CN108524953A - A kind of the specificity fluorescent labeling method and its vivo applications of food-borne probiotics - Google Patents
A kind of the specificity fluorescent labeling method and its vivo applications of food-borne probiotics Download PDFInfo
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- 235000018291 probiotics Nutrition 0.000 title claims abstract description 56
- 238000003234 fluorescent labeling method Methods 0.000 title claims abstract description 20
- 239000000523 sample Substances 0.000 claims abstract description 39
- 239000011701 zinc Substances 0.000 claims abstract description 23
- GYHNNYVSQQEPJS-UHFFFAOYSA-N Gallium Chemical compound [Ga] GYHNNYVSQQEPJS-UHFFFAOYSA-N 0.000 claims abstract description 19
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 claims abstract description 19
- 229910052733 gallium Inorganic materials 0.000 claims abstract description 19
- 229910052725 zinc Inorganic materials 0.000 claims abstract description 19
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/001—Preparation for luminescence or biological staining
- A61K49/0013—Luminescence
- A61K49/0017—Fluorescence in vivo
- A61K49/0019—Fluorescence in vivo characterised by the fluorescent group, e.g. oligomeric, polymeric or dendritic molecules
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/001—Preparation for luminescence or biological staining
- A61K49/0013—Luminescence
- A61K49/0017—Fluorescence in vivo
- A61K49/005—Fluorescence in vivo characterised by the carrier molecule carrying the fluorescent agent
- A61K49/0058—Antibodies
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- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K49/00—Preparations for testing in vivo
- A61K49/001—Preparation for luminescence or biological staining
- A61K49/0063—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres
- A61K49/0065—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres the luminescent/fluorescent agent having itself a special physical form, e.g. gold nanoparticle
- A61K49/0067—Preparation for luminescence or biological staining characterised by a special physical or galenical form, e.g. emulsions, microspheres the luminescent/fluorescent agent having itself a special physical form, e.g. gold nanoparticle quantum dots, fluorescent nanocrystals
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- C09K11/08—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials
- C09K11/67—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials containing refractory metals
- C09K11/68—Luminescent, e.g. electroluminescent, chemiluminescent materials containing inorganic luminescent materials containing refractory metals containing chromium, molybdenum or tungsten
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Abstract
The present invention relates to the specificity fluorescent labeling method of food-borne probiotics, include the following steps that (a) by solvent heat high-temperature calcination, prepares the Cr for having near-infrared fluorescent and overlength afterglow property3+Doped gallium zinc germanate long-afterglow fluorescent nano-probe；(b) by antibody surface functionalization, realize that nano-probe marks the specificity fluorescent of target probiotics.The beneficial effects of the present invention are：(1) the gallium zinc germanate long-persistence nano probe prepared by the present invention has superpower near-infrared luminous and overlength afterglow service life, excellent biocompatibility and structural stability, good size uniformity and hypotoxicity.(2) the near-infrared fluorescent label probiotics vivo biodistribution imaging technique that the present invention is developed, the lossless prebiotic thalline of visually tracer can be achieved and enter the distribution situation after organism, be of great significance for the functional site and Nutritional studies theory of opening up innovative food-borne probiotics.
Description
Technical field
The invention belongs to the research fields of food-borne probiotics, are related to the specificity fluorescent labeling method of food-borne probiotics
And its vivo applications.
Background technology
Probiotics is the pith of intestinal microflora, when it takes in enough quantity, can give consumer (place
It is main) the next great benefit of health care belt.Probiotics can greatly facilitate human body intestinal canal health, adjust the immune response of human body.Although
Have the function of great benefit and special to health, but in vivo with the relevant metabolism of probiotics, distribution and immunological regulation
Also still not very clear, function and active function to most of new species are still not enough illustrated, and need to obtain more information
It is assessed.
