MULTIPURPOSE ANALYSIS USING SECOND HARMONIC GENERATING NANOPROBES

• US 20100233820A1
• Filed: 01/20/2010
• Published: 09/16/2010
• Est. Priority Date: 01/21/2009
• Status: Active Grant

• Current Assignee: California Institute of Technology
• Original Assignee: California Institute of Technology
• Inventors: Fraser, Scott E., Masmanidis, Sotirios, Pantazis, Periklis
• Granted Patent: US 8,945,471 B2
• Time in Patent Office: Days
• US Class Current: 436/94
• CPC Class Codes: B82Y 15/00 Nanotechnology for interacting, sensing or actuating, e.g. quantum dots as markers in protein assays or molecular motors
• CPC Class Codes: B82Y 20/00 Nanooptics, e.g. quantum optics or photonic crystals
• CPC Class Codes: C12Q 1/6818 involving interaction of two or more labels, e.g. resonant energy transfer
• CPC Class Codes: C12Q 2535/101 Sanger sequencing method, i.e. oligonucleotide sequencing using primer elongation and dideoxynucleotides as chain terminators
• CPC Class Codes: C12Q 2563/155 Particles of a defined size, e.g. nanoparticles
• CPC Class Codes: G01N 2021/6441 with two or more labels
• CPC Class Codes: G01N 21/636 using an arrangement of pump beam and probe beam; using the measurement of optical non-linear properties; non-linear optics per se G02F1/35
• CPC Class Codes: G01N 21/6428 Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes" in vivo A61B5/00; immunoassay G01N33/53
• CPC Class Codes: G01N 21/6458 Fluorescence microscopy fluorescence microscopes per se G02B21/0076 and G02B21/16
• CPC Class Codes: G02F 1/37 for second-harmonic generation G02F1/3532 takes precedence
• CPC Class Codes: G02F 2202/36 Micro- or nanomaterials
• CPC Class Codes: Y10T 436/143333 Saccharide [e.g., DNA, etc.]

Abstract

Second harmonic nanoprobes for multipurpose imaging of samples and a method of using such probes to monitor nucleotide sequencing in a Multi-SHG Detection Imaging (MSDI) modality and to monitor external electric field using voltage sensitive second harmonic generating (SHG) nanoprobes are provided. The SHG nanoprobes are comprised of various kinds of nanocrystals that do not possess an inversion symmetry and therefore are capable of generating second harmonic signals that can then be detected by conventional two-photon microscopy for in vivo imaging of biological processes and structures such as cell signaling, neuroimaging, protein conformation probing, DNA conformation probing, gene transcription, virus infection and replication in cells, protein dynamics, tumor imaging and cancer therapy evaluation and diagnosis as well as quantification in optical imaging for a wide-range of biological and non-biological processes and devices.

Claims

1. A second harmonic generating (SHG) nanoprobe system for nucleotide sequencing comprising:

a plurality of distinct SHG nanoprobes, each of said distinct SHG nanoprobes having no inversion symmetry such that each SHG nanoprobe generates a distinct harmonic emission when radiated by an external excitation source at least two wavelengths; and

an external excitation source for radiating the plurality of SHG nanoprobes;

wherein each of said distinct SHG nanoprobes is assigned and linked to a distinct nucleotide type and radiated by the external excitation source at least two frequencies such that a distinct emissive fingerprint is generated for each of said nucleotide types.

2. The SHG nanoprobe system of claim 1, wherein each of the SHG nanoprobes is a nanocrystal selected from the group consisting of organic, inorganic and combinations thereof.

