Caged sensors, regulators and compounds and uses thereof

• US 20040166553A1
• Filed: 11/17/2003
• Published: 08/26/2004
• Est. Priority Date: 11/18/2002
• Status: Active Grant

• Current Assignee: Affymetrix, Inc. (Thermo Fisher Scientific Incorporated)
• Original Assignee: GenoSpectra, Inc. (Thermo Fisher Scientific Incorporated)
• Inventors: McMaster, Gary, Nguyen, Quan
• Granted Patent: US 7,541,193 B2
• Time in Patent Office: Days
• US Class Current: 435/15
• CPC Class Codes: B82Y 10/00 Nanotechnology for information processing, storage or transmission, e.g. quantum computing or single electron logic
• CPC Class Codes: B82Y 5/00 Nanobiotechnology or nanomedicine, e.g. protein engineering or drug delivery
• CPC Class Codes: C12Q 1/37 involving peptidase or proteinase
• CPC Class Codes: C12Q 1/485 involving kinase
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 2525/207 siRNA, miRNA
• CPC Class Codes: C12Q 2537/1373 Displacement by a nucleic acid
• CPC Class Codes: G01N 33/542 with steric inhibition or signal modification, e.g. fluorescent quenching
• CPC Class Codes: G01N 33/573 for enzymes or isoenzymes

Abstract

A number of caged (e.g., photoactivatable) compositions are provided, for example, caged enzyme sensors, caged binding sensors, caged nucleic acid probes, caged interfering RNAs, caged antisense nucleic acids, caged ribozymes, caged biomolecular analogs, caged transcription factors, caged antibodies, caged molecular decoys, and caged aptamers. Compositions comprising a plurality of caged components are also provided, as are labeled and optionally caged modulators. Also provided are kits for making the caged compositions and methods for making and for using the caged compositions. Related systems and devices are also provided (e.g., an uncaging device that can trigger photoactivatable assays and optionally detect signals from the assays).

Claims

1. A composition, comprising:

a cell comprising a caged sensor for detecting an activity of an enzyme, which caged sensor comprises;

a) one or more molecules collectively comprising;

i) a substrate for the enzyme, wherein the substrate is in a first state on which the enzyme can act, thereby converting the substrate to a second state, and

ii) a first label, wherein a first signal exhibited by the first label when the substrate is in its first state is distinguishable from a second signal exhibited by the first label when the substrate is in its second state; and

,

b) one or more first caging groups associated with the one or more molecules, the first caging groups inhibiting the enzyme from acting upon the substrate.

3. The composition of claim 1 or 2, wherein the first caging groups inhibit the enzyme from acting upon the substrate by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the substrate in the absence of the first caging groups.

4. The composition of claim 1 or 2, wherein the first caging groups prevent the enzyme from acting upon the substrate.

5. The composition of claim 1 or 2, wherein removal of, or an induced conformational change in, the first caging groups permits the enzyme to act upon the substrate.

6. The composition of claim 1 or 2, wherein the first label is an optically detectable label or a fluorophore;

wherein the first signal and/or the second signal is an optical signal, a fluorescent signal, a luminescent signal, a nonoptical signal, or a magnetic signal;

or wherein the first signal is a fluorescent emission at a first wavelength with a first intensity and the second signal is a fluorescent emission at the first wavelength with a second intensity substantially greater or less than the first intensity.

7. The composition of claim 1 or 2, wherein the one or more first caging groups associated with the one or more molecules are covalently attached to the one or more molecules.

8. The composition of claim 1 or 2, wherein the one or more first caging groups are removable by sonication, photoactivatable, or photolabile;

or wherein the first caging groups can be removed by exposure to light with a wavelength between about 60 nm and about 400 nm, between about 400 nm and about 700 nm, and/or between about 700 nm and about 1000 nm.

9. The composition of claim 1 or 2, wherein the first label and the substrate are physically connected.

10. The composition of claim 1 or 2, wherein the substrate comprises one or more of:

an amino acid, a polypeptide, a nitrogenous base, a nucleoside, a nucleotide, a nucleic acid, a carbohydrate, or a lipid.

13. The composition of claim 1, wherein the enzyme is an oxidoreductase, a transferase, a hydrolase, a lyase, a ligase, an isomerase, a phosphatase, a GTPase, an ATPase, a phosphodiesterase, a luciferase, an acetylase, a glycosylase, a ubiquitin-conjugating enzyme, a hydrogenase, a polymerase, a peroxidase, a protease, or a caspase.

14. The composition of claim 13, wherein the enzyme is a caspase, and wherein one polypeptide comprises the substrate for the caspase and the first label and comprises a second label or a quencher;

wherein the first label and the second label or the quencher interact to produce the first signal when the substrate is intact; and

wherein cleavage of the substrate by the caspase prevents the interaction of the first label and the second label or the quencher, thereby resulting in production of the second signal.

15. The composition of claim 14, wherein the caspase is caspase 3, the substrate comprises an Asp-Glu-Val-Asp motif, and the one or more first caging groups are located on one or more of the amino acid residues in the Asp-Glu-Val-Asp motif.

16. The composition of claim 14, wherein one of the first label and the second label or the quencher is located at the N-terminus of the polypeptide and the other of the first label and the second label or the quencher is located at the C-terminus of the polypeptide.

17. The composition of claim 14, wherein the first and second labels are fluorophores capable of exhibiting FRET, or wherein the first label is FITC and the second label is rhodamine or coumarin.

18. The composition of claim 1 or 2, wherein the enzyme is a protein kinase that phosphorylates tyrosine, serine and/or threonine.

19. The composition of claim 18, wherein one polypeptide comprises the first label and the substrate for the kinase, the substrate comprising a serine, threonine, or tyrosine residue capable of being phosphorylated by the kinase;

wherein the first label is located at the serine, threonine, or tyrosine residue and exhibits the first signal when the residue is not phosphorylated and the second signal when the residue is phosphorylated.

20. The composition of claim 19, wherein the one or more first caging groups are located on one or more amino acid residues involved in binding the kinase.

21. The composition of claim 18, wherein one polypeptide comprises the substrate for the kinase and the first label and comprises a second label or a quencher;

wherein the first label and the second label or the quencher interact to produce the first signal when the substrate is not phosphorylated; and

wherein phosphorylation of the substrate prevents the interaction of the first label and the second label or the quencher, thereby resulting in production of the second signal.

22. The composition of claim 21, wherein the one or more first caging groups are located on one or more amino acid residues involved in binding the kinase and/or on a residue that can be phosphorylated by the kinase.

23. The composition of claim 21, wherein one of the first label and the second label or the quencher is located at the N-terminus of the polypeptide and the other of the first label and the second label or the quencher is located at the C-terminus of the polypeptide.

24. The composition of claim 21, wherein the first and second labels are hydrophobic fluorophores, or wherein the first label is FITC and the second label is rhodamine or coumarin.

25. The composition of claim 21, wherein phosphorylation of the substrate triggers a conformational change in the polypeptide, the conformational change preventing the interaction of the first label and the second label or the quencher;

or wherein phosphorylation of the substrate results in binding of a phosphobinder to the phosphorylated substrate, the binding of the phosphobinder preventing the interaction of the first label and the second label or the quencher.

26. The composition of claim 25, wherein the phosphobinder is associated with one or more second caging groups, the presence of which prevents the phosphobinder from binding the phosphorylated substrate.

27. The composition of claim 26, wherein the second caging groups are removable under different conditions than the first caging groups preventing phosphorylation of the substrate.

28. The composition of claim 25, wherein the phosphobinder comprises an antibody, an SH-2 domain, a PTB domain, a 14-3-3 domain, an FHA domain, a WD40 domain and/or a WW domain.

29. The composition of claim 18, wherein one polypeptide comprises the substrate for the kinase and the first label;

wherein the polypeptide comprises a phosphobinder and a second label or a quencher;

wherein the first label and the second label or the quencher do not interact when the substrate is not phosphorylated, thereby producing the first signal; and

wherein phosphorylation of the substrate results in intramolecular binding of the phosphobinder to the phosphorylated substrate, the intramolecular binding resulting in the interaction of the first label and the second label or the quencher, thereby producing the second signal;

or, wherein a first polypeptide comprises the substrate for the kinase and the first label;

wherein a second polypeptide comprises a phosphobinder and a second label or a quencher;

wherein the first label and the second label or the quencher do not interact when the substrate is not phosphorylated, thereby producing the first signal; and

wherein phosphorylation of the substrate results in intermolecular binding of the phosphobinder to the phosphorylated substrate, the intermolecular binding resulting in the interaction of the first label and the second label or the quencher, thereby producing the second signal.

30. The composition of claim 29, wherein the one or more first caging groups are located on one or more amino acid residues involved in binding the kinase and/or on a residue that can be phosphorylated by the kinase.

31. The composition of claim 29, wherein one of the first label and the second label or the quencher is located at the N-terminus of the polypeptide and the other of the first label and the second label or the quencher is located at the C-terminus of the polypeptide.

32. The composition of claim 29, wherein the first and second labels are fluorophores capable of exhibiting FRET.

33. The composition of claim 29, wherein the phosphobinder is associated with one or more second caging groups, the presence of which prevents the phosphobinder from binding the phosphorylated substrate.

34. The composition of claim 33, wherein the second caging groups are removable under different conditions than the first caging groups preventing phosphorylation of the substrate.

35. The composition of claim 29, wherein the phosphobinder comprises an antibody, an SH-2 domain, a PTB domain, a 14-3-3 domain, an FHA domain, a WD40 domain and/or a WW domain.

