Method and device for detecting and quantifying biomolecules

• US 20030064390A1
• Filed: 09/09/2002
• Published: 04/03/2003
• Est. Priority Date: 12/13/1999
• Status: Abandoned Application

• Current Assignee: November Aktiengesellschaft Gesellschaft Fur Molekulare Medizin
• Original Assignee: November Aktiengesellschaft Gesellschaft Fur Molekulare Medizin
• Inventors: Schülein, Jurgen, Hassmann, J?ouml;rg
• Application Number:
  US10/149,873
• Publication Number:
  US 20030064390A1
• Time in Patent Office: Days
• US Class Current: 435/6
• CPC Class Codes: C12Q 1/6825 Nucleic acid detection involving sensors
• CPC Class Codes: C12Q 2563/116 electrical properties of nucleic acids, e.g. impedance, conductivity or resistance
• CPC Class Codes: G01N 33/5438 Electrodes

Abstract

The invention relates to a method for detecting and quantifying a first biomolecule in solution (5, 8), comprising the following steps: a) binding of the first biomolecule (5, 8) to a second biomolecule (3, 7) which at least along segments thereof exhibits a specific affinity to a first biomolecule and b) determination of the electrical conductivity of the complex formed from the first (3, 7) and the second biomolecule (5, 8), whereby the second biomolecule (3, 7) forms a bridge between a first 2a and a second electrode 2b.

Claims

1. A method of detecting and/or quantifying a first biomolecule (5, 8) present in a solution, having the following steps:

a) binding of the first molecule (5, 8) to a second biomolecule (3, 7) which, at least in sections, possesses specific affinity to the first biomolecule (5, 8) and

b) measuring of the electrical conductivity of the complex formed from the first (3, 7) and second biomolecules (5, 8), the second biomolecule (3, 7) forming a bridge between a first electrode (2a) and a second electrode (2b).

2. The method of claim 1, wherein the second biomolecule (5, 8) is bound before step a at least by one end (E1) to one of the electrodes (2a, 2b).

3. The method of one of the preceding claims, wherein the binding of at least one end (El) of the second biomolecule (5, 8) to one electrode (2a, 2b) is mediated by way of a spacer molecule (S) and/or a linker molecule.

4. The method of one of the preceding claims, wherein before or after step a the second biomolecule (5, 8) is bound by the other end (E2) to the second electrode (2b).

5. The method of claim 3, wherein the binding of the other end (E2) is assisted by application of a potential.

6. The method of one of the preceding claims, wherein a charge carrier (6) is bound to the other end (E2) of the second biomolecule (3, 7).

7. The method of claim 6, wherein the charge carrier (6) is a metal cluster, an organic molecule or a complexing agent which the binding of the other end (E2) to the second electrode (2b) is mediated by way of the charge carrier (6).

8. The method of one of the preceding claims, wherein step a takes place between the binding of one end (E1) of the second biomolecule (3, 7) to the first electrode (2a) and the binding of the other end (E2) of the second biomolecule (3, 7) to the second electrode (2b).

9. The method of one of the preceding claims, wherein the first electrode (2a) and the second electrode (2b) are applied to an electrically insulating substrate (1).

10. The method of one of the preceding claims, wherein prior to step b the substrate (1) is washed and/or dried and/or evacuated.

11. The method of one of the preceding claims, wherein the electrical conductivity of the complex bound between the first electrode (2a) and the second electrode (2b) is measured.

12. The method of one of the preceding claims, wherein instead of the conductivity the capacitance of the complex bound between the first electrode (2a) and the second electrode (2b) is measured.

13. The method of one of the preceding claims, wherein instead of the conductivity the impedance of the construct bound between the first electrode (2a) and the second electrode (2b) is measured.

14. The method of one of the preceding claims, wherein the distance between the first electrode (2a) and the second electrode (2b) is from 3 nm to 1 μ

m, preferably 50 nm.

15. The method of one of preceding claims, wherein the first biomolecule (3, 7) is a single-stranded DNA or RNA which is complementary to the second biomolecule (5, 8).