3. The SHG nanoprobe system of claim 2, wherein each of the nanocrystals is selected from the group consisting of BaTiO₃, SiC, ZnO, ZnS, ZnSe, ZnTe, CdS, CdSe, CdTe, GaAs, GaSb, GaP, GaN, InSb, LiNbO₃, KNbO₃, KTiOPO₄, Fe(IO₃)₃, Au, Ag, N-(4-nitrophenyl)-(L)-prolinol (NPP), urea, 4-Nitroaniline, 2-Methyl-4-nitroaniline (MNA), 3-Methyl-4-methoxy-4′

-nitrostilbene), β

-BaB2O4 (Beta-Barium Borate/BBO, LiB3O5 (Lithium Triborate/LBO), LiNbO3 (Lithium Niobate/LN), KTiOPO4 (Potassium Titanyl Phosphate/KTP), AgGaS2 (Silver Thiogallate/AGS), AgGaSe2 (Silver Gallium Selenide/AGSe), ZnGeP2 (Zinc Germanium Phosphide/ZGP), GaSe (Gallium Selenide), KH2PO4 (Potassium Dihydrogen Phosphate/KDP), NH4H2PO4 (Ammonium Dihydrogen Phosphate (ADP), KD2PO4 (Deuterated Potassium Dihydrogen Phosphate/DKDP), CsLiB6O10 (Cesium Lithium Borate/CLBO), KTiOAsO4 (Potassium Titanyl Arsenate/KTA), KNbO3 (Potassium Niobate/KN), LiTaO3 (Lithium Tantalate/LT), RbTiOAsO4 (Rubidium Titanyl Arsenate/RTA), BaTiO3 (Barium Titanate), MgBaF4 (Magnesium Barium Fluoride), GaAs (Gallium Arsenide), BiB3O6 (Bismuth Triborate/BIBO), K2Al2B2O7 (Potassium Aluminum Borate/KABO), KBe2BO3F2 (Potassium Fluoroboratoberyllate/KBBF), BaAlBO3F2 (Barium Aluminum Fluoroborate/BABF), La2CaB10O19 (Lanthanum Calcium Borate/LCB), GdCa4O(BO3)3 (Gadolinium Calcium Oxyborate/GdCOB), YCa4O (BO3)3 (Yttrium Calcium Oxyborate/YCOB), Li2B4O7 (Lithium Tetraborate/LB4), LiRbB4O7 (Lithium Rubidium Tetraborate/LRB4), CdHg(SCN)4 (Cadmium Mercury Thiocyanate/CMTC), RbTiOPO4 (Rubidium Titanyl Phosphate/RTP), LiInS2 (Lithium Thioindate/LIS), LiInSe2 (Lithium Indium Selenide/LISe), KB5O8.4H2O (Potassium Pentaborate Tetrahydrate/KB5), CsB3O5 (Cesium Triborate/CBO), C4H7D12N4PO7 (Deuterated L-Arginine Phosphate Monohydrate/DLAP), a-HIO3 (a-Iodic Acid), LiCOOH.H2O (Lithium Formate Monohydrate/LFM), CsH2AsO4 (Cesium Dihydrogen Arsenate/CDA), CsD2AsO4 (Deuterated Cesium Dihydrogen Arsenate/DCDA), RbH2PO4 (Rubidium Dihydrogen Phosphate/RDP), CsTiOAsO4 (Cesium Titanyl Arsenate/CTA), Ba2NaNb5O15 (Barium Sodium Niobate/BNN), K3Li2Nb5O15 (Potassium Lithium Niobate/KLN), CO(NH2)2 (Urea), LiIO3 (Lithium Iodate), Ag3AsS3 (Proustite), HgGa2S4 (Mercury Thiogallate), CdGeAs2 (Cadmium Germanium Arsenide/CGA), Ti3AsSe3 (Thallium Arsenic Selenide/TAS), CdSe (Cadmium Selenide), ZnO (Zinc Oxide), ZnS (Zinc Sulfide), ZnSe (Zinc Selenide), ZnTe (Zinc Telluride), CdS (Cadmium Sulfide), SiC (Silicon Carbide), GaN (Gallium Nitride), GaSb (Gallium Antimonide).

4. The SHG nanoprobe system of claim 1, wherein each of the SHG nanoprobes may include magneto-SHG particles selected from the group consisting of ferromagnetic, ferrimagnetic, paramagnetic, diamagnetic, superparamagnetic (i.e. Fe3O4), superdiamagnetic, and metamagentic superparamagnetic material.