Modern molecular biology technique such as polymerase chain reaction and denaturing gradient gel electrophoresis (PCR-DGGE) at present, in real time
The various methods such as quantitative PCR, DNA microarray, pyrosequencing and high-flux sequence determine the DNA sequence dna of biological sample, disclose
The type of probiotics, distribution and metabolism.However, above-mentioned technology usually requires to collect great amount of samples (excrement or in-vivo tissue), if
Probe/primer, for statistical analysis to a large amount of data is counted, is taken time and effort.In addition, these methods can not carry out inspection in situ
It surveys, it is virtually impossible to which true reappearance is carried out to probiotics distribution situation in organism by non-destructive and real-time in-situ monitoring.Cause
This, there is an urgent need for establish one kind can quickly, it is intuitive and there is nondestructive real-time in-situ monitoring technology, be probiotics in vivo
Distribution and functional site probe into offer technical support.
For the above research method demand and field blank, we introduce Imaging-PAM.Fluorescence imaging is as spy
The key technology of rope early diagnosis of tumor treatment is the leading edge method now in biomedical sector and modern analysis science, especially
It is the occurrence and development that non-intruding monitor tumour in live body level is imaged on using non-invasively imaged and minimally invasive vivo biodistribution, for
The prevention and early stage diagnosis and treatment of cancer have important life medical research meaning and clinical value.By this non-invasively imaged skill
Art is applied in food-borne microorganism In vivo study, can give full play to the advantage of fluorescence imaging real non-destructive monitoring, Jin Erke
Enter the distribution situation after organism and metabolism behavior with systematic research probiotics.
The image effect of fluorescence imaging depends on the luminescent properties and targeting-functionalization of contrast agent (i.e. fluorescence probe).It is modern
It is that biology penetrates window (700-1000nm and 1100- that optical detection in bio-imaging, which requires the working region of (1) fluorescence probe,
Near-infrared region 1350nm) works to increase the deep tissue penetration of detection light；(2) exciting light of fluorescence probe should not be adopted
With ultraviolet light and visible light, to avoid the autofluorescence of tissue；(3) excitating light strength of fluorescence probe should not be too strong, to drop
Low background signal increases the signal-to-noise ratio of imaging；(4) it is capable of providing real-time, whole, distributed observed result.Fluorescence nano is visited
Needle is different from small molecule image-forming contrast medium or drug, it can not only overcome space existing for routine clinical diagnosing tumor technology point
Resolution is low, contrast in tissue is low, accuracy is poor, is easy the problems such as leaky bucket model, can also solve chemistry treatment in cancer treatment method
Blood circulation time existing for method is short, therapeutic efficiency is low, lacks specificity, damages the problems such as human normal cell.In recent years
Based on different optical physics chemical properties, researcher gradually proposes several different fluorescence probes, including：Fluorescence-encoded egg
In vain, organic dyestuff, semiconductor-quantum-point, noble metal nano particles, carbon nanomaterial and up-conversion luminescent material.However these
Material all has respective defect to varying degrees, such as poor signal to noise, shortage optical stability, photobleaching, coherent light swash
Hair, luminous intensity are unstable etc..
Long-persistence nano-luminescent materials (Persistent Luminescent Nanophosphors, PLNPs) are a kind of
By high energy excite (visible light, ultraviolet light, X-ray, gamma-rays, electron beam etc.) after can be generated visible or near infrared region it is long when
Between shine special nano material.Compared to traditional nano luminescent material, PLNPs has following as bio-imaging contrast agent
Advantage：(1) long-persistence luminous nano material toxicity is relatively low, and the In vivo study of living body biological is more suitable for after surface modification；(2)
The photochemical stability of long-persistence nano material is preferable, can be to avoid the generation of photobleaching phenomenon；(3) long-persistence nano material can
With external excitation carry out optical imagery, can effectively avoid in this way organism self-luminous and excitation light source bias light at
As the influence of effect；(4) shining for long-persistence nano material can be regulated and controled by changing synthesis condition and element composition to close red
Exterior domain, the biological tissue that can increase detection light penetrate, and improve the sensitivity of optical imagery.All potential fluorescence are made a general survey of to visit
Needle, only near-infrared long after glow luminous material can be good at the needs for meeting contemporary optics bio-imaging.Therefore, by more than length
Brightness nano-probe carries out fluorescent marker to probiotics, you can realizes the metabolism point to probiotics in vivo by bio-imaging
Cloth situation is monitored in real time, and new research ideas is opened up with health research for food nutrition.
