METHODS AND ARRAYS FOR TARGET ANALYTE DETECTION AND DETERMINATION OF TARGET ANALYTE CONCENTRATION IN SOLUTION

• US 20130345078A1
• Filed: 04/25/2013
• Published: 12/26/2013
• Est. Priority Date: 02/21/2006
• Status: Abandoned Application

• Current Assignee: Trustees of Tufts College
• Original Assignee: Trustees of Tufts College
• Inventors: Walt, David R., Rissin, David M.
• Application Number:
  US13/870,596
• Publication Number:
  US 20130345078A1
• Time in Patent Office: Days
• US Class Current: 506/9
• CPC Class Codes: C12Q 1/6837 using probe arrays or probe chips C12Q1/6874 takes precedence
• CPC Class Codes: C12Q 2527/143 Concentration of primer or probe
• CPC Class Codes: G01N 2333/938 acting on beta-galactose-glycoside bonds, e.g. beta-galactosidase
• CPC Class Codes: G01N 33/543 with an insoluble carrier for immobilising immunochemicals
• CPC Class Codes: G01N 33/54346 Nanoparticles
• CPC Class Codes: G01N 33/54366 Apparatus specially adapted for solid-phase testing
• CPC Class Codes: G01N 33/54386 Analytical elements
• CPC Class Codes: G01N 33/581 with enzyme label (including co-enzymes, co-factors, enzyme inhibitors or substrates)

Abstract

Arrays of single molecules and methods of producing an array of single molecules are described. Arrays with defined volumes between 10 attoliters and 50 picoliters enable single molecule detection and quantitation.

Claims

1. A method of producing an array of single molecules, comprising:

a) providing a sample and an array, said sample comprising a molecules of target analyte in a concentration, said array comprising at least 1000 sites, each site having a defined volume between 10 attoliters and 50 picoliters; and

b) contacting said array with said sample such that the ratio of molecules of target analyte to each site of said array is less than 1;

1, so as to produce an array comprising sites containing single molecules.

2. The method of claim 1, wherein said target analyte comprises biomolecules.

3. The method of claim 2, wherein said biomolecule is selected from the group consisting of proteins, nucleic acids, lipids, and carbohydrates.

4. The method of claim 2, wherein said biomolecule is selected from the group consisting of hormones, cytokines and cellular antigens.

5. The method of claim 2, wherein said biomolecule is a receptor.

6. The method of claim 5, wherein said receptor is selected from the group consisting of neural receptors, hormonal receptors, and nutrient receptors.

7. The method of claim 2, wherein said biomolecules are labeled.

8. The method of claim 1, wherein said target analyte is an environmental pollutant.

9. The method of claim 1, wherein said target analyte is selected from the group consisting of pesticides, insecticides and toxins.

10. The method of claim 1, wherein said target analyte is a drug.

11. The method of claim 1, wherein said ratio is less than 1:

5.

12. The method of claim 1, wherein said ratio is 1:

10.

13. The method of claim 11, wherein said ratio is between 1:

5 and 1;

500.

14. The method of claim 1, wherein the percentage of sites containing a single molecule provide a measure of the concentration of molecules in said sample.

15. An array produced according to the method of claim 1.

16. The method of claim 1, further comprising, after step b), sealing the array such that the contents of each site cannot escape said site.

17. An array produced according to the method of claim 16.

18-25. -25. (canceled)

26. A method of detecting a target analyte in a sample, the method comprising:

a) providing a sample and an array, said sample comprising a biomolecule target analyte, said array comprising at least 1000 sites, each site having a defined volume between 10 attoliters and 50 picoliters;

b) contacting said array with said sample such that the ratio of biomolecule target analyte to each site of said array is less than 1;

1; and

c) determining the number of sites of said array which contain a single biomolecule.

27-44. -44. (canceled)

45. A method of detecting a target analyte in a sample, the method comprising:

a) providing a sample and an array, said sample comprising a biomolecule target analyte in a first concentration, said array comprising at least 10,000 sites, each site having a defined femtoliter volume;

b) diluting said sample to create a diluted sample, said diluted sample comprising a biomolecule target analyte in a second concentration;

c) contacting said array with said diluted sample such that the ratio of biomolecule target analyte to each site of said array is between 1;

5 and 1;

500; and

d) determining the number of sites of said array which contain a single biomolecule.

46-61. -61. (canceled)