36. The composition of claim 18, wherein one polypeptide comprises the substrate for the kinase, a second substrate, the first label, a third label, a fourth label or a quencher, and a phosphobinder;

the substrate comprising a serine, threonine, or tyrosine residue capable of being phosphorylated by the kinase;

the second substrate being associated with one or more third caging groups, the presence of which prevents phosphorylation of the second substrate;

wherein the first label is located at the serine, threonine, or tyrosine residue and exhibits the first signal when the residue is not phosphorylated and the second signal when the residue is phosphorylated;

wherein the third label and the fourth label or the quencher do not interact when the second substrate is not phosphorylated, thereby producing a third signal; and

wherein phosphorylation of the second substrate results in intramolecular binding of the phosphobinder to the phosphorylated second substrate, the intramolecular binding resulting in the interaction of the third label and the fourth label or the quencher, thereby producing a fourth signal, the fourth signal distinguishable from the first, second and third signals.

37. The composition of claim 36, wherein the second substrate is for the same kinase or for a different kinase.

38. The composition of claim 36, wherein the one or more first caging groups are located on one or more amino acid residues involved in binding the kinase that phosphorylates the first substrate;

or wherein the one or more third caging groups are located on one or more amino acid residues involved in binding the kinase and/or on a residue that can be phosphorylated by the kinase.

39. The composition of claim 38, wherein the third caging groups preventing phosphorylation of the second substrate are removable under different conditions than the first caging groups preventing phosphorylation of the substrate.

40. The composition of claim 36, wherein one of the third label and the fourth label or the quencher is located at the C-terminus of the polypeptide and the other of the third label and the fourth label or the quencher is within the polypeptide.

41. The composition of claim 36, wherein the third and fourth labels are fluorophores capable of exhibiting FRET.

42. The composition of claim 36, wherein the phosphobinder is associated with one or more second caging groups, the presence of which prevents the phosphobinder from binding the phosphorylated second substrate.

43. The composition of claim 42, wherein the second caging groups are removable under different conditions than the first caging groups preventing phosphorylation of the substrate and/or under different conditions than the third caging groups preventing phosphorylation of the second substrate.

44. The composition of claim 36, wherein the phosphobinder comprises an antibody, an SH-2 domain, a PTB domain, a 14-3-3 domain, an FHA domain, a WD40 domain and/or a WW domain.

45. The composition of claim 1 or 2, wherein the one or more molecules comprise a fifth label, the fifth label exhibiting a unique fifth signal, the fifth signal being independent of the state of the substrate.

46. The composition of claim 45, wherein the fifth label is a fluorophore or a quantum dot.

47. The composition of claim 1 or 2, wherein the one or more molecules are associated with a cellular delivery module that can mediate introduction of the sensor into a cell.

48. The composition of claim 47, wherein the cellular delivery module comprises a polypeptide, a PEP-1 peptide, an amphipathic peptide, a cationic peptide, a protein transduction domain, a protein transduction domain derived from an HIV-1 Tat protein, from a herpes simplex virus VP22 protein, or from a Drosophila antennapedia protein, or a model protein transduction domain.

49. The composition of claim 47, wherein the cellular delivery module is covalently attached to the one or more molecules.

50. The composition of claim 49, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

51. The composition of claim 47, wherein the cellular delivery module is associated with one or more fourth caging groups, the presence of which prevents the cellular delivery module from mediating introduction of the sensor into a cell.

52. The composition of claim 1 or 2, wherein the one or more molecules are associated with at least one subcellular delivery module.

53. The composition of claim 52, wherein the subcellular delivery module comprises a polypeptide, a nucleic acid, and/or a carbohydrate;

wherein the subcellular delivery module mediates localization of the sensor to one or more of;

a membrane, a mitochondrion, a peroxisome, a nucleus, an endoplasmic reticulum, a Golgi, a vesicle, a lysosome, an endosome, or a chloroplast;

wherein the subcellular delivery module comprises one or more of;

a mitochondrial matrix-targeting sequence, a nuclear localization signal, a signal peptide, an ER retention signal, a peroxisomal targeting motif, a chloroplast stromal targeting sequence, a transmembrane domain, or a lipid attachment site;

or wherein the subcellular delivery module comprises a binding domain that mediates localization of the sensor by binding to a target protein.

54. The composition of claim 52, wherein the subcellular delivery module is covalently attached to the one or more molecules.

55. The composition of claim 54, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

56. The composition of claim 52, wherein the subcellular delivery module is associated with one or more fifth caging groups, the presence of which prevents the subcellular delivery module from mediating subcellular localization of the sensor.

59. The composition of claim 1 or 2, wherein the caged sensor comprises a first oligonucleotide, the first oligonucleotide being complementary to a second oligonucleotide, the second oligonucleotide being bound to a matrix.

60. The composition of claim 59, wherein the matrix is a surface, and the second oligonucleotide is bound to the surface at a predetermined location within an array comprising other oligonucleotides;

or wherein the matrix comprises a bead.

61. A kit for making the caged sensor of claim 1 or 2, comprising a substrate, a first label, one or more first caging groups, and instructions for assembling the substrate, the first label, and the first caging groups to form the caged sensor, packaged in one or more containers;

or comprising a first label, one or more first caging groups, and instructions for assembling the first label, the first caging groups, and a substrate supplied by a user of the kit to form the caged sensor, packaged in one or more containers.

194. A system comprising the composition of claim 1, 2, 62, 93, 135, 158, or 186, comprising a fluid handling element, a plate handling element, a detector, and/or a source of uncaging energy.

2. A composition, comprising:

a caged sensor for detecting an activity of an enzyme, which caged sensor comprises;

a) one or more molecules collectively comprising;

i) a substrate for the enzyme, wherein the substrate is in a first state on which the enzyme can act, thereby converting the substrate to a second state, wherein the first state is not converted to the second state by cleavage by the enzyme, and

ii) a first label, wherein a first signal exhibited by the first label when the substrate is in its first state is distinguishable from a second signal exhibited by the first label when the substrate is in its second state; and

,

b) one or more first caging groups associated with the one or more molecules, the first caging groups inhibiting the enzyme from acting upon the substrate.

11. The composition of claim 2, comprising the enzyme, a cell, a cell comprising the caged sensor, a cell comprising the enzyme, or a cell lysate.

12. The composition of claim 2, wherein the enzyme is an oxidoreductase, a transferase, a hydrolase, a lyase, a ligase, or an isomerase.

57. The composition of claim 2, wherein the caged sensor is bound to a matrix.

58. The composition of claim 57, wherein the matrix is a surface, and the sensor is bound to the surface at a predetermined location within an array comprising other sensors;

or wherein the matrix comprises a bead.

62. A composition, comprising:

a caged sensor, which caged sensor comprises;

one or more molecules collectively comprising a binding target and a first label; and

,

one or more first caging groups associated with the one or more molecules, the first caging groups inhibiting the binding target from binding to its partner,

wherein the binding target and the partner are capable of a stable interaction.

63. The composition of claim 62, wherein the caging groups result in at least about 75%, at least about 90%, at least about 95%, or at least about 98% inhibition of binding between the binding target and the partner, as compared to the binding target in the absence of the first caging groups.

64. The composition of claim 62, wherein the caging groups prevent the binding target from binding to its partner.

65. The composition of claim 62, wherein removal of or an induced conformational change in the caging groups permits the binding target to bind to its partner.

66. The composition of claim 62, wherein a first signal exhibited by the first label when the binding target is not bound to the partner is distinguishable from a second signal exhibited by the first label when the binding target is bound to the partner.

67. The composition of claim 66, wherein the first signal is a fluorescent emission at a first wavelength with a first intensity and the second signal is a fluorescent emission at the first wavelength with a second intensity substantially greater or less than the first intensity.

68. The composition of claim 66, wherein the one or more molecules comprise a second label, the second label exhibiting a unique third signal, the third signal being independent of binding of the binding target to the partner.

69. The composition of claim 68, wherein the second label is a fluorophore or a quantum dot.

70. The composition of claim 62, wherein a first signal exhibited by the first label is not affected by binding of the binding target to the partner.

71. The composition of claim 62, wherein the binding target comprises one or more of:

an amino acid, a polypeptide, an antibody, a nitrogenous base, a nucleoside, a nucleotide, a nucleic acid, a carbohydrate, or a lipid; and

/or wherein the partner comprises one or more of;

an amino acid, a polypeptide, an antibody, a nitrogenous base, a nucleoside, a nucleotide, a nucleic acid, a carbohydrate, or a lipid.

72. The composition of claim 62, wherein the binding target comprises an antibody and the partner comprises an epitope recognized by the antibody;

wherein the binding target comprises a modulator of an activity of a protein and the partner comprises the protein;

wherein the binding target comprises a polypeptide comprising an SH-2 domain, a PTB domain, a 14-3-3 domain, an FHA domain, a WD40 domain and/or a WW domain, and the partner comprises a phosphorylated polypeptide;

or wherein the binding target comprises at least one probe nucleic acid comprising a region of complementarity to a target nucleic acid and the partner comprises the target nucleic acid.

73. The composition of claim 62, wherein the first label is an optically detectable label, a fluorophore or a quantum dot;

or wherein the first label exhibits an optical signal, a fluorescent signal, a luminescent signal, a nonoptical signal, or a magnetic signal.

74. The composition of claim 62, wherein the one or more first caging groups associated with the one or more molecules are covalently attached to the one or more molecules.

75. The composition of claim 62, wherein the one or more first caging groups are removable by sonication, photoactivatable, or photolabile.

76. The composition of claim 62, wherein the first label and the binding target are physically connected.

77. The composition of claim 62, comprising the partner, a cell, a cell comprising the sensor, a cell comprising the partner, or a cell lysate.

78. The composition of claim 62, wherein the one or more molecules are associated with a cellular delivery module that can mediate introduction of the sensor into a cell.

79. The composition of claim 78, wherein the cellular delivery module comprises a polypeptide, a PEP-1 peptide, an amphipathic peptide, a cationic peptide, a protein transduction domain, a protein transduction domain derived from an HIV-1 Tat protein, from a herpes simplex virus VP22 protein, or from a Drosophila antennapedia protein, or a model protein transduction domain.

