CN112957483A - Preparation method of intestinal targeted photoacoustic imaging contrast agent and product thereof - Google Patents
Preparation method of intestinal targeted photoacoustic imaging contrast agent and product thereof Download PDFInfo
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- 238000003384 imaging method Methods 0.000 title claims abstract description 43
- 239000002872 contrast media Substances 0.000 title claims abstract description 27
- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 230000000968 intestinal effect Effects 0.000 title claims description 16
- 239000002077 nanosphere Substances 0.000 claims abstract description 36
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 claims abstract description 31
- 210000001035 gastrointestinal tract Anatomy 0.000 claims abstract description 23
- 230000008685 targeting Effects 0.000 claims abstract description 18
- 238000000034 method Methods 0.000 claims abstract description 13
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 27
- 239000008367 deionised water Substances 0.000 claims description 25
- 229910021641 deionized water Inorganic materials 0.000 claims description 25
- FPQQSJJWHUJYPU-UHFFFAOYSA-N 3-(dimethylamino)propyliminomethylidene-ethylazanium;chloride Chemical compound Cl.CCN=C=NCCCN(C)C FPQQSJJWHUJYPU-UHFFFAOYSA-N 0.000 claims description 16
- 229920001661 Chitosan Polymers 0.000 claims description 15
- 238000006243 chemical reaction Methods 0.000 claims description 13
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 12
- 229960001484 edetic acid Drugs 0.000 claims description 12
- NQTADLQHYWFPDB-UHFFFAOYSA-N N-Hydroxysuccinimide Chemical compound ON1C(=O)CCC1=O NQTADLQHYWFPDB-UHFFFAOYSA-N 0.000 claims description 11
- RQFQJYYMBWVMQG-IXDPLRRUSA-N chitotriose Chemical compound O[C@@H]1[C@@H](N)[C@H](O)O[C@H](CO)[C@H]1O[C@H]1[C@H](N)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CO)O2)N)[C@@H](CO)O1 RQFQJYYMBWVMQG-IXDPLRRUSA-N 0.000 claims description 10
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 9
- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 claims description 9
- 238000004132 cross linking Methods 0.000 claims description 9
- 238000003756 stirring Methods 0.000 claims description 9
- 238000001132 ultrasonic dispersion Methods 0.000 claims description 9
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 claims description 8
- 238000003760 magnetic stirring Methods 0.000 claims description 8
- 239000002245 particle Substances 0.000 claims description 8
- 238000005406 washing Methods 0.000 claims description 8
- 102100024954 5-hydroxytryptamine receptor 3A Human genes 0.000 claims description 6
- 101710138027 5-hydroxytryptamine receptor 3A Proteins 0.000 claims description 6
- 102100032509 Histamine H1 receptor Human genes 0.000 claims description 6
- 101001016841 Homo sapiens Histamine H1 receptor Proteins 0.000 claims description 6
- 108010073929 Vascular Endothelial Growth Factor A Proteins 0.000 claims description 6
- 102000005789 Vascular Endothelial Growth Factors Human genes 0.000 claims description 6
- 108010019530 Vascular Endothelial Growth Factors Proteins 0.000 claims description 6
- 102000004142 Trypsin Human genes 0.000 claims description 2
- 108090000631 Trypsin Proteins 0.000 claims description 2
- 102000001400 Tryptase Human genes 0.000 claims description 2
- 108060005989 Tryptase Proteins 0.000 claims description 2
- 239000012588 trypsin Substances 0.000 claims description 2
- 241000792859 Enema Species 0.000 abstract description 3
- 239000007920 enema Substances 0.000 abstract description 3
- 229940095399 enema Drugs 0.000 abstract description 3
- 210000004347 intestinal mucosa Anatomy 0.000 abstract description 2
- 206010009944 Colon cancer Diseases 0.000 description 4
- 208000001333 Colorectal Neoplasms Diseases 0.000 description 4
- 230000031700 light absorption Effects 0.000 description 3
- 239000002086 nanomaterial Substances 0.000 description 3
- 241000699670 Mus sp. Species 0.000 description 2
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- 238000011161 development Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 239000010931 gold Substances 0.000 description 2
- 229910052737 gold Inorganic materials 0.000 description 2
- 239000002105 nanoparticle Substances 0.000 description 2
- 231100000252 nontoxic Toxicity 0.000 description 2
- 230000003000 nontoxic effect Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 206010033128 Ovarian cancer Diseases 0.000 description 1