Invention content
It will be with the luminous Cr of near-infrared (NIR) by optical bio imaging technique3+The gallium zinc germanate long afterglow of doping is received
Rice corpuscles (ZGGO PLNPs) is introduced into as optical probe in the research that food-borne probiotics is distributed in vivo.It uses
Cr3+The gallium zinc germanate long-persistence nano particle of doping has excellent afterglow property, low toxicity, good dispersibility, optical stabilization
Property and biocompatibility, can be used as a kind of tracer by antibody (gram-positive bacteria LTA antibody) to target probiotics into
Line flag, the vigor of thalline still is able to be maintained at 80% or more after label, reaches the long-term visualizing monitor to probiotics.Pass through
It takes orally after entering in Mice Body, by the optical bio imaging technique excited outside long afterglow materials, thalline after real-time research label
Absorption in body gastrointestinal tract and distribution situation.ZGGO antibody functionalized LTA:Cr3+Nano-probe can effectively and successfully
Label probiotics, and the bio distribution situation of the probiotics for real-time, non-damaging monitoring intake organism.
Specific technical solution provided by the invention is：
A kind of specificity fluorescent labeling method of food-borne probiotics, includes the following steps：
(a) by solvent heat-high-temperature calcination, the Cr for having near-infrared fluorescent and overlength afterglow property is prepared3+Doping
Gallium zinc germanate long-afterglow fluorescent nano-probe；
(b) it by antibody surface functionalization, realizes and the specificity fluorescent of target probiotics is marked.
Further, the step (a) prepares Cr3+The process of the gallium zinc germanate long-persistence nano probe of doping is as follows：
(1) metal ion solution is mixed according to a certain percentage and adjusts pH and bring it about coprecipitation reaction to obtain instead
Answer liquid；
(2) reaction solution prepared by step (1) is subjected to solvent thermal reaction in ptfe autoclave；
(3) it is calcined in Muffle furnace after slightly being ground by product centrifuge washing in step (2) and using agate mortar；
(4) by the solid powder grinding obtained by step (3), simultaneously centrifugal screening obtains the long-persistence nano spy of uniform particle diameter
Needle.
Further, each metallic element ratio of step (1) is prepared according to the chemical structural formula of nano material
Zn1.25Ga1.5Ge0.25O4:0.5%Cr3+, solvent is 2mL oleic acid and 15mL toluene, and pH is adjusted using tert-butylamine, and final pH is
7.5。Zn1.25Ga1.5Ge0.25O4:0.5%Cr3+The material that this ratio obtains, luminosity are particularly suited for grinding in organism
Study carefully.Organic solvent is to use for the first time, can improve the dispersibility of nano material；PH is adjusted using tert-butylamine, and unique, more
Mildly, the property of other compositions is not influenced.
Further, the step (2) solvent thermal reaction temperature in ptfe autoclave is 120 DEG C, the reaction time
It is 24 hours.
Further, the step (3) calcination temperature in Muffle furnace is 750 DEG C, and the reaction time is 5 hours.
Further, the step (b) marks the process of probiotics as follows by antibody surface functionalization：
(1) Cr for obtaining step (a)3+The gallium zinc germanate long-afterglow fluorescent nano-probe of doping is distributed to 2mL N, N- bis-
In methylformamide, magnetic agitation 30 minutes；
(2) 3- aminopropyl triethoxysilanes (APTES) are added in the solution of step (1), magnetic agitation 8 hours；
(3) step (2) is obtained into solution centrifugation and removes unreacted 3- aminopropyl triethoxysilanes, antibody is then added
And catalyst, room temperature are protected from light shaking 30 minutes, centrifuge washing；
(4) step (3) is obtained solution to mix with the prebiotic bacterium solution of target, is incubated.
Further, 3- aminopropyl triethoxysilane usage amounts are 20 μ L in the step (2).
Further, the antibody used in the step (3) is gram-positive bacteria LTA antibody, and 1mg antibody is dissolved in
10mL PBS (0.1M, pH 7.4)) it is modified, method of modifying is that EDC-NHS is catalyzed the reaction of carboxylic ammonia, and EDC-NHS is as catalysis
Agent.
Further, incubation conditions are that jog is incubated 1 hour at 37 DEG C in the step (4).
