Methods for assaying analytes

• US 20070048747A1
• Filed: 09/01/2005
• Published: 03/01/2007
• Est. Priority Date: 09/01/2005
• Status: Abandoned Application

• Current Assignee: Corning Incorporated
• Original Assignee: Corning Incorporated
• Inventors: Peng, Jinlin, Li, Guangshan, Lynn, Jeffrey, Leslie, Thomas
• Application Number:
  US11/219,049
• Publication Number:
  US 20070048747A1
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/54393 Improving reaction conditions or stability, e.g. by coating or irradiation of surface, by reduction of non-specific binding, by promotion of specific binding

Abstract

Described herein are methods for assaying analytes.

Claims

1. A method for performing an assay of a sample comprising an analyte, the method comprising:

a. attaching a biomolecule to a support, wherein the support comprises a substrate, a first tie layer attached to the substrate, a first polymer attached to the first tie layer, wherein the biomolecule attaches to the first polymer;

b. exposing the support with the attached biomolecule to a relative humidity of greater than 40%;

c. attaching a blocking agent to the first polymer;

d. contacting support with the sample comprising the analyte, wherein after the contacting step, the analyte is bound to the biomolecule; and

e. detecting the bound analyte.

2. The method of claim 1, wherein the assay is a high-throughput assay.

3. The method of claim 1, wherein the analyte comprises a drug, an oligonucleotide, a nucleic acid, a protein, a peptide, an antibody, an antigen, a hapten, saccharide, lipid, a small molecule, or a mixture thereof.

4. The method of claim 1, wherein the bound analyte is detected by fluorescence or non-fluorescence.

5. The method of claim 1, wherein the bound analyte is detected by surface plasmon resonance, a waveguide resonant grating system, or mass spectrometry.

6. The method of claim 1, wherein the substrate comprises a plastic, a polymeric or co-polymeric substance, a ceramic, a glass, a metal, a crystalline material, a noble or semi-noble metal, a metallic or non-metallic oxide, a transition metal, or any combination thereof.

7. The method of claim 1, wherein the substrate comprises an amine modified glass surface.

8. The method of claim 1, wherein the substrate is a microplate or a slide.

9. The method of claim 1, wherein the first tie layer is covalently attached to the outer surface of the substrate.

10. The method of claim 1, wherein the first tie layer is electrostatically attached to the outer surface of the substrate.

11. The method of claim 1, wherein the first tie layer compound comprises a straight or branched-chain aminosilane, aminoalkoxysilane, aminoalkylsilane, aminoarylsilane, aminoaryloxysilane, or a derivative or salt thereof.

12. The method of claim 1, wherein the first tie layer compound comprises 3-aminopropyl trimethoxysilane, N-(beta-aminoethyl)-3-aminopropyl trimethoxysilane, N-(beta-aminoethyl)-3-aminopropyl triethoxysilane, N′

-(beta-aminoethyl)-3-aminopropyl methoxysilane, or aminopropylsilsesquixoane.

13. The method of claim 1, wherein the first polymer is covalently attached to the first tie layer.

14. The method of claim 1, wherein the first polymer is electrostatically attached to the first tie layer.

15. The method of claim 1, wherein the first polymer comprises a copolymer.

16. The method of claim 15, wherein the copolymer is derived from maleic anhydride and a first monomer.

17. The method of claim 16, wherein the first monomer comprises styrene, tetradecene, octadecene, methyl vinyl ether, triethylene glycol methyl vinyl ether, butylvinyl ether, divinylbenzene, ethylene, acrylamide, pyrolidone, dimethylacrylamide, a polymerizable oligo(ethylene glycol) or oligo(ethylene oxide), or a combination thereof.

18. The method of claim 1, wherein the first polymer comprises poly(vinyl acetate-maleic anhydride), poly(ethylene-alt-maleic anhydride), poly(isobutylene-alt-maleic anhydride), poly(maleic anhydride-alt-1-octadecene), poly(maleic anhydride-alt-1-tetradecene), poly(maleic anhydride-alt-methyl vinyl ether), poly(triethyleneglycol methyvinyl ether-co-maleic anhydride), or any combination thereof.

19. The method of claim 1, wherein the first polymer comprises poly(styrene-co-maleic anhydride).

20. The method of claim 1, wherein the biomolecule comprises a natural, synthetic or modified oligonucleotide, a natural or modified nucleotide or nucleoside, a nucleic acid (DNA) or (RNA) or fragment thereof, a peptide comprising natural or modified amino acid, an antibody, a hapten, a biological ligand, a chelate, an aptamer, a lipid, a saccharide, a small molecule, a lectin, a modified polysaccharide, a synthetic composite macromolecule, a functionalized nanostructure, a synthetic polymer, a fluorophore, a chromophore, or a cell.

21. The method of claim 1, wherein the biomolecule comprises an oligonucleotide.

22. The method of claim 1, wherein the biomolecule comprises DNA or a fragment thereof.

23. The method of claim 1, wherein the biomolecule comprises RNA or a fragment thereof.

24. The method of claim 1, wherein the biomolecule comprises a protein, a peptide, or fragments thereof.

25. The method of claim 1, wherein the biomolecule is covalently attached to the first polymer.

26. The method of claim 1, wherein the biomolecule is electrostatically attached to the first polymer.

27. The method of claim 1, wherein the biomolecule is attached to the first polymer under a relative humidity of from about 40% to about 90%.