- 206010061535 Ovarian neoplasm Diseases 0.000 description 1
- 238000003332 Raman imaging Methods 0.000 description 1
- 244000081822 Uncaria gambir Species 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 238000004140 cleaning Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000002059 diagnostic imaging Methods 0.000 description 1
- 238000012631 diagnostic technique Methods 0.000 description 1
- 238000012377 drug delivery Methods 0.000 description 1
- 230000002496 gastric effect Effects 0.000 description 1
- 230000030136 gastric emptying Effects 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- DIHKMUNUGQVFES-UHFFFAOYSA-N n,n,n',n'-tetraethylethane-1,2-diamine Chemical compound CCN(CC)CCN(CC)CC DIHKMUNUGQVFES-UHFFFAOYSA-N 0.000 description 1
- 239000002405 nuclear magnetic resonance imaging agent Substances 0.000 description 1
- 201000008106 ocular cancer Diseases 0.000 description 1
- 230000010494 opalescence Effects 0.000 description 1
- 238000012634 optical imaging Methods 0.000 description 1
- 238000007626 photothermal therapy Methods 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 239000000523 sample Substances 0.000 description 1
- 238000007920 subcutaneous administration Methods 0.000 description 1
- -1 tetraethyl ethylenediamine oxalate Chemical compound 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/22—Echographic preparations; Ultrasound imaging preparations ; Optoacoustic imaging preparations
- A61K49/221—Echographic preparations; Ultrasound imaging preparations ; Optoacoustic imaging preparations characterised by the targeting agent or modifying agent linked to the acoustically-active agent
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/22—Echographic preparations; Ultrasound imaging preparations ; Optoacoustic imaging preparations
- A61K49/222—Echographic preparations; Ultrasound imaging preparations ; Optoacoustic imaging preparations characterised by a special physical form, e.g. emulsions, liposomes
- A61K49/225—Microparticles, microcapsules
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- Health & Medical Sciences (AREA)
- Physics & Mathematics (AREA)
- Acoustics & Sound (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Radiology & Medical Imaging (AREA)
- Epidemiology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
Abstract
The invention discloses a preparation method of a photoacoustic imaging contrast agent targeting an intestinal tract and a product thereof. The product obtained by the method can adopt enema, the nanospheres are attached to the surface of intestinal mucosa by the antigen-antibody combination principle, and meanwhile, the gold nanorods wrapped in the nanospheres can obviously improve the intensity of photoacoustic signals under the irradiation of near-infrared laser, so that targeted photoacoustic imaging of intestinal tracts is realized.
Description
Technical Field
The invention relates to the field of preparation of nano materials, in particular to a preparation method of a photoacoustic imaging contrast agent targeting an intestinal tract and a product thereof.
Background
Colorectal cancer (CRC) is the fourth most common cancer in men and the third most common cancer in women worldwide. Due to the high morbidity and mortality of CRC, the development of diagnostic techniques for CRC is of paramount importance. Some traditional medical imaging technologies are difficult to realize imaging on a molecular level, and with the increasing requirements on diagnostic accuracy, development of molecular imaging probes becomes a research hotspot in recent years. Photoacoustic imaging, an emerging imaging technology, combines the advantages of optical imaging and ultrasonic imaging, and has both high contrast and high spatial resolution, and is considered to be an imaging method with great potential.
The chitosan is a natural polysaccharide, has good biocompatibility and degradability, has no biotoxicity, is rich in amino and hydroxyl, and is easy to modify. Therefore, chitosan nanomaterials are widely used in the field of nano drug delivery, for example, Cheng J., Zhou x., Zhou w., Wu h., Zhang x., Lu q., a Novel Magnetic Contrast Agent for gastric Imaging Through Oral administration J Biomed nanotechnol., 2019,15(6),1162-71, which reports a Novel nuclear Magnetic resonance Contrast Agent that achieves multifunctional assessment of gastric emptying by loading Gd in chitosan nanoparticles and Oral administration.
