Differential labelling method

• US 20030060647A1
• Filed: 05/28/2002
• Published: 03/27/2003
• Est. Priority Date: 05/28/2001
• Status: Active Grant

• Current Assignee: Leica Biosystems Newcastle Ltd. (Danaher Corporation)
• Original Assignee: Kreatech Biotechnology BV (Danaher Corporation)
• Inventors: Talman, Eduard Gerhard, Heetebrij, Robert Jochem, Veuskens, Jacky Theo Maria, van Gijlswijk, Robertus Petrus Maria
• Granted Patent: US 7,704,755 B2
• Time in Patent Office: Days
• US Class Current: 556/137
• CPC Class Codes: G01N 33/58 involving labelled substances G01N33/53 takes precedence
• CPC Class Codes: G01N 33/84 involving inorganic compounds or pH

Abstract

The invention relates to a method for differentially labelling one or more entities, together comprising distinct reactive sites. The invention further relates to an entity that has been labelled by a method according to the invention and to a diagnostic kit comprising a labelled entity and to a diagnostic kit to employ a method according to the invention.

Claims

1. A method for differentially forming a complex of a platinum compound and one or more entities, said entities together comprising one or more sulphur-containing reactive sites and one or more nitrogen-containing reactive sites, wherein said platinum compound is reacted with said one or more entities such that substantially only sulphur containing reactive sites or substantially only nitrogen containing reactive sites are linked to said platinum compound.

2. A method according to claim 1, wherein pH is used to discriminate between reactions of sulphur containing reactive sites and reactions of nitrogen containing reactive sites.

3. A method according to claim 2, wherein the method is carried out at a pH of 5 or less, whereby substantially only sulphur containing reactive sites are linked to said platinum compound.

4. A method according to claim 1, wherein the ionic strength is used to discriminate between reactions of sulphur containing reactive sites and reactions of nitrogen containing reactive sites.

5. A method according to claim 4, wherein the ionic strength is at least 0.1 mol/l.

6. A method according to claim 5, wherein said ionic strength is realised by one or more salts comprising Cl⁻

, NO₃⁻

, HCO₃⁻

, CO₃²⁻

, SO³⁻

, I⁻

, Br⁻

, acetate, carboxylate, phosphate, sulphate, oxalate, citrate, a phosphonate or a mixture thereof.

7. A method according to claim 6, wherein said ionic strength is realised by one or more salts comprising Cl⁻

, NO₃⁻

, I⁻

, Br⁻

, acetate, or sulphate.

8. A method according to claim 1, wherein the stereochemical structure of the platinum compound is used to discriminate between reactions of sulphur containing reactive sites and reactions of nitrogen containing reactive sites.

9. A method according to claim 1, wherein said sulphur containing site or sites are first differentially shielded by a shielding moiety and thereafter said platinum compound is differentially linked to the nitrogen containing reactive site.

10. A method according to claim 1, wherein said nitrogen containing site or sites are first differentially shielded by a shielding moiety and thereafter said platinum compound is differentially linked to the sulphur containing reactive site.

11. A method according to claim 9, wherein said shielding moiety is a trans-platinum compound.

12. A method according to claim 10, wherein said shielding moiety is a trans-platinum compound.

13. A method according to any of claims 9, 10, 11, or 12, wherein the shielding moiety is selectively removed from the shielded reactive site, after the platinum compound has been reacted such that said platinum is differentially linked to said entity.

14. A method according to claim 1, wherein one or more transition metal ions are used in a concentration chosen such that said platinum compound is differentially linked to a nitrogen containing reactive site.

15. A method according to claim 14, wherein said one or more transition metal ions are selected from the group consisting of Cu(II) and Zn(II).

16. A method according to claim 1, wherein at least one of the nitrogen containing reactive sites is a histidine residue, and wherein said platinum compound is reacted with said one or more entities such that substantially only histidine residues or only non-histidine reactive sites are linked to said platinum compound.

17. A method according to claim 1, wherein said platinum compound is represented by one of the following formulas:

18. A method according to claim 1, wherein the reactive site of the entity is connected to the platinum moiety of the platinum compound through a spacer.

19. A method according to claim 1, wherein said entity comprises a primary amine, a secondary amine, a tertiary amine, an aromatic amine, a thiol, a thioether, a sulfide, a disulfide, a thioamide, a thion, an amide, an imide, an imine, an iminoether, or an azide.

20. A method according to claim 1, wherein one or more entities are chosen from the group of amino acids, peptides, oligopeptide, polypeptides, proteins, immunoglobulins, enzyms, synzyms, phospholipides, glycoproteins, nucleic acids, nucleosides, nucleotides, oligonucleotides, polynucleotides, peptide nucleic acids, peptide nucleic acid oligomers, peptide nucleic acid polymers, amines, aminoglycosides.

21. A method according to claim 20, wherein one or more entities are chosen from the group of immunoglobulins, enzymes, antibody-antigen complexes.

22. A method according to claim 1, wherein the platinum compound is further attached to a marker.

32. A method according to claim 1, wherein the marker is connected to the platinum compound through a spacer.

23. A method for labelling one or more entities through a platinum-linker, said entities together comprising one or more histidine residues and one or more other nitrogen containing reactive sites, and wherein said platinum compound is reacted with said one or more entities such that substantially only histidine residues or only non-histidine reactive sites are linked to said platinum compound.

24. An entity differentially linked to a platinum compound at one or more nitrogen containing reactive sites and/or one or more sulphur containing reactive.

25. An entity according to claim 24 further comprising a marker attached to said platinum compound.

26. An entity labelled with different markers at, respectively, nitrogen containing reactive sites and sulphur containing reactive sites.

27. A diagnostic kit comprising entities differentially linked at one or more nitrogen containing reactive sites and/or one or more sulphur containing reactive sites to a platinum compound and a marker.

28. A diagnostic kit comprising one or more preparations selected from the group consisting of platinum linkers and at least one preparation selected from the group consisting of buffers, transition metal ion preparations, preparations for adjusting the ionic strength and preparations comprising a shielding moiety.

29. A diagnostic kit according to claim 28 further comprising a marker.

30. A method for differentially labelling one or more entities through a platinum-linker, said entities together comprising one or more sulphur containing reactive sites and one or more nitrogen containing reactive sites, wherein a complex of a platinum compound and a marker is formed, and wherein said platinum compound is reacted with said one or more entities such that substantially only sulphur containing reactive sites or substantially only nitrogen containing reactive sites are linked to said platinum compound.

31. A method according to claim 30, wherein said nitrogen containing reacting sites and said sulphur containing reactive sites are labelled with different markers.

33. A method according to claim 30, wherein said marker is a fluorochrome, a phosphorescent marker, chemiluminescent marker, a specific binding pair component, a UV or visual light absorbing marker ,a radioactive marker, a colloidal dye substance, a reducing substance, a particulate sol or a metal.

34. A method according to claim 33, wherein said marker is dinitrophenol (DNP) tetramethylrhodamine, digoxygenin, a cyanine-colorant, biotin, avidin or streptavidin.

35. A method according to claim 30, wherein the labelled entity is subsequently analysed using a liquid based analyte analysis system.