80. The composition of claim 78, wherein the cellular delivery module is covalently attached to the one or more molecules.

81. The composition of claim 80, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

82. The composition of claim 78, wherein the cellular delivery module is associated with one or more second caging groups, the presence of which prevents the cellular delivery module from mediating introduction of the sensor into a cell.

83. The composition of claim 62, wherein the one or more molecules are associated with at least one subcellular delivery module.

84. The composition of claim 83, wherein the subcellular delivery module comprises a polypeptide, a nucleic acid, and/or a carbohydrate;

wherein the subcellular delivery module mediates localization of the sensor to one or more of;

a membrane, a mitochondrion, a peroxisome, a nucleus, an endoplasmic reticulum, a Golgi, a vesicle, a lysosome, an endosome, or a chloroplast;

wherein the subcellular delivery module comprises one or more of;

a mitochondrial matrix-targeting sequence, a nuclear localization signal, a signal peptide, an ER retention signal, a peroxisomal targeting motif, a chloroplast stromal targeting sequence, a transmembrane domain, or a lipid attachment site;

or wherein the subcellular delivery module comprises a binding domain that mediates localization of the sensor by binding to a target protein.

85. The composition of claim 83, wherein the subcellular delivery module is covalently attached to the one or more molecules.

86. The composition of claim 85, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

87. The composition of claim 83, wherein the subcellular delivery module is associated with one or more third caging groups, the presence of which prevents the subcellular delivery module from mediating subcellular localization of the sensor.

88. The composition of claim 62, wherein the caged sensor is bound to a matrix.

89. The composition of claim 88, wherein the matrix is a surface, and the sensor is bound to the surface at a predetermined location within an array comprising other sensors;

or wherein the matrix comprises a bead.

90. The composition of claim 62, wherein the caged sensor comprises a first oligonucleotide, the first oligonucleotide being complementary to a second oligonucleotide, the second oligonucleotide being bound to a matrix.

91. The composition of claim 90, wherein the matrix is a surface, and the second oligonucleotide is bound to the surface at a predetermined location within an array comprising other oligonucleotides;

or wherein the matrix comprises a bead.

92. A kit for making the caged sensor of claim 62, comprising a binding target, a first label, one or more first caging groups, and instructions for assembling the binding target, the first label, and the first caging groups to form a caged sensor, packaged in one or more containers;

or comprising a first label, one or more first caging groups, and instructions for assembling the first label, the first caging groups, and a binding target supplied by a user of the kit to form a caged sensor, packaged in one or more containers.

93. A composition, comprising:

a) a caged nucleic acid probe, which caged nucleic acid probe comprises;

i) at least one probe nucleic acid comprising a region of complementarity to a target nucleic acid;

ii) a first label, wherein a first signal exhibited by the first label when the probe nucleic acid is not bound to the target nucleic acid is distinguishable from a second signal exhibited by the first label when the probe nucleic acid is bound to the target nucleic acid; and

,

iii) one or more first caging groups associated with the probe nucleic acid, the first caging groups inhibiting the probe nucleic acid from binding to the target nucleic acid;

b) a caged antisense nucleic acid and a target nucleic acid, which caged antisense nucleic acid comprises;

i) an antisense nucleic acid comprising a region of complementarity to the target nucleic acid; and

,

ii) one or more first caging groups associated with the antisense nucleic acid, the first caging groups inhibiting the antisense nucleic acid from binding to and inactivating the target nucleic acid;

c) a caged biomolecular analog, which caged biomolecular analog comprises;

i) a molecule comprising one or more nonnatural nucleotides and/or one or more nonnatural amino acids; and

,

ii) one or more caging groups associated with the molecule;

d) a caged transcription factor, which caged transcription factor comprises;

i) a transcription factor; and

,

ii) one or more first caging groups associated with the transcription factor, the first caging groups inhibiting the transcription factor from regulating transcription from a cognate promoter;

e) a caged antibody, which caged antibody comprises;

i) an antibody that binds an epitope;

ii) a cellular delivery module and/or at least one subcellular delivery module associated with the antibody; and

,

iii) one or more first caging groups associated with the antibody, the first caging groups inhibiting the antibody from binding to the epitope;

f) a caged molecular decoy, which caged molecular decoy comprises;

i) a molecular decoy; and

ii) one or more first caging groups associated with the molecular decoy;

or,

g) a caged aptamer, which caged aptamer comprises;

i) an aptamer that binds a ligand; and

,

ii) one or more first caging groups associated with the aptamer, the first caging groups inhibiting the aptamer from binding to the ligand.

94. The composition of claim 93, wherein the caging groups inhibit the probe nucleic acid of (a) from binding to the target nucleic acid by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the probe nucleic acid in the absence of the first caging groups;

wherein the caging groups inhibit the antisense nucleic acid of (b) from binding to and inactivating the target nucleic acid by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the antisense nucleic acid in the absence of the first caging groups;

wherein the caging groups inhibit the transcription factor of (d) from regulating transcription from the cognate promoter by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the transcription factor in the absence of the first caging groups;

wherein the caging groups inhibit the antibody of (e) from binding to the epitope by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the antibody in the absence of the first caging groups;

or wherein the caging groups inhibit the aptamer of (g) from binding to the ligand by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the aptamer in the absence of the first caging groups.

95. The composition of claim 93, wherein the caging groups prevent the probe nucleic acid of (a) from binding to the target nucleic acid, the antisense nucleic acid of (b) from binding to and inactivating the target nucleic acid, the transcription factor of (d) from regulating transcription from the cognate promoter, the antibody of (e) from binding to the epitope, or the aptamer of (g) from binding to the ligand.

96. The composition of claim 93, wherein removal of or an induced conformational change in the caging groups permits the probe nucleic acid of (a) to bind to the target nucleic acid, the antisense nucleic acid of (b) to bind to and inactivate the target nucleic acid, the transcription factor of (d) to regulate transcription from the cognate promoter, the antibody of (e) to bind to the epitope, or the aptamer of (g) to bind to the ligand.

97. The composition of claim 93, wherein the caged biomolecular analog of (c) comprises at least a first label;

wherein the transcription factor of (d) comprises a first label, wherein a first signal exhibited by the first label when the transcription factor is not bound to the cognate promoter is distinguishable from a second signal exhibited by the first label when the transcription factor is bound to the cognate promoter;

wherein the antibody of (e) comprises a first label, wherein a first signal exhibited by the first label when the antibody is not bound to the epitope is distinguishable from a second signal exhibited by the first label when the antibody is bound to the epitope;

wherein the molecular decoy of (f) comprises a first label;

or wherein the aptamer of (g) comprises a first label, wherein a first signal exhibited by the first label when the aptamer is not bound to the ligand is distinguishable from a second signal exhibited by the first label when the aptamer is bound to the ligand.

98. The composition of claim 93, wherein the probe nucleic acid of (a) comprises a second label, the second label exhibiting a unique third signal, the third signal being independent of the probe nucleic acid binding to the target nucleic acid;

wherein the transcription factor of (d) comprises a second label, the second label exhibiting a unique third signal, the third signal being independent of the binding of the transcription factor to DNA;

wherein the antibody of (e) comprises a second label, the second label exhibiting a unique third signal, the third signal being independent of the binding of the antibody to the epitope;

or wherein the aptamer of (g) comprises a second label, the second label exhibiting a unique third signal, the third signal being independent of the binding of the aptamer to the ligand.

99. The composition of claim 98, wherein the second label is a fluorophore or a quantum dot.

100. The composition of claim 93 or 97, wherein the first label is an optically detectable label or a fluorophore;

wherein the first signal and/or the second signal is an optical signal, a fluorescent signal, a luminescent signal, a nonoptical signal, or a magnetic signal;

or wherein the first signal is a fluorescent emission at a first wavelength with a first intensity and the second signal is a fluorescent emission at the first wavelength with a second intensity substantially greater or less than the first intensity.

101. The composition of claim 93, wherein the one or more first caging groups associated with the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g) are covalently attached to the probe nucleic acid, the antisense nucleic acid, the biomolecular analog, the transcription factor, the antibody, the molecular decoy, or the aptamer.

102. The composition of claim 93, wherein the one or more first caging groups are removable by sonication, photoactivatable, or photolabile;

or wherein the first caging groups can be removed by exposure to light with a wavelength between about 60 nm and about 400 nm, between about 400 nm and about 700 nm, and/or between about 700 nm and about 1000 nm.

103. The composition of claim 93, comprising the target nucleic acid of the probe nucleic acid of (a), the cognate promoter of the transcription factor of (d), the epitope of the antibody of (e), or the ligand of the aptamer of (g);

a cell;

a cell comprising the probe nucleic acid of (a), the antisense nucleic acid and the target nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g);

or a cell lysate.

104. The composition of claim 93, wherein the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the molecular decoy of (f), or the aptamer of (g) is associated with a cellular delivery module that can mediate introduction of the probe nucleic acid, the antisense nucleic acid, the biomolecular analog, the transcription factor, the molecular decoy, or the aptamer into a cell.

105. The composition of claim 104, wherein the cellular delivery module comprises a polypeptide, a PEP-1 peptide, an amphipathic peptide, a cationic peptide, a protein transduction domain, a protein transduction domain derived from an HIV-1 Tat protein, from a herpes simplex virus VP22 protein, or from a Drosophila antennapedia protein, or a model protein transduction domain.

106. The composition of claim 104, wherein the cellular delivery module is covalently attached to the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g).

107. The composition of claim 106, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

108. The composition of claim 104, wherein the cellular delivery module is associated with one or more second caging groups, the presence of which prevents the cellular delivery module from mediating introduction of the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g) into a cell.

109. The composition of claim 93, wherein the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g) is associated with at least one subcellular delivery module.