5. The SHG nanoprobe system of claim 1, wherein each of the SHG nanoprobes may further be surrounded by a resonant material nanoshell, the nanoshell being formed from a material selected from the group consisting of gold, silver, copper, aluminum, palladium, or platinum.

6. The SHG nanoprobe system of claim 1, wherein each of the SHG nanoprobes is less than or equal to 10 μ

m.

7. The SHG nanoprobe system of claim 1, wherein the excitation source is selected from the group consisting of continuous wave, modulated and pulsed lasers.

8. The SHG nanoprobe system of claim 1, further comprising at least one exciter nanostructure, said exciter nanostructure designed to produce an enhanced local electrical field of a specified frequency when exposed to the excitation source, such that each of the SHG nanoprobes generate a second harmonic resonance emission when brought within range of the resonant electrical field of said exciter nanostructure.

9. The SHG nanoprobe system of claim 8, wherein the at least one exciter nanostructure is a metal nanostructure.

10. The SHG nanoprobe system of claim 9, wherein the metal nanostructure is a nanostructure selected from the group consisting of nanorods, nanospheres or nanoshells.

11. The SHG nanoprobe system of claim 10, wherein the metal nanostructure is made of gold, silver, copper, aluminum, palladium, or platinum.

12. The SHG nanoprobe system of claim 8, wherein each of the SHG nanoprobe and the emitter nanostructure are both attached to a single nucleotide of interest.

13. The SHG nanoprobe system of claim 8, wherein the exciter nanostructure is linked to a polymerase molecule used in the nucleotide sequencing process.

14. The SHG nanoprobe system of claim 8, wherein the exciter nanostructure is disposed on a surface of a detector used for detecting the distinct emissive fingerprints.

15. The SHG nanoprobe system of claim 1, wherein the SHG nanoprobes are functionalized such that the SHG nanoprobe may be attached to the nucleotide of interest.

16. The SHG nanoprobe system of claim 1, wherein the nucleotides are selected from the group consisting of double-stranded DNA, single-stranded DNA, DNA from plasmid, first strand cDNA, total genomic DNA, RNA, cut/end-modified DNA, in vitro transposon tagged DNA.

17. The SHG nanoprobe system of claim 1, wherein the SHG nanoprobes are voltage sensitive SHG nanoprobes, and where the external excitation source is an electric field having sufficient emissive strength to generate a harmonic emission in the SHG nanoprobes.

18. A voltage sensitive SHG nanoprobe system comprising:

at least one SHG nanoprobe, said SHG nanoprobe having no inversion symmetry such that a unique second harmonic emission is generated when radiated by an external electric field; and

a probe target capable of generating an electric field, the electric field capable of radiating the SHG nanoprobe with an emissive field strength such that the SHG nanoprobe generates a second harmonic emission.

19. The SHG nanoprobe system of claim 18, wherein the SHG nanoprobe is a nanocrystal selected from the group consisting of organic, inorganic and combinations thereof.

20. The SHG nanoprobe system of claim 19, wherein the nanocrystal is selected from the group consisting of BaTiO₃, SiC, ZnO, ZnS, ZnSe, ZnTe, CdS, CdSe, CdTe, GaAs, GaSb, GaP, GaN, InSb, LiNbO3, KNbO3, KTiOPO₄, Fe(IO₃)₃, Au, Ag, N-(4-nitrophenyl)-(L)-prolinol (NPP), urea, 4-Nitroaniline, 2-Methyl-4-nitroaniline (MNA), 3-Methyl-4-methoxy-4′

