CN105478754A - Preparation method of Cu nanometer material and cancer cell detection method - Google Patents
Preparation method of Cu nanometer material and cancer cell detection method Download PDFInfo
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- CN105478754A CN105478754A CN201610004429.7A CN201610004429A CN105478754A CN 105478754 A CN105478754 A CN 105478754A CN 201610004429 A CN201610004429 A CN 201610004429A CN 105478754 A CN105478754 A CN 105478754A
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- B—PERFORMING OPERATIONS; TRANSPORTING
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- B22F1/00—Metallic powder; Treatment of metallic powder, e.g. to facilitate working or to improve properties
- B22F1/07—Metallic powder characterised by particles having a nanoscale microstructure
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- C—CHEMISTRY; METALLURGY
- C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
- C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
- C23C18/00—Chemical coating by decomposition of either liquid compounds or solutions of the coating forming compounds, without leaving reaction products of surface material in the coating; Contact plating
- C23C18/16—Chemical coating by decomposition of either liquid compounds or solutions of the coating forming compounds, without leaving reaction products of surface material in the coating; Contact plating by reduction or substitution, e.g. electroless plating
- C23C18/18—Pretreatment of the material to be coated
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- C—CHEMISTRY; METALLURGY
- C23—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; CHEMICAL SURFACE TREATMENT; DIFFUSION TREATMENT OF METALLIC MATERIAL; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL; INHIBITING CORROSION OF METALLIC MATERIAL OR INCRUSTATION IN GENERAL
- C23C—COATING METALLIC MATERIAL; COATING MATERIAL WITH METALLIC MATERIAL; SURFACE TREATMENT OF METALLIC MATERIAL BY DIFFUSION INTO THE SURFACE, BY CHEMICAL CONVERSION OR SUBSTITUTION; COATING BY VACUUM EVAPORATION, BY SPUTTERING, BY ION IMPLANTATION OR BY CHEMICAL VAPOUR DEPOSITION, IN GENERAL
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- C23C18/16—Chemical coating by decomposition of either liquid compounds or solutions of the coating forming compounds, without leaving reaction products of surface material in the coating; Contact plating by reduction or substitution, e.g. electroless plating
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- C23C18/32—Coating with nickel, cobalt or mixtures thereof with phosphorus or boron
- C23C18/34—Coating with nickel, cobalt or mixtures thereof with phosphorus or boron using reducing agents
- C23C18/36—Coating with nickel, cobalt or mixtures thereof with phosphorus or boron using reducing agents using hypophosphites
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- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
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Abstract
The invention discloses a preparation method of a Cu nanometer material. The preparation method comprises the following steps that bar-shaped ZnO is prepared through a direct deposition method, then ZnO is plated with Ni, nickel-plated ZnO is wrapped by Cu, and finally the Cu-wrapped nickel-plated ZnO nanometer material is modified through hyaluronic acid. A cancer cell detection method comprises the following steps that the Cu nanometer material is subjected to fluorescein marking, then the Cu nanometer material and cancer cells are cultivated together, and then put under a laser microscope to be observed, and the number of the cancer cells is quantitatively detected according to the staining quantity. The Cu nanometer material prepared through the method has the beneficial effects of being easy to prepare, good in biocompatibility, capable of being combined with the cancer cell specific receptor CD44 and the like. According to the method for utilizing the Cu nanometer materials for detecting the cancer cells, only the blood of a patient needs to be collected for in-vitro detection, the cancer cells can be recognized in few samples, and the cancer can be accurately, sensitively and atraumatically diagnosed in the initial stage of illness.
Description
Technical field
The invention belongs to nano material preparation and biomedical applications field, particularly relate to the preparation method of the coated nickel plating ZnO nano material of a kind of hyaluronic acid decorated Cu and utilize the method for this material tests cancer cell.
Background technology
Cancer is one of principal disease threatening human health and life, and the incidence of disease also rises day by day.At present, a large amount of clinical fact proves, the cure rate that carninomatosis becomes the initial stage is quite high.Therefore, early diagnosis is significant to the survival rate improving cancer patient.This is in the urgent need to finding a kind of method of early diagnosis accurate, sensitive, simple to operate.
