NANOCOMPOSITES AND METHODS OF MAKING SAME

• US 20160077090A1
• Filed: 04/10/2015
• Published: 03/17/2016
• Est. Priority Date: 10/15/2013
• Status: Active Grant

• Current Assignee: The Board of Trustees of the University of Arkansas (University of Arkansas System)
• Original Assignee: The Board of Trustees of the University of Arkansas (University of Arkansas System)
• Inventors: Biris, Alexandru S., Nima, Zeid, Kumpuris, Andrew G.
• Granted Patent: US 9,810,687 B2
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: G01N 21/658 enhancement Raman, e.g. surface plasmons
• CPC Class Codes: G01N 2800/28 Neurological disorders
• CPC Class Codes: G01N 33/54346 Nanoparticles

Abstract

A nanoagent includes at least one nanocomposite. The nanocomposite includes at least one gold nanorod, a silver layer coated on an outer surface of the gold nanorod, a Raman reporter molecule layer coated on the silver layer, a pegylated layer coated on the Raman reporter molecule layer, an active layer conjugated to the pegylated layer. the active layer includes at least one of a targeting molecule configured to bind to a target of interest, and a functional molecule configured to interact with the target of interest. The silver layer has silver nanoparticles. The Raman reporter molecule layer has Raman reporter molecules that are detectable by surface enhanced Raman spectroscopy (SERS). The pegylated layer has at least one of thiolated polyethylene glycol (HS-PEG), thiolated polyethylene glycol acid (HS-PEG-COOH) and HS-PEG-NHx.

Claims

1. A nanoagent comprising at least one nanocomposite, the nanocomposite comprising:

at least one gold nanorod;

a silver layer coated on an outer surface of the gold nanorod, the silver layer comprising silver nanoparticles;

a Raman reporter molecule layer coated on the silver layer, wherein the Raman reporter molecule layer comprises the Raman reporter molecules that are detectable by surface enhanced Raman spectroscopy (SERS);

a pegylated layer coated on the Raman reporter molecule layer, comprising at least one of thiolated polyethylene glycol (HS-PEG), thiolated polyethylene glycol acid (HS-PEG-COOH) and HS-PEG-NHx; and

an active layer conjugated to the pegylated layer, the active layer comprising at least one of;

a targeting molecule configured to bind to a target of interest; and

a functional molecule configured to interact with the target of interest.

2. The nanoagent of claim 1, wherein the target of interest is at least one of a cancer cell, a pathogen, or a plant.

3. The nanoagent of claim 1, wherein the targeting molecule comprises an antibody specifically binds to the target of interest.

4. The nanoagent of claim 1, wherein the functional molecule is a virus, a phage, a drug, a growth factor, antibiotics, a gene, a plasmid, a vaccine, a plant growth agent, an anti-fungal, a fertilizer, herbicides, an antibody that specifically binds to S100 calcium-binding protein B (S100B), or other biological active molecules.

5. The nanoagent of claim 1, wherein the active layer comprises a biomarker of traumatic brain injury (TBI) or an antibody that specifically binds to the biomarker.

6. The nanoagent of claim 5, wherein the biomarker of TBI comprises at least one of S100 calcium-binding protein B (S100B), neuron-specific enolase (NSE), myelin basic protein (MBP), caspase-3, interleukins, tau protein, neurofilament light polypeptide (NEFL), neurofilament heavy polypeptide (NEFH), glial fibrillary acidic protein, amyloid precursor protein (APP), and amyloid.

7. The nanoagent of claim 1,

wherein the gold nanorod has an aspect ratio (AR) in a range of about 1-9, a length in a range of about 10-100 nm, and a diameter in a range of about 1-40 nm;

and

wherein the silver layer has a thickness in a range of about 0.5-5 nm.

8. The nanoagent of claim 1, wherein the Raman reporter molecule layer comprises 4-mercaptobenzoic acid (4MBA), p-aminothiophenol (PATP), p-nitrothiophenol (PNTP), 4-(methylsulfanyl)thiophenol (4MSTP), or other molecules with unique Raman spectra and intense Raman peak intensities.

9. The nanoagent of claim 1, wherein the HS-PEG has a molecular weight in a range of about 1.5-15 kilo Dalton (kD) and the HS-PEG-COOH has a molecular weight in a range of about 1-10 kD.

10. The nanoagent of claim 1, wherein at least one of the targeting molecule and the functional molecule is conjugated to the pegylated layer through the carboxylic group of the HS-PEG-COOH or amine group of the HS-PEG-NHx.

11. The nanoagent of claim 1, wherein the targeting molecule comprises anti-epithelial cell adhesion molecule antibody (anti-EpCAM), anti-CD44 antibody, anti-insulin-like growth factor 1 receptor antibody (anti-IGF-1), anti-Keratin 18 antibody, or one or more antibodies specific to the target of interest.