-nitrostilbene), β

-BaB2O4 (Beta-Barium Borate/BBO, LiB3O5 (Lithium Triborate/LBO), LiNbO3 (Lithium Niobate/LN), KTiOPO4 (Potassium Titanyl Phosphate/KTP), AgGaS2 (Silver Thiogallate/AGS), AgGaSe2 (Silver Gallium Selenide/AGSe), ZnGeP2 (Zinc Germanium Phosphide/ZGP), GaSe (Gallium Selenide), KH2PO4 (Potassium Dihydrogen Phosphate/KDP), NH4H2PO4 (Ammonium Dihydrogen Phosphate (ADP), KD2PO4 (Deuterated Potassium Dihydrogen Phosphate/DKDP), CsLiB6O10 (Cesium Lithium Borate/CLBO), KTiOAsO4 (Potassium Titanyl Arsenate/KTA), KNbO3 (Potassium Niobate/KN), LiTaO3 (Lithium Tantalate/LT), RbTiOAsO4 (Rubidium Titanyl Arsenate/RTA), BaTiO₃ (Barium Titanate), MgBaF4 (Magnesium Barium Fluoride), GaAs (Gallium Arsenide), BiB3O6 (Bismuth Triborate/BIBO), K2Al2B2O7 (Potassium Aluminum Borate/KABO), KBe2BO3F2 (Potassium Fluoroboratoberyllate/KBBF), BaAlBO3F2 (Barium Aluminum Fluoroborate/BABF), La2CaB10O19 (Lanthanum Calcium Borate/LCB), GdCa4O (BO3)3 (Gadolinium Calcium Oxyborate/GdCOB), YCa4O (BO3)3 (Yttrium Calcium Oxyborate/YCOB), Li2B4O7 (Lithium Tetraborate/LB4), LiRbB4O7 (Lithium Rubidium Tetraborate/LRB4), CdHg(SCN)4 (Cadmium Mercury Thiocyanate/CMTC), RbTiOPO4 (Rubidium Titanyl Phosphate/RTP), LiInS2 (Lithium Thioindate/LIS), LiInSe2 (Lithium Indium Selenide/LISe), KB5O8.4H2O (Potassium Pentaborate Tetrahydrate/KB5), CsB3O5 (Cesium Triborate/CBO), C4H7D12N4PO7 (Deuterated L-Arginine Phosphate Monohydrate/DLAP), a-HIO3 (a-Iodic Acid), LiCOOH.H2O (Lithium Formate Monohydrate/LFM), CsH2AsO4 (Cesium Dihydrogen Arsenate/CDA), CsD2AsO4 (Deuterated Cesium Dihydrogen Arsenate/DCDA), RbH2PO4 (Rubidium Dihydrogen Phosphate/RDP), CsTiOAsO4 (Cesium Titanyl Arsenate/CTA), Ba2NaNb5O15 (Barium Sodium Niobate/BNN), K3Li2Nb5O15 (Potassium Lithium Niobate/KLN), CO(NH2)2 (Urea), LiIO3 (Lithium Iodate), Ag3AsS3 (Proustite), HgGa2S4 (Mercury Thiogallate), CdGeAs2 (Cadmium Germanium Arsenide/CGA), Ti3AsSe3 (Thallium Arsenic Selenide/TAS), CdSe (Cadmium Selenide), ZnO (Zinc Oxide), ZnS (Zinc Sulfide), ZnSe (Zinc Selenide), ZnTe (Zinc Telluride), CdS (Cadmium Sulfide), SiC (Silicon Carbide), GaN (Gallium Nitride), GaSb (Gallium Antimonide).

21. The SHG nanoprobe system of claim 18, wherein each of the SHG nanoprobes may include magneto-SHG particles selected from the group consisting of ferromagnetic, ferrimagnetic, paramagnetic, diamagnetic, superparamagnetic (i.e. Fe3O4), superdiamagnetic, and metamagentic superparamagnetic material.

22. The SHG nanoprobe system of claim 18, wherein each of the SHG nanoprobes may further be surrounded by a resonant material nanoshell, the nanoshell being formed from a material selected from the group consisting of gold, silver, copper, aluminum, palladium, or platinum. The SHG nanoprobe system of claim 18, wherein the SHG nanoprobe is less than or equal to 10 μ

m.

23. The SHG nanoprobe system of claim 18, further comprising at least one exciter nanostructure, said exciter nanostructure designed to produce an enhanced local electrical field of a specified frequency when exposed to the probe target electric field, such that the SHG nanoprobe generates a second harmonic resonance emission when brought within range of the resonant electrical field of said exciter nanostructure.