At present, cancer test mode mainly contains B ultrasonic, CT, MRI, biopsy etc.B ultrasonic utilizes tangent plane ultrasonogram, can from angle, multi-faceted continuously observation is shown to focus, contribute to the position understanding tumour, with the relation of surrounding tissue organ, have valuable help to the Diagnosis and Treat of cancer, but its scope of application be less.CT, is called for short electronic computer transverse laminagraphy, can be used for checking whole body focus, can specify the scope that carninomatosis stove is invaded, whether invade the organ of surrounding, but CT equipment price costliness costly, and excessive CT can exist and brings out the danger such as malignant tumour, leukaemia.MRI, be called for short Magnetic resonance imaging art, it can be clarified a diagnosis to tumour, cardiac tumor etc. in brain deep tumor, brain stem tumor, tumor of spinal cord, mediastinum, but be equipped with in pacemaker, body have shell fragment to remain, the patient leaving metal silver brain clip etc. that performs the operation can not use.Biopsy is divided into Biopsy and aspiration biopsy two kinds of modes, and Biopsy carries out biopsy in surgical procedure, can determine tumor resection scope, helps the potential diffusion and the actual dispersion that check tumour, determines cancer staging, thus determine successive treatment scheme; Aspiration biopsy needs under the guiding of CT, B ultrasonic etc., and puncture needle is penetrated diseased region, gets living tissue and do pathological diagnosis, aspiration biopsy has the possibility causing needle track implantation, cancerous swelling hemorrhage, and it is careful to need during Clinical practice.
Summary of the invention
In view of this, an object of the present invention is to provide the preparation method of Cu nano material, the coated nickel plating ZnO nano material of hyaluronic acid decorated Cu can be obtained by the method, the advantage of described nano material both magnetic nano material and Cu nano material, have preparation simple, with low cost, do not reunite, good dispersion, the advantage that can couple together with CD44 antibody easily.
The present invention realizes above-mentioned technical purpose by following technological means:
The preparation method of Cu nano material of the present invention, comprises the following steps:
Directly depositing is utilized to prepare bar-shaped ZnO;
Ni is plated on ZnO, obtains the ZnO of nickel plating.
With the ZnO of the coated nickel plating of Cu, namely obtain the nickel plating ZnO nano material that Cu is coated;
Adopt the hyaluronic acid nickel plating ZnO nano coated to Cu to wipe to modify, it comprises the following steps: hyaluronic acid and buffer solution are placed in stirred in water bath, the nickel plating ZnO that Cu is coated is added again after making hyaluronic acid be scattered in solution, Keep agitation, adopt super filter tube to remove unnecessary hyaluronic acid, thus obtain the coated nickel plating ZnO nano material of hyaluronic acid decorated Cu.
W metal is that one has magnetic, prepares relatively simple and the good magnetic material of biocompatibility.The stable chemical nature of Cu, its nanoscale has the spectral characteristic of bioaffinity and uniqueness, and the price of Cu is cheap compared with gold and silver, form nano composite material with the magnetic Nano material that Cu is coated and effectively overcome magnetic Nano material poor stability, easy reunion, the shortcomings such as surface functional group is few, improve the physical and chemical performance of magnetic material.ZnO and Cu nanometer combines biocompatibility, anti-interference, the electric transmission speed that can improve Cu nanometer, and its surface is more easily modified.
Hyaluronic acid, have another name called hyaluronic acid or sodium hyaluronate, its specific receptor CD44 overexpression in multiple cancer cell, the CD44 acceptor on normal cell surface is then in a kind of inactive silence state, therefore realizes targeted imaging analysis to cancer cell or quantitative analysis by the combination of hyaluronic acid and its acceptor CD44.
Further, described Ni is plated on ZnO, it comprises the following steps: by ZnO sensitization in the solution of stannous chloride and hydrochloric acid, put into again in palladium chloride and hydrochloric acid solution and activate, ZnO washed with de-ionized water after activation, and then carry out chemical plating in the chemical plating pond being put into by ZnO containing nickelous sulfate, sodium hypophosphite, ammonium chloride, natrium citricum, succinic acid and lactic acid, finally the ZnO of plated nickel filtered and spend deionized water and dry, namely obtaining the ZnO of nickel plating.
