CA2198470A1 - Luciferase labelling method - Google Patents
Luciferase labelling methodInfo
- Publication number
- CA2198470A1 CA2198470A1 CA002198470A CA2198470A CA2198470A1 CA 2198470 A1 CA2198470 A1 CA 2198470A1 CA 002198470 A CA002198470 A CA 002198470A CA 2198470 A CA2198470 A CA 2198470A CA 2198470 A1 CA2198470 A1 CA 2198470A1
- Authority
- CA
- Canada
- Prior art keywords
- luciferase
- antibody
- adenosine triphosphate
- chemical entity
- luciferin
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Abstract
A method is provided for conjugating luciferase to a chemical entity, particularly to a specific binding agent such as m antibody, antigen or a nucleic acid, and more particularly an antibody, comprising (a) mixing the luciferase with one or more of D-luciferin, magnesium ions and adenosine triphosphate and (b) performing a covalent coupling reaction between the luciferase and the binding reagent using a covalent coupling reagent wherein the amount of D-luciferin, magnesium ions and/or adenosine triphosphate is sufficient to protect the luciferase activity against inhibition by the covalent coupling reagent.
Preferably the step (a) is carried out by mixing the luciferase with its substrates in solution and preferably both magnesium and adenosine triphosphate are present as magnesium adenosine triphosphate (Mg2+ATP), optionally together with D-luciferin. In a second aspect of the invention there is provided a labelled chemical entity comprising a chemical entity conjugated to active luciferase as provided by the method of the present invention. Preferably the chemical entity is a specific binding agent suitable for use in a specific binding assay, preferably being an antibody, antigen or nucleic acid. When the binding agent is a nucleic acid, it is preferably an oligonucleotide, but may be a polynucleotide or a nucleoside, and may be used as a hybridisation probe or a chain extension primer e.g. a PCR primer. Most advantageously the entity is an antibody as previous attempts to couple antibodies to luciferase have resulted in inactivity. Test kits are further provided.
Preferably the step (a) is carried out by mixing the luciferase with its substrates in solution and preferably both magnesium and adenosine triphosphate are present as magnesium adenosine triphosphate (Mg2+ATP), optionally together with D-luciferin. In a second aspect of the invention there is provided a labelled chemical entity comprising a chemical entity conjugated to active luciferase as provided by the method of the present invention. Preferably the chemical entity is a specific binding agent suitable for use in a specific binding assay, preferably being an antibody, antigen or nucleic acid. When the binding agent is a nucleic acid, it is preferably an oligonucleotide, but may be a polynucleotide or a nucleoside, and may be used as a hybridisation probe or a chain extension primer e.g. a PCR primer. Most advantageously the entity is an antibody as previous attempts to couple antibodies to luciferase have resulted in inactivity. Test kits are further provided.
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| GB9417593A GB9417593D0 (en) | 1994-09-01 | 1994-09-01 | Luciferase labelling method |
| GB9417593.2 | 1994-09-01 | ||
| PCT/GB1995/002038 WO1996007100A2 (en) | 1994-09-01 | 1995-08-30 | Luciferase labelling method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CA2198470A1 true CA2198470A1 (en) | 1996-03-07 |
| CA2198470C CA2198470C (en) | 2006-02-14 |
Family
ID=35892302
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA002198470A Expired - Fee Related CA2198470C (en) | 1994-09-01 | 1995-08-30 | Luciferase labelling method |
Country Status (1)
| Country | Link |
|---|---|
| CA (1) | CA2198470C (en) |
-
1995
- 1995-08-30 CA CA002198470A patent/CA2198470C/en not_active Expired - Fee Related
Also Published As
| Publication number | Publication date |
|---|---|
| CA2198470C (en) | 2006-02-14 |
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Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| EEER | Examination request | ||
| MKLA | Lapsed |