12. The nanoagent of claim 11,

wherein the at least one nanocomposite comprises a first nanocomposite, a second nanocomposite, a third nanocomposite, and a fourth nanocomposite;

wherein the Raman reporter molecule layer of the first nanocomposite comprises 4-mercaptobenzoic acid (4MBA), and the antibody of the first nanocomposite is anti-epithelial cell adhesion molecule antibody (anti-EpCAM);

wherein the Raman reporter molecule layer of the second nanocomposite comprises p-aminothiophenol (PATP), and the antibody of the second nanocomposite is anti-CD44 antibody;

wherein the Raman reporter molecule layer of the third nanocomposite comprises p-nitrothiophenol (PNTP), and the antibody of the third nanocomposite is anti-insulin-like growth factor 1 receptor antibody (anti-IGF-1);

wherein the Raman reporter molecule layer of the fourth nanocomposite comprises 4-(methylsulfanyl)thiophenol (4MSTP), and the antibody of the fourth nanocomposite is anti-Keratin 18 antibody; and

wherein SERS signal corresponding to each of the first, second, third and fourth nanocomposites is represented by a predetermined color.

13. A system for detecting a target of interest and monitoring conditions of the target of interest, comprising:

a nanoagent of claim 1;

a surface enhanced Raman spectrometer configured to provide an incident radiation signal to the target of interest, and to collect SERS signals generated by the Raman reporter molecule layer in response to the incident radiation signal; and

a processing unit for processing the SERS signals collected by the surface enhanced Raman spectrometer, so as to detect and monitor the conditions of the target of interest.

14. A method of making a nanocomposite, comprising:

forming at least one gold nanorod;

coating a silver layer on an outer surface of the gold nanorod;

assembling a Raman reporter molecule layer on the coated silver layer;

coating a pegylated layer on the assembled Raman reporter molecule layer;

conjugating the coated pegylated layer with an active layer, the active layer comprising at least one of a targeting molecule configured to bind to the target of interest and a functional molecule configured to interact with the target of interest.

15. The method of claim 14, wherein the step of forming the at least one gold nanorod comprises:

mixing a first exadecyltrimethylammoniumbromide (CTAB) solution with a silver nitrate solution to form a first mixture;

adding a first HAuCl₄ to the first mixture to form a second mixture;

adding a first ascorbic acid to the second mixture to form a third mixture;

adding a seed solution to the third mixture to form a fourth mixture; and

centrifuging the fourth mixture to form a first precipitate, wherein the first precipitate comprises the gold nanorod.

16. The method of claim 15, wherein the seed solution is prepared by:

mixing a second CTAB solution with a second HAuCl₄ to form a fifth mixture; and

adding NaBH₄ to the fifth mixture and stirring to form the seed solution.

17. The method of claim 16, wherein the step of coating the silver layer comprises:

dispersing the gold nanorod in a third CTAB solution by sonication to form a sixth mixture;

adding a polyvinylpyrrolidone (PVP) solution and AgNO₃ to the sixth mixture and gently mixing to form a seventh mixture;

adding a second ascorbic acid to the seventh mixture to form an eighth mixture;

adding NaOH solution to the eighth mixture to form a ninth mixture, such that the pH of the ninth mixture is elevated to about pH 9, and a silver ion reduction reaction is initiated; and

centrifuging the ninth mixture to form a second precipitate, wherein the second precipitate comprises the gold nanorod coated with the silver layer.

18. The method of claim 17, wherein the step of assembling the Raman reporter molecule layer comprises:

dispersing the gold nanorod coated with the silver layer in distilled water to form a tenth mixture;

dissolving the Raman reporter molecule comprising 4-mercaptobenzoic acid (4-MBA), p-aminothiophenol (PATP), p-nitrothiophenol (PNTP), or 4-(methylsulfanyl)thiophenol (4-MSTP), in ethanol to form a reporter solution;

adding the reporter solution to the tenth mixture and stirring to form an eleventh mixture; and

centrifuging the eleventh mixture to form a third precipitate, wherein the third precipitate comprises the gold nanorod coated with the silver layer, and assembled with the Raman report molecule layer.

19. The method of claim 18, wherein the step of coating the pegylated layer comprises:

dispersing the gold nanorod with the coated silver layer and the assembled Raman reporter molecule layer in thiolated polyethylene glycol acid (HS-PEG-COOH) solution and vigorously stirring to form a twelfth mixture, wherein the HS-PEG-COOH solution comprises about 2 mg/ml HS-PEG and about 2 mM NaCl;

adding thiolated polyethylene glycol (HS-PEG) to the twelfth mixture and keep at about 5°

C. overnight to form a thirteenth mixture; and

centrifuging the thirteenth mixture to form a fourth precipitate, wherein the fourth precipitate comprises the gold nanorod coated with the silver layer, assembled with the Raman report molecule layer, and coated with the pegylated layer.