110. The composition of claim 109, wherein the subcellular delivery module comprises a polypeptide, a nucleic acid, and/or a carbohydrate;

wherein the subcellular delivery module mediates localization of the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g) to one or more of;

a membrane, a mitochondrion, a nucleus, or a chloroplast;

wherein the subcellular delivery module comprises one or more of;

a mitochondrial matrix-targeting sequence, a nuclear localization signal, or a chloroplast stromal targeting sequence;

or wherein the subcellular delivery module comprises a binding domain that mediates localization of the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g) by binding to a target protein.

111. The composition of claim 109, wherein the subcellular delivery module is covalently attached to the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g).

112. The composition of claim 111, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

113. The composition of claim 109, wherein the subcellular delivery module is associated with one or more third caging groups, the presence of which prevents the subcellular delivery module from mediating subcellular localization of the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g).

114. The composition of claim 93, wherein the caged probe nucleic acid of (a), antisense nucleic acid of (b), biomolecular analog of (c), transcription factor of (d), antibody of (e), molecular decoy of (f), or aptamer of (g) is bound to a matrix.

115. The composition of claim 114, wherein the matrix is a surface, and the probe nucleic acid of (a), the antisense nucleic acid of (b), the biomolecular analog of (c), the transcription factor of (d), the antibody of (e), the molecular decoy of (f), or the aptamer of (g) is bound to the surface at a predetermined location within an array comprising other nucleic acid probes, antisense nucleic acids, biomolecular analogs, transcription factors, antibodies, molecular decoys, or aptamers;

or wherein the matrix comprises a bead.

116. The composition of claim 93, wherein the caged probe nucleic acid of (a), antisense nucleic acid of (b), biomolecular analog of (c), transcription factor of (d), antibody of (e), molecular decoy of (f), or aptamer of (g) comprises a first oligonucleotide, the first oligonucleotide being complementary to a second oligonucleotide, the second oligonucleotide being bound to a matrix.

117. The composition of claim 116, wherein the matrix is a surface, and the second oligonucleotide is bound to the surface at a predetermined location within an array comprising other oligonucleotides;

or wherein the matrix comprises a bead.

118. The composition of claim 93, wherein the at least one probe nucleic acid of (a) comprises one probe nucleic acid or a plurality of probe nucleic acids.

119. The composition of claim 93, wherein the probe nucleic acid of (a) comprises an oligodeoxyribonucleotide or a peptide nucleic acid.

120. The composition of claim 93, wherein the probe nucleic acid of (a) comprises a molecular beacon.

121. The composition of claim 93, wherein the at least one probe nucleic acid of (a) comprises a first probe nucleic acid and a second probe nucleic acid, the first probe nucleic acid comprising a first region of complementarity to the target nucleic acid and the first label, the second probe nucleic acid comprising a second region of complementarity to the target nucleic acid and a third label, the first label being an acceptor fluorophore, and the third label being a donor fluorophore;

whereby binding of the first and second probe nucleic acids to the target nucleic acid brings the donor and acceptor fluorophores into proximity, permitting energy transfer between them and resulting in the second signal.

122. The composition of claim 121, wherein the first probe nucleic acid and/or the second probe nucleic acid forms a stem-loop structure when not bound to the target nucleic acid.

123. The composition of claim 122, wherein the first probe nucleic acid and/or the second probe nucleic acid comprises a quencher.

124. The composition of claim 93, wherein the antisense nucleic acid of (b) comprises an RNA or a peptide nucleic acid.

125. The composition of claim 93, wherein the caged biomolecular analog of (c) comprises one or more natural nucleotides and/or one or more natural amino acids.

126. The composition of claim 93, wherein the caged biomolecular analog of (c) is chemically synthesized.

127. The composition of claim 93, wherein the transcription factor of (d) is a transcriptional activator or a transcriptional repressor;

comprises one or more of;

a leucine zipper motif, a helix-loop-helix motif, an HMG domain, a helix-turn-helix motif, a homeo domain, a winged helix motif, a paired box domain, or a TEA domain;

comprises at least one zinc finger motif;

or comprises at least one Cys₂His₂-type zinc finger motif.

128. The composition of claim 93, comprising a cell comprising the transcription factor of (d), wherein the cell is a prokaryotic cell, a eubacterial cell, an archaebacterial cell, an Escherichia coli, a eukaryotic cell, a mammalian cell, or a yeast cell.

129. The composition of claim 93, wherein the antibody of (e) is a monoclonal antibody or a single-chain antibody.

130. The composition of claim 93, wherein the molecular decoy of (f) comprises one or more of:

an amino acid, a polypeptide, a nitrogenous base, a nucleoside, a nucleotide, a nucleic acid, a carbohydrate, or a lipid.

131. The composition of claim 130, wherein the molecular decoy is a nucleic acid comprising a double-stranded DNA comprising a transcription factor binding sequence, a nucleic acid comprising a viral assembly sequence, or a protein that binds a pathogenic protein or pathogenic nucleic acid.

132. The composition of claim 93, comprising the target nucleic acid of (a), the epitope of (e), or the ligand of (g).

133. A kit for making the caged nucleic acid probe, the caged antisense nucleic acid, the caged biomolecular analog, the caged transcription factor, the caged antibody, the caged molecular decoy, or the caged aptamer of claim 93, comprising:

a probe nucleic acid, a first label, one or more first caging groups, and instructions for assembling the probe nucleic acid, the first label, and the first caging groups to form a caged nucleic acid probe, packaged in one or more containers;

a first label, one or more first caging groups, and instructions for assembling the first label, the first caging groups, and a probe nucleic acid supplied by a user of the kit to form a caged nucleic acid probe, packaged in one or more containers;

an antisense nucleic acid, one or more first caging groups, and instructions for assembling the antisense nucleic acid and the first caging groups to form the caged antisense nucleic acid, packaged in one or more containers;

one or more first caging groups and instructions for assembling the first caging groups and an antisense nucleic acid supplied by a user of the kit to form the caged antisense nucleic acid, packaged in one or more containers;

a biomolecular analog, one or more caging groups, and instructions for assembling the biomolecular analog and the caging groups to form the caged biomolecular analog, packaged in one or more containers;

one or more caging groups and instructions for assembling the caging groups and a biomolecular analog supplied by a user of the kit to form the caged biomolecular analog, packaged in one or more containers;

a transcription factor, one or more first caging groups, and instructions for assembling the transcription factor and the first caging groups to form the caged transcription factor, packaged in one or more containers;

one or more first caging groups and instructions for assembling the first caging groups and a transcription factor supplied by a user of the kit to form the caged transcription factor, packaged in one or more containers;

an antibody, one or more first caging groups, one or more cellular delivery modules and/or at least one subcellular delivery module, and instructions for assembling the antibody, the first caging groups, and the delivery modules to form the caged antibody, packaged in one or more containers;

one or more first caging groups, one or more cellular delivery modules and/or at least one subcellular delivery module, and instructions for assembling the first caging groups, the delivery modules, and an antibody supplied by a user of the kit to form the caged antibody, packaged in one or more containers;

a molecular decoy, one or more first caging groups, and instructions for assembling the molecular decoy and the first caging groups to form the caged molecular decoy, packaged in one or more containers;

one or more first caging groups and instructions for assembling the first caging groups and a molecular decoy supplied by a user of the kit to form the caged molecular decoy, packaged in one or more containers;

an aptamer, one or more first caging groups, and instructions for assembling the aptamer and the first caging groups to form the caged aptamer, packaged in one or more containers;

or one or more first caging groups and instructions for assembling the first caging groups and an aptamer supplied by a user of the kit to form the caged aptamer, packaged in one or more containers.

134. A kit for using the caged transcription factor of claim 93, comprising the caged transcription factor and instructions for using the caged transcription factor, packaged in one or more containers.

135. A cell, comprising:

a) a caged ribozyme, which caged ribozyme comprises;

i) a ribozyme, and,

ii) one or more first caging groups associated with the ribozyme, the first caging groups inhibiting the ribozyme from exhibiting an enzymatic activity, or

b) a caged antibody, which caged antibody comprises;

i) an antibody that binds an epitope, and

ii) one or more first caging groups associated with the antibody, the first caging groups inhibiting the antibody from binding to the epitope.

136. The cell of claim 135, wherein the caging groups inhibit the enzymatic activity of the ribozyme of (a) by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the ribozyme in the absence of the first caging groups, or wherein the caging groups inhibit binding between the antibody of (b) and the epitope by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the antibody in the absence of the first caging groups.

137. The cell of claim 135, wherein the caging groups prevent the ribozyme of (a) from exhibiting the enzymatic activity, or wherein the caging groups prevent the antibody of (b) from binding to the epitope.

138. The cell of claim 135, wherein removal of or an induced conformational change in the caging groups permits the ribozyme of (a) to exhibit the enzymatic activity or the antibody of (b) to bind to the epitope.

139. The cell of claim 135, wherein the one or more first caging groups associated with the ribozyme of (a) or the antibody of (b) are covalently attached to the ribozyme or antibody.

140. The cell of claim 135, wherein the one or more first caging groups are removable by sonication, photoactivatable, or photolabile;

or wherein the first caging groups can be removed by exposure to light with a wavelength between about 60 nm and about 400 nm, between about 400 nm and about 700 nm, and/or between about 700 nm and about 1000 nm.

141. The cell of claim 135, wherein the ribozyme of (a) or the antibody of (b) is associated with a cellular delivery module that can mediate introduction of the ribozyme or antibody into the cell.

142. The cell of claim 141, wherein the cellular delivery module comprises a polypeptide, a PEP-1 peptide, an amphipathic peptide, a cationic peptide, a protein transduction domain, a protein transduction domain derived from an HIV-1 Tat protein, from a herpes simplex virus VP22 protein, or from a Drosophila antennapedia protein, or a model protein transduction domain.

143. The cell of claim 141, wherein the cellular delivery module is covalently attached to the ribozyme of (a) or the antibody of (b).