24. The SHG nanoprobe system of claim 23, wherein the at least one exciter nanostructure is a metal nanostructure.

25. The SHG nanoprobe system of claim 24, wherein the metal nanostructure is a nanostructure selected from the group consisting of nanorods, nanospheres or nanoshells.

26. The SHG nanoprobe system of claim 24, wherein the metal nanostructure is made of gold, silver, copper, aluminum, palladium, or platinum.

27. The SHG nanoprobe system of claim 23, wherein each of the SHG nanoprobe and the emitter nanostructure are attached to a single probe target of interest.

28. The SHG nanoprobe system of claim 18, wherein the electric field is a physiologically generated field.

29. A method of nucleic acid sequence comprising:

attaching at least one distinct SHG nanoprobe to each of a plurality of distinct nucleotide types of interest, wherein each of the SHG nanoprobes has no inversion symmetry, said distinct SHG nanoprobes being selected such that each generates a distinct second harmonic emission when radiated by an excitation source at at least two frequencies;

sequencing the nucleotides;

radiating the sequenced nucleotides with an external excitation source at at least two excitation frequencies; and

detecting the second harmonic emission from the SHG nanoprobe to obtain a distinct emissive fingerprint for each nucleotide.

30. The method of claim 29, wherein each of the SHG nanoprobes is a nanocrystal selected from the group consisting of organic, inorganic and combinations thereof.

31. The method of claim 29, wherein each of the SHG nanoprobes is one of either directly attached to or introduced into the distinct nucleotide.

32. The method of claim 29, wherein each of the SHG nanoprobes is conjugated to a probe molecule that is sensitive to the presence of the nucleotide of interest.

33. The method of claim 29, wherein the excitation source is selected from the group consisting of continuous wave, modulated and pulsed lasers.

34. The method of claim 29, further comprising interspersing an exciter nanostructure with the nucleotides, said exciter nanostructure designed to produce an enhanced electrical field of a specified frequency when exposed to an excitation source, wherein each of the SHG nanoprobes generates a distinct second harmonic resonance emission when brought within range of the enhanced electrical field of said exciter nanostructure.

35. The method of claim 34, wherein the exciter nanostructure is a metal nanostructure selected from the group consisting of nanorods, nanospheres or nanoshells.

36. The method of claim 34, wherein each of the SHG nanoprobe and the emitter nanostructure are attached to a single nucleotide of interest.

37. The method of claim 29, wherein the SHG nanoprobes are voltage sensitive SHG nanoprobes, and where the external excitation source is an electric field having sufficient emissive strength to generate a harmonic emission in the SHG nanoprobes.

38. A method of measuring an external electric field comprising:

dispersing at least one SHG nanoprobe in the vicinity of a target of interest interest, wherein the SHG nanoprobe has no inversion symmetry such that said SHG nanoprobe generates a second harmonic emission when radiated by an electric field, and wherein the target of interest is capable of generating an electric field of sufficient emissive strength to generate a second harmonic emission from the SHG nanoprobe, and;

exposing the SHG nanoprobe to the target generated electric field; and

monitoring the second harmonic emission from the SHG nanoprobe.

39. The method of claim 38, wherein the SHG nanoprobe is a nanocrystal selected from the group consisting of organic, inorganic and combinations thereof.

40. The method of claim 38, further comprising interspersing an exciter nanostructure with the SHG nanoprobe, said exciter nanostructure designed to produce an enhanced electrical field of a specified frequency when exposed to an excitation source, wherein the SHG nanoprobe generates a second harmonic resonance emission when brought within range of the enhanced electrical field of said exciter nanostructure.

41. The method of claim 40, wherein the exciter nanostructure is a metal nanostructure selected from the group consisting of nanorods, nanospheres or nanoshells.

42. The method of claim 40, wherein each of the SHG nanoprobe and the emitter nanostructure are attached to a single target of interest.

43. The method of claim 38, wherein the electric field is a physiologically generated field.