Biodetectors targeted to specific ligands
First Claim
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1. A biodetector for the detection of a selected substance, said biodetector comprising:
- a signal converting element, comprising (i) an extracellular ligand-specific binding domain which specifically binds said selected substance, wherein said ligand-specific binding domain comprises an epitope-binding fragment of an antibody, and (ii) an intracellular signal transforming domain of PhoQ, wherein binding of said substance to said epitope-binding fragment of said ligand-specific binding domain activates said intracellular signal transforming domain providing an activated intracellular signal transforming domain;
a transducer, wherein (i) said transducer has an inactive form and an active form which are distinct from each other, and (ii) said activated intracellular signal transforming domain converts said inactive form of said transducer into said active form of said transducer;
a transcription control element comprising the phoN promoter, wherein expression mediated by said transcription control element is activated by said active form of said transducer; and
a reporter gene operatively linked to said transcription control element, wherein expression of said reporter gene mediated by said transcription control element causes expression of a reporter gene product that provides a detectable signal, wherein said detectablen signal is detected optically by bioluminescence detection or fluorescence detection.
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Abstract
The present invention relates to biodetectors for detecting and quantifying molecules in liquid, gas, or matrices. More specifically, the present invention relates to biodetectors comprising a molecular switching mechanism to express a reporter gene upon interaction with target substances. The invention further relates to methods using such biodetectors for detecting and quantifying selected substances with high specificity and high sensitivity.
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Citations
18 Claims
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1. A biodetector for the detection of a selected substance, said biodetector comprising:
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a signal converting element, comprising (i) an extracellular ligand-specific binding domain which specifically binds said selected substance, wherein said ligand-specific binding domain comprises an epitope-binding fragment of an antibody, and (ii) an intracellular signal transforming domain of PhoQ, wherein binding of said substance to said epitope-binding fragment of said ligand-specific binding domain activates said intracellular signal transforming domain providing an activated intracellular signal transforming domain;
a transducer, wherein (i) said transducer has an inactive form and an active form which are distinct from each other, and (ii) said activated intracellular signal transforming domain converts said inactive form of said transducer into said active form of said transducer;
a transcription control element comprising the phoN promoter, wherein expression mediated by said transcription control element is activated by said active form of said transducer; and
a reporter gene operatively linked to said transcription control element, wherein expression of said reporter gene mediated by said transcription control element causes expression of a reporter gene product that provides a detectable signal, wherein said detectablen signal is detected optically by bioluminescence detection or fluorescence detection. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18)
at least about 1000 biodetectors of claim 8, wherein the extracellular ligand-specific binding domain of each of said biodetectors comprises a different antibody fragment, a reporter gene operatively linked to said transcription control element, wherein expression of said reporter gene mediated by said transcription control element causes expression of a reporter gene product that provides a detectable signal, wherein said detectable signal is detected optically by means selected from the group consisting of bioluminescence detection and fluorescence detection. -
14. An ordered array of the library of biodetectors of claim 13.
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15. The biodetector of claim 1, wherein said detectable signal is detectable by fluorescence detection.
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16. The biodetector of claim 15, wherein said reporter gene encodes a fluorescent protein.
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17. The biodetector of claim 8, wherein said bacterial cell comprises a luciferase operon.
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18. The biodetector of claim 1, wherein said bioluminescence detection or fluorescence detection is performed using a charge coupled device camera.
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Specification