Method of identifying energy transfer sensors for analytes
First Claim
1. A method of identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer, said method comprising:
- a) obtaining a predetermined analyte binding ligand from a combinatorial library comprising ligands, said analyte binding ligand having been predetermined by contacting the combinatorial library with a first analyte-analogue and selecting a ligand to which the first analyte-analogue binds; and
b) attaching a label to at least one of said analyte binding ligand and a second analyte-analogue, said label comprising at least one of a first component and a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair such that non-radiative fluorescence resonance energy transfer occurs when said second analyte-analogue is bound to said analyte binding ligand, and a change in non-radiative fluorescence resonance energy transfer occurs when said second analyte-analogue is not bound to said analyte binding ligand.
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Abstract
A method of identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer (FRET) using a combinatorial library. The method includes a) obtaining an analyte binding ligand from a combinatorial library that includes ligands, and b) attaching a label at least one of the analyte binding ligand and an analyte-analogue with at least one of a first component and a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair (FRET pair) such that FRET occurs when the analyte-analogue is bound to the analyte binding ligand, and a change in FRET occurs when the analyte-analogue is not bound to the analyte binding ligand.
The method also includes contacting a combinatorial library of ligands, which are optionally labeled with a component of a FRET pair, with analyte-analogue, which is optionally labeled with a component of a FRET pair, and detecting the presence of FRET.
90 Citations
55 Claims
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1. A method of identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer, said method comprising:
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a) obtaining a predetermined analyte binding ligand from a combinatorial library comprising ligands, said analyte binding ligand having been predetermined by contacting the combinatorial library with a first analyte-analogue and selecting a ligand to which the first analyte-analogue binds; and
b) attaching a label to at least one of said analyte binding ligand and a second analyte-analogue, said label comprising at least one of a first component and a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair such that non-radiative fluorescence resonance energy transfer occurs when said second analyte-analogue is bound to said analyte binding ligand, and a change in non-radiative fluorescence resonance energy transfer occurs when said second analyte-analogue is not bound to said analyte binding ligand. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12)
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13. A method of identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer, said method comprising:
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a) contacting a combinatorial library with an analyte-analogue, said combinatorial library comprising ligands;
b) identifying at least one ligand to which said analyte-analogue binds, said ligand being the analyte binding ligand; and
c) attaching a label to at least one of said analyte binding ligand and said analyte-analogue, said label comprising at least one of a first component and a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair such that non-radiative fluorescence resonance energy transfer occurs when said analyte-analogue is bound to said analyte binding ligand, and a change in non-radiative fluorescence resonance energy transfer occurs when said analyte-analogue is not bound to said analyte binding ligand. - View Dependent Claims (14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24)
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25. A method of identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer, said method comprising:
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a) contacting a combinatorial library comprising a plurality of ligands with an analyte-analogue such that said analyte-analogue binds to at least one of said ligands to form an analyte-ligand binding pair, said ligands comprising a first label comprising a first component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair, at least one of said analyte-analogue and said ligands comprising a second label comprising a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair; and
b) detecting an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer. - View Dependent Claims (26, 27, 28, 29, 30, 31, 32, 33)
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34. A method of identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer, said method comprising:
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a) determining a constant region on a ligand at which to attach at least one component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair;
b) obtaining a predetermined analyte binding ligand from a combinatorial library comprising ligands comprising said predetermined constant region, said analyte binding ligand having been predetermined by contacting the combinatorial library with a first analyte-analogue, and selecting an analyte binding ligand capable of binding the first analyte-analogue; and
c) attaching a label comprising at least one of a first component and a second component of said non-radiative fluorescence resonance energy transfer donor-acceptor pair to at least one of said analyte binding ligand and a second analyte-analogue such that non-radiative fluorescence resonance energy transfer occurs when said second analyte-analogue is bound to said analyte binding ligand, and a change in non-radiative fluorescence resonance energy transfer occurs when said second analyte-analogue is not bound to said analyte binding ligand. - View Dependent Claims (35, 36, 37, 38, 39, 40, 41, 42)
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43. A method of identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer, said method comprising:
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a) determining a region on an analyte-analogue at which to attach a component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair;
b) preparing an analyte-analogue comprising said predetermined region;
c) contacting a combinatorial library comprising ligands with said analyte-analogue;
d) identifying a ligand to which said analyte-analogue binds, said ligand being the analyte binding ligand; and
e) attaching a label comprising at least one of a first component and a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair to at least one of said analyte binding ligand and said analyte-analogue such that non-radiative fluorescence resonance energy transfer occurs when said analyte-analogue is bound to said analyte binding ligand, and a change in non-radiative fluorescence resonance energy transfer when said analyte-analogue is not bound to said analyte binding ligand. - View Dependent Claims (44, 45, 46)
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47. A method of identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer, said method comprising:
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a) identifying a linking moiety to which at least one component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair binds;
b) obtaining a predetermined analyte binding ligand from a combinatorial library comprising ligands, said analyte binding ligand having been predetermined by contacting the combinatorial library with a first analyte-analogue, and selecting an analyte binding ligand capable of binding the first analyte-analogue; and
c) attaching a label to said linking moiety, said label comprising a first component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair;
d) attaching a label comprising a second component of said non-radiative fluorescence resonance energy transfer donor-acceptor pair to at least one of said analyte binding ligand and a second analyte-analogue; and
e) attaching said linking moiety to said analyte binding ligand, wherein non-radiative fluorescence resonance energy transfer occurs when said second analyte-analogue is bound to said analyte binding ligand, and a change in non-radiative fluorescence resonance energy transfer when said second analyte-analogue is not bound to said analyte binding ligand. - View Dependent Claims (48)
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49. A method of screening a combinatorial library, said method comprising
a) preparing a combinatorial library comprising ligands comprising a first component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair; -
b) contacting said combinatorial library with an analyte-analogue comprising a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair; and
c) identifying an analyte-ligand binding pair that exhibits non-radiative fluorescence resonance energy transfer.
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50. A sensor comprising:
an analyte-ligand binding pair comprising a) a first analyte-analogue, and b) a predetermined analyte binding ligand, said analyte binding ligand having been predetermined by contacting a combinatorial library with a second analyte-analogue and selecting a ligand to which the second analyte-analogue binds, c) a label comprising a first component and a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair, said analyte-ligand binding pair exhibiting non-radiative fluorescence resonance energy transfer when the first analyte-analogue is bound to said analyte binding ligand, and a change in non-radiative fluorescence resonance energy transfer when the first analyte-analogue is not bound to said analyte binding ligand. - View Dependent Claims (51, 52, 53, 54)
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55. A method of making a sensor, said method comprising:
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a) selecting an analyte-analogue;
b) attaching a label comprising a first component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair to an analyte-analogue;
c) selecting an analyte binding ligand from a combinatorial library, said analyte binding ligand being capable of binding with said analyte-analogue;
d) attaching a label comprising a second component of a non-radiative fluorescence resonance energy transfer donor-acceptor pair to said analyte binding ligand; and
e) encapsulating said labeled analyte binding ligand and said labeled analyte-analogue, said sensor exhibiting either non-radiative fluorescence resonance energy transfer when said analyte-analogue is bound to said analyte binding ligand, and a change in non-radiative fluorescence resonance energy transfer when said analyte-analogue is not bound to said analyte binding ligand, or being free from non-radiative fluorescence resonance energy transfer when said analyte-analogue is bound to said analyte binding ligand, and exhibiting non-radiative fluorescence resonance energy transfer when said analyte-analogue is bound to said analyte binding ligand.
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Specification