Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-3)Gal(β1-4)GlcNAcβ, and Neu5Ac(α2-6)Gal(β1-4)GlcNAc(β1-3)Gal(β1-4)GlcNAc(β1-3)Gal(β1-4)GlcNAcβ.