144. The cell of claim 143, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

145. The cell of claim 141, wherein the cellular delivery module is associated with one or more second caging groups, the presence of which prevents the cellular delivery module from mediating introduction of the ribozyme of (a) or the antibody of (b) into the cell.

146. The cell of claim 135, wherein the ribozyme of (a) or the antibody of (b) is associated with at least one subcellular delivery module.

147. The cell of claim 146, wherein the subcellular delivery module comprises a polypeptide, a nucleic acid, and/or a carbohydrate;

wherein the subcellular delivery module mediates localization of the ribozyme of (a) or the antibody of (b) to one or more of;

a membrane, a mitochondrion, a peroxisome, a nucleus, an endoplasmic reticulum, a Golgi, a vesicle, a lysosome, an endosome, or a chloroplast;

wherein the subcellular delivery module comprises one or more of;

a mitochondrial matrix-targeting sequence, a nuclear localization signal, a signal peptide, an ER retention signal, a peroxisomal targeting motif, a chloroplast stromal targeting sequence, a transmembrane domain, or a lipid attachment site;

or wherein the subcellular delivery module comprises a binding domain that mediates localization of the ribozyme of (a) or the antibody of (b) by binding to a target protein.

148. The cell of claim 146, wherein the subcellular delivery module is covalently attached to the ribozyme of (a) or the antibody of (b).

149. The cell of claim 148, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

150. The cell of claim 146, wherein the subcellular delivery module is associated with one or more third caging groups, the presence of which prevents the subcellular delivery module from mediating subcellular localization of the ribozyme of (a) or the antibody of (b).

151. The cell of claim 135, wherein the ribozyme of (a) comprises a first label, wherein a first signal exhibited by the first label when the ribozyme is not active is distinguishable from a second signal exhibited by the first label when the ribozyme is active;

or wherein the antibody of (b) comprises a first label, wherein a first signal exhibited by the first label when the antibody is not bound to the epitope is distinguishable from a second signal exhibited by the first label when the antibody is bound to the epitope.

152. The cell of claim 151, wherein the first label is an optically detectable label or a fluorophore;

wherein the first signal and/or the second signal is an optical signal, a fluorescent signal, a luminescent signal, a nonoptical signal, or a magnetic signal;

or wherein the first signal is a fluorescent emission at a first wavelength with a first intensity and the second signal is a fluorescent emission at the first wavelength with a second intensity substantially greater or less than the first intensity.

153. The cell of claim 135, wherein the cell is a member of an array of cells.

154. The cell of claim 135, wherein the antibody of (b) is a monoclonal antibody or a single-chain antibody.

155. The cell of claim 135, wherein the antibody of (b) comprises a second label, the second label exhibiting a unique third signal, the third signal being independent of the binding of the antibody to the epitope.

156. The cell of claim 155, wherein the second label is a fluorophore or a quantum dot.

157. A kit for making the caged ribozyme or the caged antibody of claim 135, comprising:

a ribozyme, one or more first caging groups, and instructions for assembling the ribozyme and the first caging groups to form the caged ribozyme, packaged in one or more containers;

one or more first caging groups and instructions for assembling the first caging groups and a ribozyme supplied by a user of the kit to form the caged ribozyme, packaged in one or more containers;

an antibody, one or more first caging groups, and instructions for assembling the antibody and the first caging groups to form the caged antibody, packaged in one or more containers;

or one or more first caging groups and instructions for assembling the first caging groups and an antibody supplied by a user of the kit to form the caged antibody, packaged in one or more containers.

158. A composition, comprising a labeled modulator, which labeled modulator comprises

a modulator of an activity of a protein;

and,

a first label, wherein a first signal exhibited by the first label when the modulator is not bound to the protein is distinguishable from a second signal exhibited by the first label when the modulator is bound to the protein.

159. The composition of claim 158, wherein the modulator is an activator, an inhibitor, or a small molecule.

160. The composition of claim 158, wherein the protein is an enzyme, an oxidoreductase, a transferase, a hydrolase, a lyase, a ligase, an isomerase, a kinase, a protease, a phosphatase, a GTPase, an ATPase, a phosphodiesterase, a luciferase, an acetylase, a glycosylase, a ubiquitin-conjugating enzyme, a hydrogenase, a polymerase, or a peroxidase.

161. The composition of claim 160, wherein the enzyme is protein kinase C and the modulator is the inhibitor HB89.

162. The composition of claim 158, wherein the first label is an optically detectable label or a fluorophore;

wherein the first signal and/or the second signal is an optical signal, a fluorescent signal, a luminescent signal, a nonoptical signal, or a magnetic signal;

or wherein the first signal is a fluorescent emission at a first wavelength with a first intensity and the second signal is a fluorescent emission at the first wavelength with a second intensity substantially greater or less than the first intensity.

163. The composition of claim 158, comprising one or more first caging groups associated with the modulator, the first caging groups inhibiting the modulator from binding to the protein and/or affecting the activity of the protein.

164. The composition of claim 163, wherein the caging groups inhibit the modulator from binding to the protein and/or affecting the activity of the protein by at least about 75%, at least about 90%, at least about 95%, or at least about 98%, as compared to the modulator in the absence of the first caging groups.

165. The composition of claim 163, wherein the first caging groups prevent the modulator from binding to the protein and/or affecting the activity of the protein.

166. The composition of claim 163, wherein removal of or an induced conformational change in the first caging groups permits the modulator to bind to the protein and/or to affect the activity of the protein.

167. The composition of claim 163, wherein the one or more first caging groups associated with the modulator are covalently attached to the modulator.

168. The composition of claim 163, wherein the one or more first caging groups are removable by sonication, photoactivatable, or photolabile.

169. The composition of claim 158, comprising the protein, a cell, a cell comprising the labeled modulator, a cell comprising the protein, or a cell lysate.

170. The composition of claim 158, wherein the modulator comprise a second label, the second label exhibiting a unique third signal, the third signal being independent of the binding of the modulator to the protein.

171. The composition of claim 170, wherein the second label is a fluorophore or a quantum dot.

172. The composition of claim 158, wherein the modulator is associated with a cellular delivery module that can mediate introduction of the modulator into a cell.

173. The composition of claim 172, wherein the cellular delivery module comprises a polypeptide, a PEP-1 peptide, an amphipathic peptide, a cationic peptide, a protein transduction domain, a protein transduction domain derived from an HIV-1 Tat protein, from a herpes simplex virus VP22 protein, or from a Drosophila antennapedia protein, or a model protein transduction domain.

174. The composition of claim 172, wherein the cellular delivery module is covalently attached to the modulator.

175. The composition of claim 174, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

176. The composition of claim 172, wherein the cellular delivery module is associated with one or more second caging groups, the presence of which prevents the cellular delivery module from mediating introduction of the modulator into a cell.

177. The composition of claim 158, wherein the modulator is associated with at least one subcellular delivery module.

178. The composition of claim 177, wherein the subcellular delivery module comprises a polypeptide, a nucleic acid, and/or a carbohydrate;

wherein the subcellular delivery module mediates localization of the modulator to one or more of;

a membrane, a mitochondrion, a peroxisome, a nucleus, an endoplasmic reticulum, a Golgi, a vesicle, a lysosome, an endosome, or a chloroplast;

wherein the subcellular delivery module comprises one or more of;

a mitochondrial matrix-targeting sequence, a nuclear localization signal, a signal peptide, an ER retention signal, a peroxisomal targeting motif, a chloroplast stromal targeting sequence, a transmembrane domain, or a lipid attachment site;

or wherein the subcellular delivery module comprises a binding domain that mediates localization of the modulator by binding to a target protein.

179. The composition of claim 177, wherein the subcellular delivery module is covalently attached to the modulator.

180. The composition of claim 179, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

181. The composition of claim 177, wherein the subcellular delivery module is associated with one or more third caging groups, the presence of which prevents the subcellular delivery module from mediating subcellular localization of the modulator.

182. The composition of claim 158, wherein the labeled modulator is bound to a matrix.

183. The composition of claim 182, wherein the matrix is a surface, and the labeled modulator is bound to the surface at a predetermined location within an array comprising other labeled modulators;

or wherein the matrix comprises a bead.

184. A kit for making the labeled modulator of claim 158, comprising a modulator, a first label, and instructions for assembling the modulator and the first label to form the labeled modulator, packaged in one or more containers;

or a first label and instructions for assembling the first label and a modulator supplied by a user of the kit to form the labeled modulator, packaged in one or more containers.

185. A kit for making the caged labeled modulator of claim 163, comprising a modulator, a first label, one or more first caging groups, and instructions for assembling the modulator, the first label, and the first caging groups to form the caged labeled modulator, packaged in one or more containers;

or a first label, one or more first caging groups, and instructions for assembling the first label, the first caging groups, and a modulator supplied by a user of the kit to form the caged labeled modulator, packaged in one or more containers.

186. A composition comprising at least a first caged component and a second caged component.

187. The composition of claim 186, wherein the first caged component can be uncaged by exposure to energy of a first type and the second caged component can be uncaged by exposure to energy of a second type different from the first type.

188. The composition of claim 187, wherein the energy of the first type is light of a first wavelength and the energy of the second type is light of a second wavelength.

189. The composition of claim 186, wherein the composition comprises three or more caged components.

190. The composition of claim 186, wherein the caged components comprise one or more of:

a caged sensor, a caged nucleic acid probe, a caged modulator, a caged interfering RNA, a caged antisense nucleic acid, a caged ribozyme, a caged biomolecular analog, a caged transcription factor, a caged molecular decoy, a caged antibody, a caged aptamer, a caged small molecule, a caged nucleoside triphosphate, a caged chelating agent, or a caged metal ion.

191. The composition of claim 186, comprising a cell or a cell comprising the caged components.

192. The composition of claim 186, wherein the first and second caged components are not physically connected, or wherein the first and second caged components are physically connected.