28. The method of claim 1, wherein the relative humidity is from about 55% to about 80%.

29. The method of claim 1, wherein the relative humidity is from about 60% to about 75%.

30. The method of claim 1, wherein during step (b), the substrate is exposed to a relative humidity of from 50% to 85% from 2 hours to 24 hours.

31. The method of claim 1, wherein during step (b), the substrate is exposed to a relative humidity of from 65% to 80% of from 8 hours to 16 hours.

32. The method of claim 1, wherein prior to step (a), the biomolecule is in a solution comprising a low boiling aprotic solvent, a surfactant, and a buffer, wherein the pH of the solution is from 7 to 10.

33. The method of claim 1, wherein a plurality of biomolecules are present on the support and wherein the biomolecules are on discrete and defined locations on the support to produce an array.

34. The method of claim 33, wherein the array comprises at least 96 distinct and defined locations.

35. The method of claim 33, wherein the support comprises at least 192 distinct and defined locations.

36. The method of claim 33, wherein the distinct and defined locations are from 50 to 1000 mm apart from each other.

37. The method of claim 1, wherein the blocking agent comprises at least one nucleophilic group, the first polymer comprises at least one electrophilic group, and the blocking agent is attached to the first polymer by a reaction between the electrophilic group and the nucleophilic group.

38. The method of claim 1, wherein the blocking agent comprises an amine group or a salt thereof.

39. The method of claim 1, wherein the blocking agent comprises 2-(2-aminoethoxy)ethanol, N,N-dimethyl ethylenediamine, ethanolamine, ethylenediamine, 4,7,10-trioxa-1,13-tridecanediamine, PEG amine, Tris hydrochloride, diethylaminoethyl-cellulose, diethylaminoethyl-cellulose amine, diethylaminoethyl-cellulose dextran, bovine serum albumin, chicken egg albumin, dry milk, pluronic or any combination thereof.

40. The method of claim 1, wherein the blocking agent comprises 2-(2-aminoethoxy)ethanol.

41. The method of claim 1, wherein the blocking agent comprises N,N-dimethyl ethylenediamine.

42. The method of claim 1, wherein the blocking agent comprises 2-(2-aminoethoxy)ethanol or N,N-dimethyl ethylenediamine in a solution of pH from 7 to 10.

43. The method of claim 1, wherein the blocking agent is covalently attached to the first polymer.

44. The method of claim 1, wherein the blocking agent is electrostatically attached to the first polymer.

45. The method of claim 1, wherein the sample of the analyte further comprises an enhancer.

46. The method of claim 45, wherein the enhancer comprises a low molecular weight poly(vinylpolypyrrolidone) polymer, low molecular weight dextran sulfate, Ficoll, low molecular weight polyethylene glycol, or any combination thereof.

47. The method of claim 45, wherein enhancer comprises poly(vinylpolypyrrolidone) having a molecular weight of about 10 kDa (PVP10), about 29 kDa (PVP29), about 50 kDa (PVP50), or about 55 kDa (PVP55), polyethyleneglycol having a molecular weight of about 8 kDa, or dextran sulfate having a molecular weight of about 8 kDa.

48. The method of claim 45, wherein the enhancer is present in the sample at from about 0.1% to about 30% by volume.

49. The method of claim 45, wherein the enhancer is present in the sample at from about 3% to about 4% by volume.

50. The method of claim 45, wherein the enhancer is present in the sample at from about 5% to about 10% by volume.

51. The method of claim 45, wherein the sample has a salt concentration of from 0.5×

SSC to 20×

SSC.

52. The method of claim 1, wherein the substrate is glass, the tie layer derived from gamma-aminopropylsilane, the first polymer is poly(styrene-co-maleic anhydride), the biomolecule is an oligonucleotide, wherein the relative humidity is from about 60% to about 75%, and the blocking agent is 2-(2-aminoethoxy)ethanol or N,N-dimethyl ethylenediamine.

53. A method for performing an assay of a sample comprising an analyte, the method comprising:

a. attaching a biomolecule to a support, wherein the support comprises a substrate comprising a first polymer, wherein the biomolecule attaches to the first polymer;

b. exposing the support with the attached biomolecule to a relative humidity of greater than 40%;

c. attaching a blocking agent to the first polymer;

d. contacting support with the sample comprising the analyte, wherein after the contacting step, the analyte is bound to the biomolecule; and

e. detecting the bound analyte.

54. The method of claim 53, wherein the substrate is derived from maleic anhydride and a first monomer.

55. The method of claim 54, wherein the first monomer comprises styrene, tetradecene, octadecene, methyl vinyl ether, triethylene glycol methyl vinyl ether, butylvinyl ether, divinylbenzene, ethylene, acrylamide, pyrolidone, dimethylacrylamide, a polymerizable oligo(ethylene glycol) or oligo(ethylene oxide), or a combination thereof.

56. The method of claim 53, wherein the first polymer comprises poly(vinyl acetate-maleic anhydride), poly(ethylene-alt-maleic anhydride), poly(isobutylene-alt-maleic anhydride), poly(maleic anhydride-alt-1-octadecene), poly(maleic anhydride-alt-1-tetradecene), poly(maleic anhydride-alt-methyl vinyl ether), poly(triethyleneglycol methyvinyl ether-co-maleic anhydride), or any combination thereof.

57. The method of claim 53, wherein the first polymer comprises poly(styrene-co-maleic anhydride).