Further, the ZnO of the coated nickel plating of described Cu, it comprises the following steps: moved in deionized water by the nickel plating ZnO of preparation, add polyethylene glycol, and under ultrasound condition, dispersion obtains the suspension of nickel plating ZnO; Cuprous oxide powder is moved in deionized water and fully dissolves, obtain cuprous oxide suspension; Cuprous oxide suspension is mixed with nickel plating ZnO suspension, through standing, filtration, washing, alcohol wash post-drying, namely obtains the nickel plating ZnO nano material that Cu is coated.
Two of object of the present invention is to provide the cancer cell detection method of Cu nano material, detected by the method, only need gather the blood of patient, and just can identify cancer cell in few sample, realizing the ill initial stage can accurate, sensitive, AT cancer diagnosis.
The present invention realizes above-mentioned technical purpose by following technological means:
A kind of cancer cell detection method of the present invention, comprises the following steps:
A) fluorescein-labelled, described Cu nano material is dissolved in deionized water, adds fluorescein isothiocynate and mark;
B) cultivation of cancer cell, 80% ~ 90% cancer cell converged is reached after going down to posterity, be inoculated in orifice plate, after hatching 24h, use alkaline phosphatase brine, add the hyaluronic acid decorated Cu nanomaterial solution of marked by fluorescein isothiocyanate, and after adopting paraformaldehyde solution fixing, use alkaline phosphatase brine, leave standstill after adding 2% potassium ferrocyanide solution of equivalent and 2% hydrogen chloride mixed liquor, lucifuge hatching 18 ~ 30min, wash away dyeing liquor, core fast red is redyed, and removes supernatant, again washs with alkaline phosphatase salting liquid; Adopt each 1 time of 50%, 75%, 85%, 95% and 100% Gradient elution using ethanol, transparent 2 times of dimethylbenzene, each 10 ~ 15min; Neutral gum mounting, observes under being placed in laser confocal microscope, and the cell membrane of tumour cell and cytoplasm have Positive fluorescence to dye, and quantitatively can detect the quantity of cancer cell according to dyeing quantity.
Further, the particle size range of described hyaluronic acid decorated Cu nano material is 40-60nm, and hyaluronic acid is 0.4 ~ 0.5mmol/g in the carrying capacity on the ZnO nano material surface of the coated plating Ni of Cu.
Further, described cancer cell is the one in lung carcinoma cell, HCC, breast cancer cell or stomach cancer cell.
Beneficial effect of the present invention: the advantage of (1) Cu nano material both magnetic nano material and Cu nano material, have preparation simple, with low cost, do not reunite, the advantages such as good dispersion; (2) with hyaluronic acid, finishing is carried out to Cu nano material, not only can improve its stability, and make it surface-functionalized, can couple together with CD44 antibody more easily; (3) blood that the method utilizing hyaluronic acid decorated Cu nano material to detect cancer cell only need gather patient carries out vitro detection, and just can cancer cell be identified in few sample, realizing the ill initial stage can accurate, sensitive, AT cancer diagnosis.
Detailed description of the invention
Below will be described in detail technical scheme of the present invention by specific embodiment:
The preparation method of embodiment 1Cu nano material
The preparation method of hyaluronic acid decorated Cu nano material of the present invention, comprises the following steps:
Directly depositing is utilized to prepare bar-shaped ZnO;
Ni is plated on ZnO, it comprises the following steps: by ZnO sensitization in the solution of stannous chloride and hydrochloric acid, put into again in palladium chloride and hydrochloric acid solution and activate, ZnO washed with de-ionized water after activation, and then carry out chemical plating in the chemical plating pond being put into by ZnO containing nickelous sulfate, sodium hypophosphite, ammonium chloride, natrium citricum, succinic acid and lactic acid, finally the ZnO of plated nickel filtered and spend deionized water and dry, namely obtaining the ZnO of nickel plating.
