Agent and method for detecting chemical substances

• US 6,468,752 B1
• Filed: 11/13/2000
• Issued: 10/22/2002
• Est. Priority Date: 03/03/1998
• Status: Expired due to Fees

• Current Assignee: November Aktiengesellschaft Gesellschaft Fur Molekulare Medizin
• Original Assignee: November Aktiengesellschaft Gesellschaft Fur Molekulare Medizin
• Inventors: Bertling, Wolf
• Primary Examiner(s): Jones, W. Gary
• Assistant Examiner(s): Taylor, Janell E.
• Application Number:
  US09/623,581
• Time in Patent Office: 708 Days
• Field of Search: 435/6, 435/91.1, 435/91.2, 435/5, 536/24.3, 935/77, 935/78
• US Class Current: 435/6.11
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: G01N 33/54306 Solid-phase reaction mechanisms

Abstract

The invention relates to an agent for detecting chemical substances, comprising: a first polynucleotide or peptide sequence having a first fluorophore molecule, the first end of said sequence being bonded on a solid phase and a second polynucleotide or peptide sequence having a second fluorophore molecule, the second end of said sequence having a group that is sensitive to the chemical substance to be detected. The first polynucleotide or peptide sequence can be hybridized to the second polynucleotide or peptide sequence in such a way that a spatial relationship enabling an interaction betweeen the first and the second fluorophore molecule is produced. When the chemical substance including a physical property is attached to the group and the influence of an external force is exerted on the physical property, the spatial relation can be eliminated, thereby generating or abolishing a fluorescent reaction.

Claims

1. A method for the detection of chemical substances, comprising:

providing an agent comprising a first polynucleotide or peptide sequence having a first fluorophoric molecule, wherein first end of said first polynucleotide or peptide sequence is bonded to a solid phase, and a second polynucleotide or peptide sequence having a second fluorophoric molecule, wherein a second end of said second polynucleotide or peptide sequence has a group which is sensitive to the chemical substance to be detected, said chemical substance comprising a physical property sensitive to an external force, the first polynucleotide or peptide sequence being hybridizable to the second polynucleotide or peptide sequence such that a spatial relationship making possible an interaction between the first and the second fluorophoric molecule is producible, and the spatial relationship being terminatable and thus a fluorescence reaction being producible upon addition of the chemical substance to the group and action of an external force;

contacting the agent with the chemical substance to be detected;

applying the external force; and

detecting the presence or absence of the fluorescence reaction.

2. The method according to claim 1, the first and the second polynucleotide or peptide sequence being linked to give a molecule.

3. The method according to claim 1, the spatial relationship being designed in the form of a secondary structure.

4. The method according to claim 3, the secondary structure being a hairpin loop, a helix or a pleated sheet.

5. The method according to claim 4, the first fluorophoric molecule being bonded to a first loop section and the second fluorophoric molecule being bonded oppositely to a second loop section of the hairpin loop at a distance making possible an interaction.

6. The method according to claim 1, the solid phase being plastic.

7. The method according to claim 6, the plastic comprising polycarbonate, trimethylthiophene, thiophene, triaminobenzene or polycarbene.

8. The method according to claim 6, the plastic being electrically conductive.

9. The method according to claim 1, the first fluorophoric molecule being an acceptor molecule and the second fluorophoric molecule being a donor molecule.

10. The method according to claim 9, the acceptor molecule being 6-carboxytetramethylrhodamine, tetramethylrhodamine, fluorescein, DABCYL or Bodipy F1.

11. The method according to claim 9, the donor molecule being 6-carboxyfluorescene, fluorescein, IADEANS, EDANS or Bodipy F1.

12. The method according to claim 1, the first fluorophoric molecule being replaced by a quencher.

13. The method according to claim 1, the polynucleotide sequence being DNA, PTO or PNA.

14. The method according to claim 1, the group being an antibody, a receptor, an antigen, or a ligand.

15. The method according to claim 14, wherein a substance which contains a recognizing group suitable for recognizing the chemical substance and having a superparamagnetic particle bonded thereto is added to the sample containing the chemical substance.

16. The method according to claim 15, the chemical substance to be detected being bonded to the group which is sensitive to the chemical substance and to the recognizing group.

17. The method according to claim 16, a magnetic field being applied such that the superparamagnetic particle moves away from the solid phase and the secondary structure is abolished or altered thereby.

18. The method according to claim 1, the fluorescence being detected by means of a fluorometer connected to a data processing device.

19. The method according to claim 18, the concentration of the chemical substance to be detected being determined from the temporal alteration of the fluorescence intensity.

20. An agent for the detection of chemical substances, having a first polynucleotide or peptide sequence having a first fluorophoric molecule, wherein a first end of said first polynucleotide or peptide sequence is bonded to a solid phase, and a second polynucleotide or peptide sequence having a second fluorophoric molecule, wherein a second end of said second polynucleotide or peptide sequence has a group which is sensitive to the chemical substance to be detected, said chemical substance comprising a physical property sensitive to an external force, the first polynucleotide or peptide sequence being hybridizable to the second polynucleotide or peptide sequence such that a spatial relationship making possible an interaction between the first and the second fluorophoric molecule is producible, and the spatial relationship being terminatable and thus a fluorescence reaction being producible in the presence of the chemical substance and action of said external force, wherein the spatial relationship is designed in the form of a secondary structure, wherein the secondary structure is a hairpin loop, a helix or a pleated sheet.

21. An agent for the detection of chemical substances, having a first polynucleotide or peptide sequence having a first fluorophoric molecule, wherein a first end of said first polynucleotide or peptide sequence is bonded to a solid phase, and a second polynucleotide or peptide sequence having a second fluorophoric molecule, wherein a second end of said second polynucleotide or peptide sequence has a group which is sensitive to the chemical substance to be detected, said chemical substance comprising a physical property sensitive to an external force, the first polynucleotide or peptide sequence being hybridizable to the second polynucleotide or peptide sequence such that a spatial relationship making possible an interaction between the first and the second fluorophoric molecule is producible, and the spatial relationship being terminatable and thus a fluorescence reaction being producible in the presence of the chemical substance and action of said external force, wherein the solid phase is plastic, wherein the plastic is electrically conductive.