16. The method of one of the preceding claims, wherein the second biomolecule (5, 8) is formed with sections of double-strandedness, the double-stranded section(s) being formed preferably from DNA and/or RNA.

17. The method of one of the preceding claims, wherein a single-stranded section has been inserted into the second biomolecule (5, 8).

18. The method of one of the preceding claims, wherein the single-stranded section is formed from DNA, RNA or PNA.

19. The method of one of the preceding claims, wherein a protein or a peptide is associated with into the second biomolecule (5, 8).

20. The method of one of the preceding claims, wherein following step b a force is exerted on the first biomolecule (5, 8) by means of an applied voltage in order to remove the first biomolecule (5, 8) from the second biomolecule (3, 7).

21. The method of one of the preceding claims, wherein following step b the substrate (1) is washed in order to remove the first biomolecule (5, 8) from the second biomolecule (3, 7).

22. A device for detecting and/or quantifying a first biomolecule (5, 8) present in a solution, wherein

aa) a first electrode (2a) and a second electrode (2b) are applied on an electrically insulating substrate (1),

bb) bound at least to the first electrode (2a) by its one end (E1) is a second biomolecule (3, 7) which, at least in sections, possesses a specific affinity to the first biomolecule (5, 8), and wherein

cc) the distance between the first electrode (2a) and the second electrode (2b) is chosen such that by binding the other end (E2) of the second biomolecule (3, 7) it is possible to produce a bridge between the first electrode (2a) and the second electrode (2b).

23. The device of claim 22, wherein the distance between the first electrode (2a) and the second electrode (2b) is from 3 nm to 1 μ

m, preferably 50 nm.

24. The device of claim 22 or 23, wherein at least one end (E1) of the second biomolecule (5, 8) is bound to one electrode (2a, 2b) by way of a direct coupling, a spacer molecule (S) and/or a linker molecule (L).

25. The device of one of claims 22 to 24, wherein a charge carrier (6) has been bound to the other end (E2) of the second biomolecule (3, 7).

26. The device of one of claims 22 to 25, wherein the charge carrier (6) is a metal cluster, an organic molecule or a completing agent and the binding of the other end (E2) to the second electrode (2b) can be mediated by way of the charge carrier (6).

27. The device of one of claims 22 to 26, wherein the substrate (1) is produced from ceramic, from silicon compounds, preferably silicon with an oxide layer, from mica or from an electrically insulating polymer matrix.

28. The device of one of claims 22 to 27, wherein a means for measuring the electrical conductivity of the construct formed from the first biomolecule (3, 7) and the second biomolecule (5, 8), said means being connected to the first electrode (2a) and the second electrode (2b), is provided.

29. The device of claim 28, wherein instead of the conductivity the capacitance of the construct formed from the first biomolecule (5, 8) and the second biomolecule (3, 7) can be measured by means of said means.

30. The device of claim 28, wherein instead of the conductivity the impedance of the construct formed from the first biomolecule (5, 8) and the second biomolecule (3, 7) can be measured by means of said means.

31. The device of one of claims 22 to 30, wherein the first biomolecule (5, 8) is a single-stranded DNA or RNA which is complementary to the second biomolecule (3, 7).

32. The device of claim 31, wherein the second biomolecule (3, 7) is formed with sections of double-strandedness, the double-stranded sections being formed preferably from DNA and/or RNA.

33. The device of claim 32, wherein a single-stranded section has been inserted into the second biomolecule (3, 7).

34. The device of one of claims 22 to 33, wherein the single-stranded section is formed from DNA, RNA or PNA.

35. The device of one of claims 22 to 34, wherein a protein or a peptide is inserted into the second biomolecule (3, 7).

36. The device of one of claims 22 to 35, wherein a large number of first electrodes (2a) and second electrodes (2b) are mounted on the substrate (1).

37. The device of one of claims 22 to 36, wherein the second biomolecules (3, 7) bound to the first electrodes (2a) are different from one another, so that simultaneous detection and/or quantification of a large number of first biomolecules (5, 8) is possible.