A kind of vivo applications of the specificity fluorescent labeling method of food-borne probiotics are entered by oral in organism,
By the optical bio imaging technique excited outside long-persistence nano probe body, the metabolism of real time monitoring probiotics in vivo was distributed
Journey.
The application of the present invention is to be entered in organism by oral, and skill is imaged by the optical bio excited outside long afterglow materials
Art, the metabolism distributed process of real time monitoring probiotics in vivo.
Conventional fluorescence labeling material, such as organic dyestuff, fluorescin, semiconductor-quantum-point etc., is respectively present fluorescence
The defects of stability is poor, biological imaging signal-to-noise ratio is low, toxicity is high, application is limited.Near-infrared prepared by the present invention
Long-persistence nano material, it is excellent to have that luminous efficiency is high, fluorescence lifetime is long, toxicity is relatively low, photochemical stability is good, synthesis is controllable etc.
Gesture, and carry out optical imagery can be excited in vitro, it can effectively avoid the bias light of the self-luminous and excitation light source of organism
Influence to imaging effect, improves the sensitivity of optical imagery, therefore is highly suitable for the fluorescent marker of probiotics, thus into
One step realizes that probiotics the real time monitoring of behavior and is probed into vivo.
The beneficial effects of the present invention are：
(1) the gallium zinc germanate long-persistence nano probe prepared by the present invention has the superpower near-infrared luminous and overlength afterglow longevity
Life, excellent biocompatibility and structural stability, good size uniformity and hypotoxicity fit through surface antibody modification
Specific marker target probiotics.
(2) the near-infrared fluorescent label probiotics vivo biodistribution imaging technique that the present invention is developed is, it can be achieved that lossless visual
Change the ground prebiotic thalline of tracer and enter the distribution situation after organism, for opening up the functional site of innovative food-borne probiotics
And Nutritional studies theory is of great significance.
Description of the drawings：
Fig. 1：The transmission electron microscope picture of gallium zinc germanate long-persistence nano probe
Fig. 2：The biocompatibility of gallium zinc germanate long-afterglow fluorescent nano-probe
Fig. 3：The hypotoxicity of gallium zinc germanate long-afterglow fluorescent nano-probe：(a) cytotoxicity；
Fig. 4：The hypotoxicity of gallium zinc germanate long-afterglow fluorescent nano-probe：(b) live body toxicity
Fig. 5：The transmission electron microscope picture of fluorescence labeling probiotics
Specific implementation mode
In order to keep features described above and advantage of the present invention more clear and be readily appreciated that, below in conjunction with attached drawing to the present invention's
Embodiment is described in further detail.
Embodiment 1
A kind of specificity fluorescent labeling method of food-borne probiotics
(1) by solvent heat-high-temperature calcination, the Cr for having near-infrared fluorescent and overlength afterglow property is prepared3+
The gallium zinc germanate long-afterglow fluorescent nano-probe of doping.
Accurately weigh 0.013mmol Cr (NO3)3·9H2O and 3.01mmol Zn (NO3)2·6H2O solves homogeneously in 10mL
The 0.6mol L configured-1Ga(NO3)3Magnetic agitation in solution makes it be uniformly mixed, is total to tert-butylamine tune pH to 7.5
Precipitation continues magnetic agitation 2h, and being ultrasonically treated 30min makes it be uniformly mixed, and the white emulsion of formation is placed in polytetrafluoroethyl-ne
120 DEG C of hydro-thermal reactions are for 24 hours in alkene reaction kettle.It after taking out after naturally cool to room temperature, is added in 2 times of volume ethanols, is uniformly mixed,
Precipitation is generated, 7000rpm 10min centrifugations are collected sediment and washed 2-3 times with absolute ethyl alcohol.The precipitation centrifuged is placed in 80
In DEG C vacuum drying chamber after dry 3h, after slightly being ground using agate mortar, it is put in 750 DEG C of calcining 5h in Muffle furnace.The length prepared
Twilight sunset nano-probe material is ground 2~3 times with ethyl alcohol is wet, is resuspended in 5mmol L-1NaOH solution in, and magnetic agitation mistake
Night.4500rmp centrifugations 7min takes supernatant liquid with 3500rmp centrifugations 15min again after taking upper layer turbid solution, by the upper liquid of acquisition
Body freeze-drying is final to obtain the long-persistence nano probe material that grain size is 30~80nm.