193. The composition of claim 192, wherein the first and second caged components are connected by a photolabile linker.

195. A composition comprising a caged component, which caged component comprises a component, one or more first caging groups associated with the component, and a cellular delivery module associated with the component, which cellular delivery module can mediate introduction of the component into a cell.

196. The composition of claim 195, wherein the component comprises:

a) one or more molecules collectively comprising

i) a substrate for the enzyme, wherein the substrate is in a first state on which the enzyme can act, thereby converting the substrate to a second state, and

ii) a first label, wherein a first signal exhibited by the first label when the substrate is in its first state is distinguishable from a second signal exhibited by the first label when the substrate is in its second state;

b) one or more molecules collectively comprising a binding target and a first label, wherein the binding target and the partner are capable of a stable interaction;

c) a nucleic acid probe, comprising at least one probe nucleic acid comprising a region of complementarity to a target nucleic acid and a first label, wherein a first signal exhibited by the first label when the probe nucleic acid is not bound to the target nucleic acid is distinguishable from a second signal exhibited by the first label when the probe nucleic acid is bound to the target nucleic acid;

d) a biomolecular analog comprising a molecule comprising one or more nonnatural nucleotides and/or one or more nonnatural amino acids;

e) a transcription factor;

f) an antibody;

g) a molecular decoy;

h) an aptamer;

i) a ribozyme;

or

j) a modulator of an activity of a protein.

197. The composition of claim 195, wherein the cellular delivery module comprises a polypeptide, a PEP-1 peptide, an amphipathic peptide, a cationic peptide, a protein transduction domain, a protein transduction domain derived from an HIV-1 Tat protein, from a herpes simplex virus VP22 protein, or from a Drosophila antennapedia protein, a model protein transduction domain, a lipid, or a myristoyl group.

198. The composition of claim 195, wherein the cellular delivery module is covalently attached to the one or more molecules.

199. The composition of claim 198, wherein the covalent attachment is reversible by exposure to light of a preselected wavelength.

200. The composition of claim 195, wherein the cellular delivery module is associated with one or more second caging groups, the presence of which prevents the cellular delivery module from mediating introduction of the component into a cell.

201. A method of assaying an activity of an enzyme, the method comprising:

contacting the enzyme and a caged sensor by introducing the caged sensor into a cell, the caged sensor comprising one or more molecules collectively comprising;

a) a substrate for the enzyme, wherein the substrate is in a first state on which the enzyme can act, thereby converting the substrate to a second state, and a first label, wherein a first signal exhibited by the first label when the substrate is in its first state is distinguishable from a second signal exhibited by the first label when the substrate is in its second state, and

b) one or more caging groups associated with the one or more molecules, the caging groups inhibiting the enzyme from acting upon the substrate;

initiating the assay by exposing the enzyme and the caged sensor to uncaging energy of a first type, whereby exposure to the uncaging energy frees the sensor from inhibition by the first caging groups; and

,

assaying the activity of the enzyme.

204. The method of claim 201 or 203, comprising introducing a vector encoding the enzyme into the cell.

205. The method of claim 201 or 202, wherein the one or more caging groups prevent the enzyme from acting upon the substrate, and wherein removal of or an induced conformational change in the one or more caging groups permits the enzyme to act upon the substrate.

206. The method of claim 201 or 202, wherein exposing the enzyme and the caged sensor to uncaging energy of a first type comprises sonicating the enzyme and the caged sensor or exposing the enzyme and the caged sensor to light of a first wavelength.

207. The method of claim 206, wherein exposing the enzyme and the caged sensor to light of a first wavelength comprises exposing one or more preselected areas to the light.

208. The method of claim 207, wherein the one or more preselected areas comprise one or more wells of a multiwell plate;

wherein the one or more preselected areas comprise a plurality of the wells of a multiwell plate, and wherein exposing the preselected areas to the light comprises exposing the plurality of wells to the light simultaneously;

wherein the one or more preselected areas comprise at least about 12, at least about 24, at least about 48, at least about 96, at least about 384, or at least about 1536 wells of a multiwell plate;

wherein the one or more preselected areas comprise one or more channels of a microfluidic chip;

wherein the one or more preselected areas comprise a plurality of the channels of the microfluidic chip, and wherein exposing the preselected areas to the light comprises exposing the plurality of channels to the light simultaneously;

wherein the one or more preselected areas comprise one or more spots of a microarray;

wherein the one or more preselected areas comprise a plurality of the spots of the microarray, and wherein exposing the preselected areas to the light comprises exposing the plurality of spots to the light simultaneously;

or, wherein the one or more preselected areas comprise one or more regions of a cell, a tissue, or a body of an organism.

209. The method of claim 206, wherein exposing the enzyme and the caged sensor to light of a first wavelength comprises exposing the enzyme and the caged sensor to light wherein the intensity of the light and the duration of exposure to the light are controlled such that a first portion of the caged sensor is uncaged and a second portion of the caged sensor remains caged;

comprising repeating the assay by exposing the enzyme and the caged sensor to light of the first wavelength again.

210. The method of claim 209, wherein the first portion is a selected amount.

211. The method of claim 201 or 202, comprising detecting the first and/or the second signal.

212. The method of claim 201 or 202, wherein the caged sensor is bound to a matrix, or wherein the caged sensor comprises a first oligonucleotide, the first oligonucleotide being complementary to a second oligonucleotide, the second oligonucleotide being bound to a matrix.

213. The method of claim 201 or 202, comprising contacting the enzyme and a second caged component comprising one or more second caging groups, and exposing the second caged component to uncaging energy of a second type, whereby exposure to the uncaging energy frees the second component from inhibition by the second caging groups.

214. The method of claim 213, wherein the second caged component comprises a nucleoside triphosphate, ATP, a metal ion, a polypeptide, a nucleic acid, a carbohydrate, a lipid, a phosphobinder, or an antibody.

215. The method of claim 213, wherein exposing the second caged component to uncaging energy of a second type comprises exposing the second caged component to light of a second wavelength, different from the first and/or third wavelength.

216. The method of claim 201 or 202, comprising contacting the enzyme with a third caged component, wherein the third caged component comprises a third component required for termination of the assay and one or more third caging groups associated with the third component, the presence of the third caging groups inhibiting the third component from terminating the assay;

and comprising exposing the third caged component to uncaging energy of a third type, whereby exposure to the uncaging energy frees the third component from inhibition by the third caging groups.

217. The method of claim 216, wherein exposing the third caged component to uncaging energy of a third type comprises exposing the third caged component to light of a third wavelength, different from the first and/or second wavelength.

218. The method of claim 216, wherein the third component comprises an inhibitor, a chelating agent, EGTA or EDTA.

219. The method of claim 201 or 202, comprising contacting the enzyme with a fourth caged component, wherein the fourth caged component comprises a modulator and one or more fourth caging groups associated with the modulator;

and comprising exposing the fourth caged component to uncaging energy of a fourth type, whereby exposure to the uncaging energy frees the modulator from inhibition by the fourth caging groups.

220. The method of claim 201 or 202, comprising contacting a second enzyme with a fifth caged component, wherein the fifth caged component comprises a caged sensor for the second enzyme, the caged sensor comprising a sensor for the second enzyme and one or more fifth caging groups associated with the sensor;

comprising exposing the fifth caged component to uncaging energy of a fifth type, whereby exposure to the uncaging energy frees the sensor from inhibition by the fifth caging groups.

221. The method of claim 201 or 202, wherein contacting the enzyme with one or more reagents comprises introducing the first caged component into a subcellular compartment, a tissue, or an organism.

202. A method of assaying an activity of an enzyme, the method comprising:

contacting the enzyme and a caged sensor, the caged sensor comprising;

a) one or more molecules collectively comprising a substrate for the enzyme, wherein the substrate is in a first state on which the enzyme can act, thereby converting the substrate to a second state, wherein the first state is not converted to the second state by cleavage by the enzyme, and a first label, wherein a first signal exhibited by the first label when the substrate is in its first state is distinguishable from a second signal exhibited by the first label when the substrate is in its second state; and

,

b) one or more caging groups associated with the one or more molecules, the caging groups inhibiting the enzyme from acting upon the substrate;

initiating the assay by exposing the enzyme and the caged sensor to uncaging energy of a first type, whereby exposure to the uncaging energy frees the sensor from inhibition by the first caging groups; and

,

assaying the activity of the enzyme.

203. The method of claim 202, wherein contacting the enzyme and the caged sensor comprises introducing the caged sensor into a cell.

222. A method of assaying an intermolecular association between a first molecule and a second molecule which are capable of a stable interaction, the method comprising:

contacting the first and second molecules, wherein one or more caging groups are associated with at least the first molecule, the caging groups inhibiting the association of the first and second molecules;

initiating the assay by exposing the first and second molecules to uncaging energy, whereby exposure to the uncaging energy frees the first and/or second molecule from inhibition by the caging groups; and

,

assaying the association between the first and second molecules.

223. The method of claim 222, wherein exposing the first and second molecules to uncaging energy comprises sonicating the first and second molecules or exposing the first and second molecules to light of a first wavelength.

224. The method of claim 223, wherein exposing the first and second molecules to light of a first wavelength comprises exposing one or more preselected areas to the light.

225. The method of claim 224, wherein the one or more preselected areas comprise one or more wells of a multiwell plate;

wherein the one or more preselected areas comprise a plurality of the wells of a multiwell plate, and wherein exposing the preselected areas to the light comprises exposing the plurality of wells to the light simultaneously;

wherein the one or more preselected areas comprise at least about 12, at least about 24, at least about 48, at least about 96, at least about 384, or at least about 1536 wells of a multiwell plate;

wherein the one or more preselected areas comprise one or more channels of a microfluidic chip;

wherein the one or more preselected areas comprise a plurality of the channels of the microfluidic chip, and wherein exposing the preselected areas to the light comprises exposing the plurality of channels to the light simultaneously;

wherein the one or more preselected areas comprise one or more spots of a microarray;

wherein the one or more preselected areas comprise a plurality of the spots of the microarray, and wherein exposing the preselected areas to the light comprises exposing the plurality of spots to the light simultaneously;

or, wherein the one or more preselected areas comprise one or more regions of a cell, a tissue, or a body of an organism.