20. The method of claim 19, wherein the step of conjugating the coated pegylated layer with the targeting molecule comprises:

suspending the gold nanorod with the coated silver layer, the assembled Raman reporter molecule layer, and the coated with pegylated layer in phosphate-buffered saline (PBS) by sonicating to form a suspending mixture;

adding N-hydroxysuccinimide (NHS) and 1N-ethyl-N′

-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) to the suspending mixture and stirring to form a fourteenth mixture;

washing the fourteenth mixture by centrifuging twice using PBS to obtain a fifth precipitate;

dispending the fifth precipitate in PBS to form a fifteen mixture;

adding molecules of the targeting molecule to the fifteenth mixture and mixing thoroughly to form a sixteen mixture, wherein the targeting molecule comprises anti-EpCAM, anti-CD44, anti-IGF-1 Receptor β

, anti-Keratin 18, or one or more antibodies specific to the target of interest; and

stirring the sixteenth mixture at room temperature to form the nanocomposite.

21. A nanocomposite, comprising:

a nanostructure formed by at least one nanomaterial; and

an active layer conjugated to the nanostructure, and comprising at least one of;

a targeting molecule configured to bind to a target of interest; and

a functional molecule configured to interact with a target of interest.

22. The nanocomposite of claim 21, wherein the nanostructure has a spherical shape, a tubular shape, a cylindrical shape, or a rod-like triangular shape.

23. The nanocomposite of claim 21, wherein the nanomaterial comprises at least one of silver coated gold rods, quantum dots, nanowires, nanotubes, nanofibers, and fullerenes.

24. The nanocomposite of claim 21, wherein the nanomaterial comprises gold, silver, gold/silver, copper, iron, Fe_xO_y, TiO₂, SiO₂, and carbon.

25. The nanocomposite of claim 21, wherein the target of interest is at least one of a cancer cell, a pathogen, or a plant, and the functional molecule is a virus, a phage, a drug, a growth factor, antibiotics, a gene, a plasmid, a vaccine, a plant growth agent, an anti-fungal, a fertilizer, herbicides, an antibody that specifically binds to S100 calcium-binding protein B (S100B), or other biological active molecules.

26. The nanocomposite of claim 25, wherein the target of interest is the cancer cell, and the functional molecule is the virus capable of reproducing in the cancer cell.

27. The nanocomposite of claim 25, wherein the target of interest is the plant, and the functional molecule is the growth factor capable of promoting growth of the plant.

28. The nanocomposite of claim 21, wherein the active layer comprises a biomarker of traumatic brain injury (TBI) or an antibody that specifically binds to the biomarker.

29. The nanocomposite of claim 28, wherein the biomarker of TBI comprises at least one of S100 calcium-binding protein B (S100B), neuron-specific enolase (NSE), myelin basic protein (MBP) caspase-3, interleukins, tau protein, neurofilament light polypeptide (NEFL), neurofilament heavy polypeptide (NEFH), glial fibrillary acidic protein, amyloid precursor protein (APP), and amyloid.

30. The nanocomposite of claim 21, wherein the nanostructure comprises:

at least one gold nanorod; and

a silver layer surrounding the at least one gold nanorod, the silver layer comprising silver nanoparticles.

31. The nanocomposite of claim 30,

wherein the gold nanorod has an aspect ratio (AR) in a range of about 1-9, a length in a range of about 10-100 nm, and a diameter in a range of about 1-40 nm;

and

wherein the silver layer has a thickness in a range of about 0.5-5 nm.

32. The nanocomposite of claim 21, further comprising a reporter layer disposed between the nanomaterial and the active layer, wherein the reporter layer is detectable by at least one of surface enhanced Raman spectroscopy (SERS), magnetic resonance imaging (MRI), x-ray radiography, computed tomography (CT), and infrared spectroscopy (IR).

33. The nanocomposite of claim 32, wherein the reporter layer comprises 4-mercaptobenzoic acid (4MBA), p-aminothiophenol (PATP), p-nitrothiophenol (PNTP), 4-(methylsulfanyl)thiophenol (4MSTP), or other molecules with unique Raman spectra and intense Raman peak intensities.

34. The nanocomposite of claim 32, further comprising a pegylated layer disposed between the reporter layer and the active layer, wherein the pegylated layer comprises at least one of thiolated polyethylene glycol (HS-PEG), thiolated polyethylene glycol acid (HS-PEG-COOH) and HS-PEG-NHx.

35. The nanocomposite of claim 21, wherein the targeting molecule is molecule of anti-epithelial cell adhesion molecule antibody (anti-EpCAM), anti-CD44 antibody, anti-insulin-like growth factor 1 receptor antibody (anti-IGF-1), anti-Keratin 18 antibody, or one or more antibodies specific to the target of interest.

36. The nanocomposite of claim 35, wherein the functional molecule is conjugated to at least one of the pegylated layer and the targeting molecule.