(2) it by antibody surface functionalization, realizes and the specificity fluorescent of target probiotics is marked.
The obtained long-persistence nano probe of 5mg steps (1) accurately is weighed, its ultrasound is resuspended in 2mL N, N- dimethyl
In formamide (DMF), 20 μ L 3- aminopropyl triethoxysilanes (APTES) are then added dropwise, reaction solution is acute at 80 DEG C
Strong stirring is for 24 hours to form the PLNPs (NH2-ZGGO) of surface amination.1mg antibody is dissolved in 10mL PBS (0.1M, pH
7.4) in, NHS (15mg) and EDC (15mg) is then added.It is anti-with complete activation that said mixture is incubated to 30min at room temperature
Carboxyl on body.By the way that NHs of the 10mg through ultrasonic disperse is added into 10mL PBS (0.1M, pH 7.4) buffer solution2-ZGGO
In PLNPs suspension.Above two solution is mixed, after being protected from light jog under room temperature and being incubated 4h, by gained mix products
(antibody-ZGGO) at 4 DEG C 7000rpm centrifuge 5min, after washed twice with PBS buffer solution, the ZGGO for obtaining antibody modification is glimmering
Light probe.Lactobacillus casei is added thereto, jog is incubated 1h at 37 DEG C, completes fluorescent marker.
Embodiment 2
A kind of specificity fluorescent labeling method of food-borne probiotics, step and method are substantially the same manner as Example 1, different
Place is that probiotics is bifidobacterium lactis V9, accordingly obtains the bifidobacterium lactis V9 of fluorescent marker.
Embodiment 3
A kind of specificity fluorescent labeling method of food-borne probiotics, step and method are substantially the same manner as Example 1, different
Place is that probiotics is lactobacillus reuteri, accordingly obtains the lactobacillus reuteri of fluorescent marker.
Embodiment 4
A kind of specificity fluorescent labeling method of food-borne probiotics, step and method are substantially the same manner as Example 1, different
Place is that probiotics is lactobacillus acidophilus, accordingly obtains the lactobacillus acidophilus of fluorescent marker.
Fig. 1 is the transmission electron microscope picture of gallium zinc germanate long-persistence nano particle, and it is good to show that the fluorescent nano probe prepared has
Good size uniformity, size are suitable for biomarker in 50nm or so.
Fig. 2 is the Study on Biocompatibility of gallium zinc germanate long-afterglow fluorescent nano-probe, and common large biological molecule is to it
Photoluminescent property shows that the fluorescent nano probe prepared has good biocompatibility, is suitable for without significant impact
Living imaging is studied.
Fig. 3 is the Toxicity test of gallium zinc germanate long-afterglow fluorescent nano-probe, and three kinds of cell strains of nano-probe pair do not have
Apparent toxicity is monitored by control mice group and experimental group weight, it was demonstrated that probe does not have significant bio-toxicity yet, finally
Show that nano-probe toxicity is relatively low.
Fig. 4 is the transmission electron microscope picture of fluorescence labeling probiotics, shows to combine by antibody modification and antibody surface, nanometer is visited
Needle pass flag is to phage surface.
The foregoing is merely the preferred embodiments of the invention, are not intended to limit the invention creation, all at this
Within the spirit and principle of innovation and creation, any modification, equivalent replacement, improvement and so on should be included in the invention
Protection domain within.
Claims (10)
1. a kind of specificity fluorescent labeling method of food-borne probiotics, which is characterized in that include the following steps：
(a) by solvent heat-high-temperature calcination, the Cr for having near-infrared fluorescent and overlength afterglow property is prepared3+The gallium germanium of doping
Sour zinc long-afterglow fluorescent nano-probe；
(b) it by antibody surface functionalization, realizes and the specificity fluorescent of target probiotics is marked.
2. a kind of specificity fluorescent labeling method of food-borne probiotics according to claim 1, which is characterized in that described
Step (a) prepares Cr3+The process of the gallium zinc germanate long-persistence nano probe of doping is as follows：
(1) metal ion solution is mixed according to a certain percentage and adjusts pH and bring it about coprecipitation reaction to be reacted
Liquid；
(2) reaction solution prepared by step (1) is subjected to solvent thermal reaction in ptfe autoclave；
(3) it is calcined in Muffle furnace after slightly being ground by product centrifuge washing in step (2) and using agate mortar；
(4) by the solid powder grinding obtained by step (3), simultaneously centrifugal screening obtains the long-persistence nano probe of uniform particle diameter.