226. The method of claim 223, wherein exposing the first and second molecules to light of a first wavelength comprises exposing the first and second molecules to light wherein the intensity of the light and the duration of exposure to the light are controlled such that a first portion of the caged first molecule is uncaged and a second portion of the caged first molecule remains caged;

comprising repeating the assay by exposing the first and second molecules to light of the first wavelength again.

227. The method of claim 226, wherein the first portion is a selected amount.

228. The method of claim 222, wherein contacting the first and second molecules comprises introducing the caged first molecule into a cell, a subcellular compartment, a tissue, or an organism.

229. The method of claim 222, wherein the first molecule comprises a first label, wherein a first signal exhibited by the first label when the first molecule is not bound to the second molecule is distinguishable from a second signal exhibited by the first label when the first molecule is bound to the second molecule;

the method comprising detecting the first and/or second signal.

230. A method of detecting a target nucleic acid, the method comprising:

contacting the target nucleic acid and at least one caged probe nucleic acid, the caged probe nucleic acid comprising a probe nucleic acid and one or more caging groups associated with the probe nucleic acid, the probe nucleic acid comprising a region of complementary to a first strand of the target nucleic acid, the caging groups inhibiting the probe nucleic acid from binding to the first strand of the target nucleic acid;

exposing the caged probe nucleic acid and the target nucleic acid to uncaging energy, whereby exposure to the uncaging energy frees the probe nucleic acid from inhibition by the caging groups and permits the probe nucleic acid to bind to the first strand of the target nucleic acid; and

,

detecting a binding-dependent signal from the probe nucleic acid.

231. The method of claim 230, wherein exposing the caged probe nucleic acid and the target nucleic acid to uncaging energy comprises exposing the caged probe nucleic acid and the target nucleic acid to light of a first wavelength or sonicating the caged probe nucleic acid and the target nucleic acid.

232. The method of claim 230, wherein contacting the caged probe nucleic acid and the target nucleic acid comprises introducing the caged probe nucleic acid into a cell or a subcellular compartment.

233. A method of selectively inhibiting expression of a target nucleic acid in a cell, the method comprising:

introducing a caged antisense nucleic acid into the cell, the caged antisense nucleic acid comprising an antisense nucleic acid comprising a region of complementarity to the target nucleic acid and one or more caging groups associated with the antisense nucleic acid, the caging groups inhibiting the antisense nucleic acid from binding to and inactivating the target nucleic acid; and

,

exposing the cell to uncaging energy, whereby exposure to the uncaging energy frees the antisense nucleic acid from inhibition by the caging groups and permits the antisense nucleic acid to inactivate the target nucleic acid.

234. The method of claim 233, wherein exposing the cell to uncaging energy comprises exposing the cell to light of a first wavelength.

235. A method of activating a ribozyme, the method comprising:

providing a caged ribozyme, the caged ribozyme comprising a ribozyme and one or more caging groups associated with the ribozyme, the caging groups inhibiting the ribozyme from exhibiting an enzymatic activity;

introducing the caged ribozyme into a cell; and

,

exposing the caged ribozyme to uncaging energy, whereby exposure to the uncaging energy frees the ribozyme from inhibition by the caging groups and permits the ribozyme to exhibit the enzymatic activity.

236. The method of claim 235, wherein exposing the ribozyme to uncaging energy comprises exposing the ribozyme to light of a first wavelength or sonicating the ribozyme.

237. The method of claim 235, comprising introducing the caged ribozyme into a subcellular compartment.

238. A method of using a biomolecular analog, the method comprising:

providing a caged biomolecular analog, the caged biomolecular analog comprising a biomolecular analog and one or more caging groups associated with the biomolecular analog, the biomolecular analog comprising a molecule comprising one or more nonnatural nucleotides and/or one or more nonnatural amino acids; and

,

exposing the caged biomolecular analog to uncaging energy, whereby exposure to the uncaging energy frees the biomolecular analog from inhibition by the caging groups.

239. The method of claim 238, wherein exposing the caged biomolecular analog to uncaging energy comprises exposing the biomolecular analog to light of a first wavelength or sonicating the biomolecular analog.

240. The method of claim 238, comprising introducing the caged biomolecular analog into a cell or a subcellular compartment.

241. A method of regulating the expression of a gene comprising a first promoter in a cell, the method comprising:

introducing a caged transcription factor into the cell, the caged transcription factor comprising

a transcription factor capable of regulating transcription from the first promoter, and

one or more caging groups associated with the transcription factor, the caging groups inhibiting the transcription factor from regulating transcription from the first promoter; and

,

exposing the cell to uncaging energy, whereby exposure to the uncaging energy frees the transcription factor from inhibition by the caging groups and permits the transcription factor to regulate transcription from the first promoter.

242. The method of claim 241, wherein exposing the cell to uncaging energy comprises exposing the cell to light of a first wavelength.

243. The method of claim 241, wherein the transcription factor is a transcriptional activator.

244. A method of regulating transcription of a gene, the method comprising:

introducing a caged direct transcriptional modulator into a cell, wherein the caged transcriptional modulator comprises a transcriptional modulator and one or more caging groups associated with the transcriptional modulator, the caging groups inhibiting the transcriptional modulator from affecting expression of the gene; and

,

exposing the cell to uncaging energy, whereby exposure to the uncaging energy frees the transcriptional modulator from inhibition by the caging groups and permits the transcriptional modulator to regulate transcription of the gene.

245. A method of using a labeled modulator, the method comprising:

contacting a protein and the labeled modulator, the protein having an activity, and the labeled modulator comprising a modulator of the activity of the protein and a first label, wherein a first signal exhibited by the first label when the modulator is not bound to the protein is distinguishable from a second signal exhibited by the first label when the modulator is bound to the protein; and

,

detecting the first and/or second signal.

246. The method of claim 245, wherein the labeled modulator comprises one or more caging groups associated with the labeled modulator, the caging groups inhibiting the labeled modulator from binding to the protein and/or affecting the activity of the protein, the method comprising exposing the protein and the caged labeled modulator to uncaging energy, whereby exposure to the uncaging energy frees the labeled modulator from inhibition by the caging groups and permits the labeled modulator to bind to and affect the activity of the protein.

247. The method of claim 246, wherein exposing the caged labeled modulator to uncaging energy comprises exposing the caged labeled modulator to light of a first wavelength or sonicating the caged labeled modulator.

248. The method of claim 245, comprising introducing the labeled modulator into a cell or a subcellular compartment.

249. The method of claim 245, comprising introducing the labeled modulator into a cell, wherein detecting the first and/or second signal comprises determining the location of the labeled modulator within the cell.

250. A method of using a caged antibody, comprising:

introducing a caged antibody into a cell, the caged antibody comprising an antibody that binds an epitope and one or more caging groups associated with the antibody, the caging groups inhibiting the antibody from binding to the epitope; and

,

exposing the cell to uncaging energy, whereby exposure to the uncaging energy frees the antibody from inhibition by the caging groups and permits the antibody to bind the epitope.

251. The method of claim 250, wherein exposing the cell to uncaging energy comprises exposing the cell to light of a first wavelength.

252. The method of claim 250, wherein the antibody comprises a first label, wherein a first signal exhibited by the first label when the antibody is not bound to the epitope is distinguishable from a second signal exhibited by the first label when the antibody is bound to the epitope;

the method comprising detecting the first and/or second signal.

253. The method of claim 252, wherein detecting the first and/or second signal comprises determining the location of the antibody within the cell.

254. A method of using a molecular decoy, the method comprising:

contacting a caged molecular decoy and a protein, the caged molecular decoy comprising a molecular decoy that binds the protein and one or more caging groups associated with the molecular decoy, the caging groups inhibiting the molecular decoy from binding to the protein; and

,

exposing the caged molecular decoy to uncaging energy, whereby exposure to the uncaging energy frees the molecular decoy from inhibition by the caging groups and permits the molecular decoy to bind the protein.

255. The method of claim 254, wherein exposing the caged molecular decoy to uncaging energy comprises exposing the molecular decoy to light of a first wavelength or sonicating the molecular decoy.

256. The method of claim 254, comprising introducing the caged molecular decoy into a cell or a subcellular compartment.

257. A method of using an aptamer, the method comprising:

contacting a caged aptamer and a ligand, the caged aptamer comprising an aptamer that binds the ligand and one or more caging groups associated with the aptamer, the caging groups inhibiting the aptamer from binding to the ligand; and

,

exposing the caged aptamer to uncaging energy, whereby exposure to the uncaging energy frees the aptamer from inhibition by the caging groups and permits the aptamer to bind the ligand.

258. The method of claim 257, wherein exposing the caged aptamer to uncaging energy comprises exposing the aptamer to light of a first wavelength or sonicating the aptamer.

259. The method of claim 257, comprising introducing the caged aptamer into a cell or a subcellular compartment.

260. A method of using a plurality of caged components, the method comprising:

contacting at least a first caged component and a second caged component, wherein the first caged component can be uncaged by exposure to uncaging energy of a first type and the second caged component can be uncaged by exposure to uncaging energy of a second type different from the first type;

exposing the caged components to uncaging energy of the first type; and

,

exposing the caged components to uncaging energy of the second type.

261. The method of claim 260, wherein exposing the caged components to uncaging energy of the first type comprises exposing the caged components to light of a first wavelength, and/or wherein exposing the caged components to uncaging energy of the second type comprises exposing the caged components to light of a second wavelength.