The gold nanorod is a representative gold nanomaterial, has the light absorption characteristic of high intensity in a near infrared region, and is a research hotspot in the fields of photoacoustic imaging, photothermal therapy and the like. For example, Jokerst J., Cole J., Van de Sompel D., Gambir S.S., Gold nanoparticles for ocular cancer detection with photoacoustic imaging and restriction analysis via Raman imaging in living mice, ACS nano., 6(11), 10366-77, which prepares Gold nanorods of different aspect ratios, enabling photoacoustic imaging of subcutaneous ovarian cancer in mice.
Therefore, the gold nanorods are loaded in the chitosan nanospheres, and the targeted antibodies are used for modifying the surfaces of the nanospheres to obtain the intestinal targeted photoacoustic imaging contrast agent. The targeted photoacoustic imaging of the intestinal tract is realized by utilizing the safe and nontoxic carrier characteristic of the chitosan nanosphere, the near infrared light absorption characteristic of the gold nanorod and the active targeting characteristic of the antibody.
Disclosure of Invention
Aiming at the defects in the prior art, the invention aims to provide a preparation method of a photoacoustic imaging contrast agent for targeting an intestinal tract.
Yet another object of the present invention is to: an intestinal tract targeted photoacoustic imaging contrast agent product prepared by the method is provided.
The purpose of the invention is realized by the following scheme: a preparation method of a targeted intestinal photoacoustic imaging contrast agent comprises the steps of coating a gold nanorod in a chitosan nanosphere, coupling a targeted antibody to the surface of the nanosphere through EDC/NHS reaction to obtain the intestinal targeted photoacoustic imaging contrast agent, and at least comprises the following preparation steps:
a. dissolving 30 mg of chitosan oligosaccharide and 17mg of Ethylene Diamine Tetraacetic Acid (EDTA) in 10 mL of deionized water, adding 2-5 mg of gold nanorods, and performing ultrasonic dispersion uniformly;
b. dropwise adding absolute ethyl alcohol under magnetic stirring until the system is discolored, adding 30 mu L of 25% glutaraldehyde solution for crosslinking for 4 h, and centrifugally collecting nanospheres to obtain the chitosan nanospheres coated with gold nanorod shells;
c. dissolving 100 μ L of the target antibody in 10 mL of deionized water, adding 12 mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 8 mg of N-hydroxysuccinimide (NHS) to activate the carboxyl group of the antibody at room temperature for 2-4 h;
d. and (c) adding 10-15 mg of the nanospheres obtained in the step (b) into the reaction system, stirring for 4 hours, centrifuging and collecting, and washing with deionized water for three times to remove unreacted antibodies to obtain a product.
The diameter of the gold nanorod is 40-100 nm.
The length-diameter ratio of the gold nanorods is 2-5.
The target Antibody is one of a 5-hydroxytryptamine receptor 3 Antibody (Anti-5-HT3R), a vascular endothelial growth factor Antibody (Anti-VEGF), a histamine receptor H1 Antibody (Anti-HRH 1) and a Tryptase Polyclonal Antibody (Polyclonal Antibody to trypsin).
The invention provides a photoacoustic imaging contrast agent for targeting an intestinal tract, which is prepared according to any one of the methods, wherein the average particle size of the obtained product is 173-228 nm, under the irradiation of laser with 808nm, the temperature of 200 mu g/mL product is raised to 45-52 ℃ within 1 min, and the targeting antibodies are targeting intestinal tract 5-hydroxytryptamine receptor 3, targeting intestinal tract histamine receptor H1 and targeting vascular endothelial growth factor.
Respectively weighing chitosan oligosaccharide and tetraethyl ethylenediamine, adding the chitosan oligosaccharide and the tetraethyl ethylenediamine oxalate into deionized water, fully stirring and dissolving, then adding a certain amount of gold nanorods into the solution, performing ultrasonic dispersion, then dropwise adding absolute ethyl alcohol until the reaction system becomes a opalescence, and adding a glutaraldehyde solution for crosslinking. And (3) centrifuging and cleaning the product, and then connecting the product with a targeting antibody to obtain the photoacoustic imaging contrast agent for targeting the intestinal tract, so as to realize intestinal tract targeting photoacoustic imaging.