3. a kind of specificity fluorescent labeling method of food-borne probiotics according to claim 2, which is characterized in that described
Each metallic element ratio of step (1) prepares Zn according to the chemical structural formula of nano material1.25Ga1.5Ge0.25O4:0.5%Cr3+, molten
Agent is 2mL oleic acid and 15mL toluene, and pH, which is adjusted, uses tert-butylamine, final pH 7.5.
4. a kind of specificity fluorescent labeling method of food-borne probiotics according to claim 2, which is characterized in that the step
Suddenly (2) solvent thermal reaction temperature in ptfe autoclave is 120 DEG C, and the reaction time is 24 hours.
5. a kind of specificity fluorescent labeling method of food-borne probiotics according to claim 2, which is characterized in that the step
Suddenly (3) calcination temperature in Muffle furnace is 750 DEG C, and the reaction time is 5 hours.
6. a kind of specificity fluorescent labeling method of food-borne probiotics according to claim 1, which is characterized in that described
Step (b) marks the process of probiotics as follows by antibody surface functionalization：
(1) Cr for obtaining step (a)3+The gallium zinc germanate long-afterglow fluorescent nano-probe of doping is distributed to 2mL N, N- dimethyl
In formamide, magnetic agitation 30 minutes；
(2) 3- aminopropyl triethoxysilanes are added in the solution of step (1), magnetic agitation 8 hours；
(3) step (2) is obtained into solution centrifugation and removes unreacted 3- aminopropyl triethoxysilanes, antibody is then added and urges
Agent, room temperature are protected from light shaking 30 minutes, centrifuge washing；
(4) step (3) is obtained solution to mix with the prebiotic bacterium solution of target, is incubated.
7. a kind of specificity fluorescent labeling method of food-borne probiotics according to claim 6, which is characterized in that described
3- aminopropyl triethoxysilane usage amounts are 20 μ L in step (2).
8. a kind of specificity fluorescent labeling method of food-borne probiotics according to claim 6, which is characterized in that described
Antibody used in step (3) is gram-positive bacteria LTA antibody, and 1mg antibody is dissolved in 10mL PBS and is modified, is repaiied
Decorations method is that EDC-NHS is catalyzed carboxyl-amino reaction.
9. a kind of specificity fluorescent labeling method of food-borne probiotics according to claim 6, which is characterized in that described
Incubation conditions are that jog is incubated 1 hour at 37 DEG C in step (4).
10. a kind of vivo applications of the specificity fluorescent labeling method of food-borne probiotics, which is characterized in that entered by oral
In organism, by the optical bio imaging technique excited outside long-persistence nano probe body, real time monitoring probiotics is in vivo
It is metabolized distributed process.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109781687A (en) * | 2019-01-29 | 2019-05-21 | 南开大学 | A kind of preparation method and its vivo applications of composite fluorescence nano-probe |
| CN110665480A (en) * | 2019-09-25 | 2020-01-10 | 南开大学 | Preparation method and application of long-afterglow molecularly imprinted polymer |
| CN111956186A (en) * | 2020-08-19 | 2020-11-20 | 湖南大学 | Kit for monitoring activity degree of signal path in body |
-
2018
- 2018-04-25 CN CN201810380388.0A patent/CN108524953A/en active Pending
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109781687A (en) * | 2019-01-29 | 2019-05-21 | 南开大学 | A kind of preparation method and its vivo applications of composite fluorescence nano-probe |
| CN110665480A (en) * | 2019-09-25 | 2020-01-10 | 南开大学 | Preparation method and application of long-afterglow molecularly imprinted polymer |
| CN111956186A (en) * | 2020-08-19 | 2020-11-20 | 湖南大学 | Kit for monitoring activity degree of signal path in body |
| CN111956186B (en) * | 2020-08-19 | 2022-07-01 | 湖南大学 | Kit for monitoring activity degree of signal path in body |
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Application publication date: 20180914 |