With the ZnO of the coated nickel plating of Cu, it comprises the following steps: moved in deionized water by the nickel plating ZnO of preparation, add polyethylene glycol, and under ultrasound condition, dispersion obtains the suspension of nickel plating ZnO; Cuprous oxide powder is moved in deionized water and fully dissolves, obtain cuprous oxide suspension; Cuprous oxide suspension is mixed with nickel plating ZnO suspension, through standing, filtration, washing, alcohol wash post-drying, namely obtains the nickel plating ZnO nano material that Cu is coated.
The hyaluronic acid nickel plating ZnO nano material coated to Cu is adopted to modify, it comprises the following steps: hyaluronic acid and buffer solution are placed in stirred in water bath, the nickel plating ZnO that Cu is coated is added again after making hyaluronic acid be scattered in solution, Keep agitation, adopt super filter tube to remove unnecessary hyaluronic acid, thus obtain the coated nickel plating ZnO nano material of hyaluronic acid decorated Cu.
Embodiment 2 cancer cell detection method
Cancer cell detection method of the present invention comprises the following steps:
A) fluorescein-labelled, described Cu nano material is dissolved in deionized water, add fluorescein isothiocynate to mark, the particle size range of described Cu nano material is 40-60nm, and hyaluronic acid is 0.4 ~ 0.5mmol/g in the carrying capacity of Cu nano-material surface;
B) cultivation of cancer cell, described cancer cell is the one in lung carcinoma cell, HCC, breast cancer cell or stomach cancer cell, and concrete way is: after going down to posterity, reach 80% ~ 90% cancer cell converged, with 5 × 10
5individual/hole is inoculated in orifice plate, 30 ~ 37 DEG C, after hatching 24h under 5%CO2 condition, use alkaline phosphatase brine, add the copper nanomaterial solution of marked by fluorescein isothiocyanate, after hatching 24h, fully wash with alkaline phosphatase salting liquid, every hole adds after 1ml4% paraformaldehyde solution fixes 20min, use alkaline phosphatase brine, 5min is left standstill after adding 2% potassium ferrocyanide solution of equivalent and 2% hydrogen chloride mixed liquor, under 37 DEG C of conditions, lucifuge hatching 30min, deionized water washes away dyeing liquor, core fast red is redyed, remove supernatant, again wash with alkaline phosphatase salting liquid.Adopt each 1 time of 50%, 75%, 85%, 95% and 100% Gradient elution using ethanol, transparent 2 times of dimethylbenzene, each 10min.Neutral gum mounting, observes under being placed in inverted microscope and takes pictures.
What finally illustrate is, above embodiment is only in order to illustrate technical scheme of the present invention and unrestricted, although with reference to preferred embodiment to invention has been detailed description, those of ordinary skill in the art is to be understood that, can modify to technical scheme of the present invention or equivalent replacement, and not departing from aim and the scope of technical solution of the present invention, it all should be encompassed in the middle of right of the present invention.
Claims (6)
- The preparation method of 1.Cu nano material, is characterized in that, comprises the following steps:Directly depositing is utilized to prepare bar-shaped ZnO;Ni is plated on ZnO, obtains the ZnO of nickel plating;With the ZnO of the coated nickel plating of Cu, namely obtain the nickel plating ZnO nano material that Cu is coated;The hyaluronic acid nickel plating ZnO nano material coated to Cu is adopted to modify, it comprises the following steps: hyaluronic acid and buffer solution are placed in stirred in water bath, the nickel plating ZnO that Cu is coated is added again after making hyaluronic acid be scattered in solution, Keep agitation, adopt super filter tube to remove unnecessary hyaluronic acid, thus obtain the coated nickel plating ZnO nano material of hyaluronic acid decorated Cu.
- 2. the preparation method of Cu nano material according to claim 1, it is characterized in that, described Ni is plated on ZnO, it comprises the following steps: by ZnO sensitization in the solution of stannous chloride and hydrochloric acid, put into again in palladium chloride and hydrochloric acid solution and activate, ZnO washed with de-ionized water after activation, and then carry out chemical plating in the chemical plating pond being put into by ZnO containing nickelous sulfate, sodium hypophosphite, ammonium chloride, natrium citricum, succinic acid and lactic acid, finally the ZnO of plated nickel filtered and spend deionized water and dry, namely obtaining the ZnO of nickel plating.