The photoacoustic imaging contrast agent with the target intestinal tract, which is formed by adopting the method, has the advantages of simple method, good water dispersibility of the obtained product, high imaging contrast ratio and the like. The using method of the product is enema, the nanospheres are attached to the surface of intestinal mucosa by the antigen-antibody combination principle, and meanwhile, the gold nanorods wrapped in the nanospheres can obviously improve the strength of photoacoustic signals under the irradiation of near-infrared laser, so that targeted photoacoustic imaging of intestinal tracts is realized. The targeted photoacoustic imaging of the intestinal tract is realized by utilizing the safe and nontoxic carrier characteristic of the chitosan nanosphere, the near infrared light absorption characteristic of the gold nanorod and the active targeting characteristic of the antibody.
Compared with the prior art, the invention has the following advantages:
(1) the preparation method is simple, the reaction is easy to control, the stability is good, and the application prospect is wide.
(2) The obtained product has excellent water dispersibility and biocompatibility and high imaging contrast.
(3) The use mode is an enema mode, the operation is easy, and the toxicity to organisms is low.
Detailed Description
The present invention will be described in detail with reference to specific examples. The following examples will assist those skilled in the art in further understanding the invention, but are not intended to limit the invention in any way. It should be noted that variations and modifications can be made by persons skilled in the art without departing from the spirit of the invention. All falling within the scope of the present invention.
Example 1
A targeted intestinal photoacoustic imaging contrast agent is prepared by coating a gold nanorod in a chitosan nanosphere, coupling a targeted antibody to the surface of the nanosphere through EDC/NHS reaction to obtain the intestinal targeted photoacoustic imaging contrast agent and preparing the intestinal targeted photoacoustic imaging contrast agent according to the following preparation steps:
a. dissolving 30 mg of chitosan oligosaccharide and 17mg of EDTA in 10 mL of deionized water, adding 2 mg of gold nanorods, and performing ultrasonic dispersion uniformly;
b. dropwise adding absolute ethyl alcohol under magnetic stirring until the color of the system changes into a milky color, adding 30 mu L of 25% glutaraldehyde solution for crosslinking for 4 h, and centrifugally collecting nanospheres to obtain the chitosan nanospheres coated with gold nanorod shells;
c. dissolving 100 μ L of Anti-5-HT3R in 10 mL deionized water, adding 12 mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 8 mg of N-hydroxysuccinimide (NHS) to activate the carboxyl group of the antibody at room temperature for 2-4 h;
d. and (c) adding 10 mg of the nanospheres obtained in the step (b) into the reaction system, stirring for 4 hours, centrifuging and collecting, and washing with deionized water for three times to remove unreacted antibodies to obtain a product.
The average particle size of the product is 228 nm, and under the irradiation of laser with 808nm, 200 mug/mL of the product is heated to 45 ℃ within 1 min, and the product is targeted to the intestinal tract 5-hydroxytryptamine receptor 3.
Example 2:
an intestinal tract targeted photoacoustic imaging contrast agent prepared by the following steps, similar to the steps of the example:
a. dissolving 30 mg of chitosan oligosaccharide and 17mg of Ethylene Diamine Tetraacetic Acid (EDTA) in 10 mL of deionized water, adding 5 mg of gold nanorods, and performing ultrasonic dispersion uniformly;
b. dropwise adding absolute ethyl alcohol under magnetic stirring until the color of the system changes into a milky color, adding 30 mu L of 25% glutaraldehyde solution for crosslinking for 4 h, and centrifugally collecting nanospheres to obtain the chitosan nanospheres coated with gold nanorod shells;
c. dissolving 100 μ L of Anti-HRH1 in 10 mL of deionized water, adding 12 mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 8 mg of N-hydroxysuccinimide (NHS) to activate the carboxyl group of the antibody at room temperature for 2-4 h;
d. and (c) adding 10 mg of the nanospheres obtained in the step (b) into the reaction system, stirring for 4 hours, centrifuging and collecting, and washing with deionized water for three times to remove unreacted antibodies to obtain a product.