262. The method of claim 260, comprising introducing the caged components into a cell prior to exposing them to uncaging energy.

263. A method of determining whether a test compound affects an activity of a GPCR, the method comprising:

providing a cell comprising the GPCR;

introducing at least one caged component into the cell, the caged component comprising a component and one or more caging groups associated with the component;

contacting the cell and the test compound;

exposing the cell to uncaging energy, whereby exposure to the uncaging energy frees the component from inhibition by the caging groups; and

,

detecting a signal from the component, the signal providing an indication of the activity of the GPCR.

264. The method of claim 263, wherein the one or more caging groups are photoactivatable or photolabile.

265. The method of claim 263, wherein the caged component is selected from the group consisting of:

a caged sensor for detecting an activity of an enzyme, a caged nucleic acid probe, and a caged interfering RNA sensor.

266. The method of claim 263, comprising contacting the cell and an activating compound, the activating compound increasing the activity of the GPCR.

267. A method of detecting modification of a substrate of interest, the method comprising:

introducing an initial substrate into a cell, wherein the initial substrate is caged by association with one or more caging moiety that inhibits modification of the substrate in the cell;

uncaging the initial substrate in the cell by dissociating part or all of the caging moiety from the substrate to provide an uncaged substrate of interest, or by changing the conformation of the initial substrate or caging moiety, or a combination thereof;

permitting the uncaged substrate of interest to be modified in the cell;

releasing the resulting modified substrate from the cell, or fixing the cell to fix the modified substrate in the cell;

contacting one or more modification detection reagents to the modified substrate, whereby said contact of the modification detection reagent to the modified substrate results in a detectable signal; and

,

detecting the detectable signal.

268. The method of claim 267, wherein the modified substrate is captured on one or more capture substrate prior to detecting the detectable signal.

269. The method of claim 268, wherein the modified substrate comprises one or more affinity moiety and the capture substrate comprises one or more affinity capture moiety, wherein the modified substrate is contacted to the capture substrate, whereby the affinity capture moiety binds the affinity moiety.

270. The method of claim 269, wherein the affinity moiety comprises:

avidin, biotin, a nucleic acid, an antibody, an antibody ligand, a polypeptide, a polypeptide ligand, a protein, a protein ligand, a His tag, a His tag binding moiety, Glutathione, GST, an HA tag, or a nucleic acid.

271. The method of claim 269, wherein the capture substrate comprises:

a microwell plate, a bead, an affinity column, or a microscope slide.

272. The method of claim 267, wherein the initial substrate is introduced into the cell by co-transfecting, into the cell, the initial substrate with one or more vector that comprises or encodes a substrate modification component, which substrate modification component modifies the uncaged substrate of interest.

273. The method of claim 272, wherein the substrate modification component comprises a catalytic molecule selected from the group consisting of:

an enzyme, a ribozyme, an oxidoreductase, a transferase, a hydrolase, a lyase, a ligase, an isomerase, a kinase, a phosphatase, a GTPase, an ATPase, a phosphodiesterase, a luciferase, an acetylase, a glycosylase, a ubiquitin-conjugating enzyme, a hydrogenase, a polymerase, a peroxidase, a protease, and a caspase.

274. The method of claim 273, wherein the uncaged substrate of interest is a substrate for the catalytic molecule.

275. The method of claim 267, wherein the uncaged substrate of interest is a specific substrate.

276. The method of claim 267, wherein the uncaged substrate of interest is a generic substrate.

277. The method of claim 272, wherein the vector comprises a nucleic acid that encodes the substrate modification component.

278. The method of claim 267, wherein the substrate is associated with a cellular delivery agent to enhance transport of the substrate into the cell, wherein the cellular delivery agent is selected from the group consisting of:

a polypeptide, a PEP-1 peptide, an MPG peptide, lipofectamine, an amphipathic peptide, a cationic peptide, a homopolymer of D-arginine, a homopolymer of histidine, a homopolymer of lysine, a protein transduction domain, an HIV-1 Tat protein, a herpes simplex virus VP22 protein, and a Drosophila antennapedia protein.

279. The method of claim 267, wherein uncaging the substrate comprises exposing the cell to one or more of:

light, a change in pH, a change in heat, or sonic energy.

280. The method of claim 267, wherein permitting the substrate to be modified in the cell comprises incubating the cell with the substrate for between about 5 seconds and about 240 minutes after uncaging the substrate prior to said releasing or fixing.

281. The method of claim 267, wherein releasing the modified substrate comprises lysing the cell.

282. The method of claim 267, wherein the detection reagent comprises a phosphobinder, an antibody that specifically binds to the modified substrate, a nucleic acid that specifically binds to the modified substrate, an aptamer that specifically binds to the modified substrate, or a combination thereof.

283. The method of claim 267, wherein detecting the signal comprises performing a homogeneous assay, a heterogeneous assay, FRET, Q-FRET, TR-FRET, or fluorescence polarization.

284. The method of claim 267, wherein the detection reagent comprises a detection reagent label comprising a donor or acceptor moiety, and the modified substrate comprises a substrate label comprising a donor or acceptor moiety, wherein binding of the detection reagent to the modified substrate results in an increase or decrease in signal from the detection reagent label or the substrate label, or both.

285. The method of claim 284, wherein the donor or acceptor moiety on the detection reagent label transfers energy to the substrate label, or wherein the substrate label transfers energy to the detection reagent label, wherein said energy transfer is dependent on proximity of the detection reagent label and the substrate label.

286. The method of claim 267, wherein the signal is detected using an optical detector.

287. The method of claim 267, comprising adding one or more actual or potential modulator of a substrate modification component to the cell and determining whether or to what extent the actual or potential substrate modulator modulates an activity of the substrate modification component on the uncaged substrate of interest.

288. The method of claim 287, wherein the actual or potential modulator is incubated in the cell for between about 0.5 hours and about 72 hours prior to uncaging the substrate.

289. The method of claim 267, wherein the cell is treated with trypsin and seeded into a microwell plate prior to uncaging the initial substrate.

290. A composition, comprising:

a cellular delivery agent, a caged substrate and a vector that comprises or encodes a substrate modification component, which substrate modification component modifies an uncaged substrate corresponding to the caged substrate.

291. The composition of claim 290, wherein the caged or uncaged substrate is an enzyme substrate that comprises a target site that is bound or modified by the substrate modification component, and wherein the caged or uncaged substrate comprises an affinity capture component.

292. The composition of claim 290, wherein the substrate modification component is selected from the group consisting of:

an enzyme, a ribozyme, an oxidoreductase, a transferase, a hydrolase, a lyase, a ligase, an isomerase, a kinase, a phosphatase, a GTPase, an ATPase, a phosphodiesterase, a luciferase, an acetylase, a glycosylase, a ubiquitin-conjugating enzyme, a hydrogenase, a polymerase, a peroxidase, a protease, and a caspase; and

wherein the caged substrate, when in uncaged form, is modified by the substrate modification component.

293. The composition of claim 290, wherein the cellular delivery agent comprises one or more of:

a polypeptide, a PEP-1 peptide, an MPG peptide, lipofectamine, an amphipathic peptide, a cationic peptide, a homopolymer of D-arginine, a homopolymer of histidine, a homopolymer of lysine, a protein transduction domain, an HIV-1 Tat protein, a herpes simplex virus VP22 protein, and a Drosophila antennapedia protein.

294. The composition of claim 290, comprising a cell that permits entry of the cellular delivery agent.

295. The composition of claim 290, wherein the vector comprises a nucleic acid that encodes the substrate modification component.

296. A system comprising:

an affinity capture substrate;

a cell or cell lysate comprising a modified enzyme substrate comprising an affinity capture moiety that can be specifically bound to the affinity capture substrate, wherein the modified enzyme substrate comprises a modification detection site that indicates whether a substrate modification component has acted upon the substrate;

a modification detection reagent that specifically binds to the modification detection site, wherein binding of the modification detection reagent to the modified substrate produces a detectable signal; and

,

a detector that detects the detectable signal during operation of the system.

297. The system of claim 296, wherein the affinity capture substrate comprises:

a microwell plate, a bead, an affinity column, or a microscope slide.

298. The system of claim 296, wherein the affinity capture substrate comprises an affinity moiety that comprises avidin, biotin, a nucleic acid, an antibody, an antibody ligand, a polypeptide, a polypeptide ligand, a protein, a protein ligand, a His tag, a His tag binding moiety, Glutathione, GST, an HA tag, or a nucleic acid.

299. The system of claim 296, wherein the modification detection reagent comprises a phosphobinder, an antibody that specifically binds to the modified substrate, a nucleic acid that specifically binds to the modified substrate, an aptamer that specifically binds to the modified substrate, or a combination thereof.

300. The system of claim 296, wherein the detector comprises a microscope, a CCD array, a camera, a spectrophotometer, or any combination thereof.

301. The system of claim 296, wherein the cell or cell lysate is treated with one or more actual or potential modulators of enzyme activity prior to detection by the modification detection reagent.

302. A system comprising:

a cell comprising a caged substrate of interest and a modification component;

an actual or potential modulator of the modification component;

a source of uncaging energy;

a cell lysis or fixation module;

a reporter that detects modifications to the substrate of interest;

a detector that detects a signal from the reporter; and

,

a computer comprising system memory or computer readable media comprising system instructions that direct the system to;

incubate the cell with the actual or potential modulator for a user selected period of time,

uncage the caged substrate of interest by exposing the cell to the source of uncaging energy for a selected period of time,

incubate the resulting energy-exposed cell that comprises a resulting uncaged substrate of interest for a user selected period of time,

lyse or fix the cell with the cell lysis or fixation module,

contact the resulting fixed cell or cell lysate with the reporter, and

detect the signal from the reporter.

303. A kit comprising:

an affinity capture substrate, a caged substrate comprising an affinity capture moiety, and detection reagent that detects one or more modification to the caged substrate when in uncaged form.