- 3. the preparation method of Cu nano material according to claim 1, it is characterized in that, the ZnO of the coated nickel plating of described Cu, it comprises the following steps: moved into by the nickel plating ZnO of preparation in deionized water, add polyethylene glycol, under ultrasound condition, dispersion obtains the suspension of nickel plating ZnO; Cuprous oxide powder is moved in deionized water and fully dissolves, obtain cuprous oxide suspension; Cuprous oxide suspension is mixed with nickel plating ZnO suspension, through standing, filtration, washing, alcohol wash post-drying, namely obtains the nickel plating ZnO nano material that Cu is coated.
- 4. as the cancer cell detection method in claim 1-3 as described in any one, it is characterized in that, comprise the following steps:A) fluorescein-labelled, Cu nano material is dissolved in deionized water, adds fluorescein isothiocynate and mark;B) cultivation of cancer cell, 80% ~ 90% cancer cell converged is reached after going down to posterity, be inoculated in orifice plate, after hatching 24h, use alkaline phosphatase brine, add the solution of the Cu nano material of marked by fluorescein isothiocyanate, and after adopting paraformaldehyde solution fixing, use alkaline phosphatase brine, leave standstill after adding 2% potassium ferrocyanide solution of equivalent and 2% hydrogen chloride mixed liquor, lucifuge hatching 18 ~ 30min, wash away dyeing liquor, core fast red is redyed, and removes supernatant, again washs with alkaline phosphatase salting liquid; Adopt Gradient elution using ethanol, transparent 2 times of dimethylbenzene, each 10 ~ 15min; Neutral gum mounting, is placed in laser capture microdissection Microscopic observation, and the cell membrane of tumour cell and cytoplasm have Positive fluorescence to dye, and quantitatively can detect the quantity of cancer cell according to dyeing quantity.
- 5. cancer cell detection method as claimed in claim 4, is characterized in that: the particle size range of described Cu nano material is 40-60nm, and hyaluronic acid is 0.4-0.5mmol/g in the carrying capacity on the coated nickel plating ZnO nano material surface of Cu.
- 6. cancer cell detection method as claimed in claim 4, is characterized in that: described cancer cell is the one in lung carcinoma cell, HCC, breast cancer cell or stomach cancer cell.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114099677A (en) * | 2022-01-26 | 2022-03-01 | 广东工业大学 | Preparation method and application of copper ferrite nano material |
Citations (4)
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| CN101634629A (en) * | 2009-08-27 | 2010-01-27 | 电子科技大学 | Cancer cell detection method based on optical stretching and optical fiber interference sensing technology |
| CN102072894A (en) * | 2009-11-25 | 2011-05-25 | 欧普图斯(苏州)光学纳米科技有限公司 | Nano-structure-based spectrum detecting method for detecting chemical and biochemical impurities |
| CN103502465A (en) * | 2011-03-31 | 2014-01-08 | 国立大学法人弘前大学 | Method for detecting cancer cell using fluorescently labeled L-glucose derivative, and cancer cell-imaging agent comprising fluorescently labeled L-glucose derivative |
| JP2014227397A (en) * | 2013-05-24 | 2014-12-08 | 独立行政法人国立高等専門学校機構 | Ferromagnetic/non-magnetic complex |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101634629A (en) * | 2009-08-27 | 2010-01-27 | 电子科技大学 | Cancer cell detection method based on optical stretching and optical fiber interference sensing technology |
| CN102072894A (en) * | 2009-11-25 | 2011-05-25 | 欧普图斯(苏州)光学纳米科技有限公司 | Nano-structure-based spectrum detecting method for detecting chemical and biochemical impurities |
| CN103502465A (en) * | 2011-03-31 | 2014-01-08 | 国立大学法人弘前大学 | Method for detecting cancer cell using fluorescently labeled L-glucose derivative, and cancer cell-imaging agent comprising fluorescently labeled L-glucose derivative |
| JP2014227397A (en) * | 2013-05-24 | 2014-12-08 | 独立行政法人国立高等専門学校機構 | Ferromagnetic/non-magnetic complex |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN114099677A (en) * | 2022-01-26 | 2022-03-01 | 广东工业大学 | Preparation method and application of copper ferrite nano material |
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