The average particle size of the product is 173 nm, and under the irradiation of laser light with 808nm, 200 μ g/mL of the product is heated to 48 ℃ within 1 min, and the product is targeted to the intestinal histamine receptor H1.
Example 3:
an intestinal tract targeted photoacoustic imaging contrast agent prepared by the following steps, similar to the steps of the example:
a. dissolving 30 mg of chitosan oligosaccharide and 17mg of EDTA in 10 mL of deionized water, adding 10 mg of gold nanorods, and performing ultrasonic dispersion uniformly;
b. dropwise adding absolute ethyl alcohol under magnetic stirring until the color of the system changes into a milky color, adding 30 mu L of 25% glutaraldehyde solution for crosslinking for 4 h, and centrifugally collecting nanospheres;
c. dissolving 100 μ L of Anti-VEGF in 10 mL of deionized water, adding 12 mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 8 mg of N-hydroxysuccinimide (NHS) to activate the antibody carboxyl for 2-4 h at room temperature;
d. adding 15 mg of nanosphere into the reaction system, stirring for 4 h, centrifuging and collecting, and washing with deionized water for three times to remove unreacted antibody to obtain the product.
The average particle size of the product is 215 nm, and under the irradiation of laser of 808nm, the temperature of 200 mug/mL product is raised to 52 ℃ within 1 min, and the product targets the vascular endothelial growth factor.
Claims (8)
1. A preparation method of a photoacoustic imaging contrast agent targeting an intestinal tract is characterized in that gold nanorods are coated in chitosan nanospheres, and then a targeting antibody is coupled to the surfaces of the nanospheres through EDC/NHS reaction to obtain the photoacoustic imaging contrast agent targeting the intestinal tract, and at least comprises the following preparation steps:
a. dissolving 30 mg of chitosan oligosaccharide and 17mg of Ethylene Diamine Tetraacetic Acid (EDTA) in 10 mL of deionized water, adding 2-5 mg of gold nanorods, and performing ultrasonic dispersion uniformly;
b. dropwise adding absolute ethyl alcohol under magnetic stirring until the system is discolored, adding 30 mu L of 25% glutaraldehyde solution for crosslinking for 4 h, and centrifugally collecting nanospheres to obtain the chitosan nanospheres coated with gold nanorod shells;
c. dissolving 100 μ L of the target antibody in 10 mL of deionized water, adding 12 mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 8 mg of N-hydroxysuccinimide (NHS) to activate the carboxyl group of the antibody at room temperature for 2-4 h;
d. and (c) adding 10-15 mg of the nanospheres obtained in the step (b) into the reaction system, stirring for 4 hours, centrifuging and collecting, and washing with deionized water for three times to remove unreacted antibodies to obtain a product.
2. The method for preparing a photoacoustic imaging contrast agent for intestinal targeting according to claim 1, wherein the diameter of the gold nanorods is 40-100 nm.
3. The method for preparing the intestinal tract-targeted photoacoustic imaging contrast agent according to claim 1, wherein the length-to-diameter ratio of the gold nanorods is 2 to 5.
4. The method for preparing the intestinal tract-targeted photoacoustic imaging contrast agent according to claim 1, wherein the targeting Antibody is one of 5-hydroxytryptamine receptor 3 Antibody (Anti-5-HT3R), vascular endothelial growth factor Antibody (Anti-VEGF), histamine receptor H1 Antibody (Anti-HRH 1), and Tryptase Polyclonal Antibody (Polyclonal Antibody to trypsin).
5. The method for preparing the intestinal tract targeted photoacoustic imaging contrast agent according to claims 1 to 4, comprising the following steps:
a. dissolving 30 mg of chitosan oligosaccharide and 17mg of EDTA in 10 mL of deionized water, adding 2 mg of gold nanorods, and performing ultrasonic dispersion uniformly;
b. dropwise adding absolute ethyl alcohol under magnetic stirring until the color of the system changes into a milky color, adding 30 mu L of 25% glutaraldehyde solution for crosslinking for 4 h, and centrifugally collecting nanospheres to obtain the chitosan nanospheres coated with gold nanorod shells;
c. dissolving 100 μ L of Anti-5-HT3R in 10 mL deionized water, adding 12 mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 8 mg of N-hydroxysuccinimide (NHS) to activate the carboxyl group of the antibody at room temperature for 2-4 h;
d. and (c) adding 10 mg of the nanospheres obtained in the step (b) into the reaction system, stirring for 4 hours, centrifuging and collecting, and washing with deionized water for three times to remove unreacted antibodies to obtain a product.
The average particle size of the product is 228 nm, and under the irradiation of laser with 808nm, 200 mug/mL of the product is heated to 45 ℃ within 1 min, and the product is targeted to the intestinal tract 5-hydroxytryptamine receptor 3.
6. The method for preparing the intestinal tract targeted photoacoustic imaging contrast agent according to claims 1 to 4, comprising the following steps:
a. dissolving 30 mg of chitosan oligosaccharide and 17mg of Ethylene Diamine Tetraacetic Acid (EDTA) in 10 mL of deionized water, adding 5 mg of gold nanorods, and performing ultrasonic dispersion uniformly;
b. dropwise adding absolute ethyl alcohol under magnetic stirring until the color of the system changes into a milky color, adding 30 mu L of 25% glutaraldehyde solution for crosslinking for 4 h, and centrifugally collecting nanospheres to obtain the chitosan nanospheres coated with gold nanorod shells;
c. dissolving 100 μ L of Anti-HRH1 in 10 mL of deionized water, adding 12 mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 8 mg of N-hydroxysuccinimide (NHS) to activate the carboxyl group of the antibody at room temperature for 2-4 h;
d. and (c) adding 10 mg of the nanospheres obtained in the step (b) into the reaction system, stirring for 4 hours, centrifuging and collecting, and washing with deionized water for three times to remove unreacted antibodies to obtain a product.
The average particle size of the product is 173 nm, and under the irradiation of laser light with 808nm, 200 μ g/mL of the product is heated to 48 ℃ within 1 min, and the product is targeted to the intestinal histamine receptor H1.
7. The method for preparing the intestinal tract targeted photoacoustic imaging contrast agent according to claims 1 to 4, comprising the following steps:
a. dissolving 30 mg of chitosan oligosaccharide and 17mg of EDTA in 10 mL of deionized water, adding 10 mg of gold nanorods, and performing ultrasonic dispersion uniformly;
b. dropwise adding absolute ethyl alcohol under magnetic stirring until the color of the system changes into a milky color, adding 30 mu L of 25% glutaraldehyde solution for crosslinking for 4 h, and centrifugally collecting nanospheres;
c. dissolving 100 μ L of Anti-VEGF in 10 mL of deionized water, adding 12 mg of 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDC) and 8 mg of N-hydroxysuccinimide (NHS) to activate the antibody carboxyl for 2-4 h at room temperature;
d. adding 15 mg of nanosphere into the reaction system, stirring for 4 h, centrifuging and collecting, and washing with deionized water for three times to remove unreacted antibody to obtain the product.
The average particle size of the product is 215 nm, and under the irradiation of laser of 808nm, the temperature of 200 mug/mL product is raised to 52 ℃ within 1 min, and the product targets the vascular endothelial growth factor.
8. An intestinal targeted photoacoustic imaging contrast agent, which is characterized by being prepared according to the method of any one of claims 1 to 7, wherein the average particle size of the obtained product is 173-228 nm, and under the irradiation of laser light at 808nm, 200 μ g/mL of the product is heated to 45-52 ℃ within 1 min, and the targeted antibodies are intestinal targeted 5-hydroxytryptamine receptor 3, intestinal histamine receptor H1 and vascular endothelial growth factor.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN114306276A (en) * | 2021-12-28 | 2022-04-12 | 上海纳米技术及应用国家工程研究中心有限公司 | Preparation method of multifunctional intestinal tract diagnosis and treatment preparation and product thereof |
| WO2022179307A1 (en) * | 2021-02-24 | 2022-09-01 | 上海纳米技术及应用国家工程研究中心有限公司 | Method for preparing multifunctional intestinal diagnosis and treatment formulation, and product thereof |
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