CN107754723A - A kind of fluorocarbon surfactant available for high flux numeral droplet PCR and its preparation method and application - Google Patents
A kind of fluorocarbon surfactant available for high flux numeral droplet PCR and its preparation method and application Download PDFInfo
- Publication number
- CN107754723A CN107754723A CN201711015652.2A CN201711015652A CN107754723A CN 107754723 A CN107754723 A CN 107754723A CN 201711015652 A CN201711015652 A CN 201711015652A CN 107754723 A CN107754723 A CN 107754723A
- Authority
- CN
- China
- Prior art keywords
- phase fluid
- numeral
- high flux
- fluorocarbon surfactant
- anionicsite
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C09—DYES; PAINTS; POLISHES; NATURAL RESINS; ADHESIVES; COMPOSITIONS NOT OTHERWISE PROVIDED FOR; APPLICATIONS OF MATERIALS NOT OTHERWISE PROVIDED FOR
- C09K—MATERIALS FOR MISCELLANEOUS APPLICATIONS, NOT PROVIDED FOR ELSEWHERE
- C09K23/00—Use of substances as emulsifying, wetting, dispersing, or foam-producing agents
- C09K23/16—Amines or polyamines
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- C08G65/00—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule
- C08G65/02—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring
- C08G65/04—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring from cyclic ethers only
- C08G65/22—Cyclic ethers having at least one atom other than carbon and hydrogen outside the ring
- C08G65/223—Cyclic ethers having at least one atom other than carbon and hydrogen outside the ring containing halogens
- C08G65/226—Cyclic ethers having at least one atom other than carbon and hydrogen outside the ring containing halogens containing fluorine
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08G—MACROMOLECULAR COMPOUNDS OBTAINED OTHERWISE THAN BY REACTIONS ONLY INVOLVING UNSATURATED CARBON-TO-CARBON BONDS
- C08G65/00—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule
- C08G65/02—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring
- C08G65/26—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring from cyclic ethers and other compounds
- C08G65/2618—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring from cyclic ethers and other compounds the other compounds containing nitrogen
- C08G65/2621—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring from cyclic ethers and other compounds the other compounds containing nitrogen containing amine groups
- C08G65/2624—Macromolecular compounds obtained by reactions forming an ether link in the main chain of the macromolecule from cyclic ethers by opening of the heterocyclic ring from cyclic ethers and other compounds the other compounds containing nitrogen containing amine groups containing aliphatic amine groups
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6844—Nucleic acid amplification reactions
- C12Q1/6851—Quantitative amplification
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Life Sciences & Earth Sciences (AREA)
- Medicinal Chemistry (AREA)
- Polymers & Plastics (AREA)
- Zoology (AREA)
- Wood Science & Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biotechnology (AREA)
- Biochemistry (AREA)
- Immunology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Analytical Chemistry (AREA)
- Physics & Mathematics (AREA)
- Biophysics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Materials Engineering (AREA)
- Emulsifying, Dispersing, Foam-Producing Or Wetting Agents (AREA)
- Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
Abstract
The present invention relates to a kind of fluorocarbon surfactant available for high flux numeral droplet PCR and its preparation method and application.The surfactant passes through formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite is combined in water phase and an oil phase interface electrostatic interaction and obtained：Wherein, x is that 2 ~ 15, y is that 15 ~ 45, z is that 2 ~ 15, n is 20 ~ 80, and the mass ratio of the cationic moiety and anionicsite is 1:1~16.Fluorocarbon surfactant provided by the invention available for high flux numeral droplet PCR passes through formula（I）And formula（II）Shown cationic moiety and anionicsite combines in water phase and an oil phase interface electrostatic interaction to be obtained, purity is high, and has good bio-compatibility and stability, can reduce the interfacial tension between water phase and an oil phase, prevent the fusion between drop, the drop stablized.In addition, the preparation method of the surfactant provided by the invention can obtain the fluorocarbon surfactant of high-purity, preparation process is simple.
Description
Technical field
The invention belongs to surfactant field, and in particular to a kind of fluorine carbon table available for high flux numeral droplet PCR
Face activating agent and its preparation method and application.
Background technology
In bioanalysis and application, oil-based liquid used has to meet several basic conditions simultaneously：Biological nothing
Toxicity, good permeability, hydrophobic, lipophobic.This conventional class I liquid I usually solvent containing carbon fluorine chain, the boundary of carbon fluorous solvent in itself
Face tension force is much lower compared with water.This proposes more harsh requirement to surfactant.First, surfactant allows for surely
Determine drop；Secondly, surfactant must be biologically inert, i.e., surfactant can form a biology in the inner surface of water droplet
Inert inner surface, the biotic component in aqueous phase will not be damaged；Finally, surfactant must also be nonionic table
Face activating agent, otherwise electrostatic interaction can occur with the opposite charges in DNA/RNA, albumen or other biological molecule.
Fluorocarbon surfactant is one of very important kind in special surfactant, in many special fields
（Such as fire-fighting, leather, electrical industry）, wherein there is the irreplaceable important use of many hc-surfactants（Such as：It is resistance to
More than 300 °C of high temperature, antistatic property etc.）.The special performance of fluorocarbon surfactant is often summarized as " three high " " two hate ",
That is high surface, high stability and high chemical stability；Fluorine-containing alkyl not only hydrophobic but also had hated oil.Fluorocarbon surfactant has very high
Heat resistance and good chemical stability, it can resist the effect of strong oxidizer, strong acid and highly basic, and in these solution
In be maintained to good surface-active, therefore can be used in the solution under some extreme conditions.Fluorine surfactant
It is only the 0.01 ~ 0.1 of hc-surfactant to reach with the concentration of the same surface tension of hc-surfactant.Fluorocarbon surface is lived
Property agent is in many industry extensive uses such as chemical industry, machinery, weaving, papermaking, coating, glass, ceramics, household supplies.
PCR（Polymer Chain Reaction, PCR）It is a kind of external simplified condition Imitating DNA
The method of the molecule rapid amplifying replicated in vivo.Micro- reaction that droplet type round pcr is reacted by the use of Water-In-Oil structure as PCR
Device, enter performing PCR amplification.The characteristics of droplet type PCR maximums is that can form the independent reaction space of huge number to enter performing PCR
Amplification, " microreactor " of each drop as its own PCR, does not produce cross pollution each other.Its key technology is
Before PCR reactions, the aqueous solution including reacted constituents all PCR is injected into the oil phase surface of rotation at a high speed, forms nothing
The stable small water droplet of number.Surfactant plays conclusive effect to the stability of small water droplet.
The fluorocarbon surfactant for being presently used for stablizing the microlayer model in droplet type PCR is mainly using chemical synthesis
Mode obtains, and main Types include PFPE or polyether block nonionic surfactant etc..For example business procurement is arrived
EA surfactants（Raindance, the U.S.）, Pico-Surf（Dolomite, Britain）Deng these surfactants all may be used
With for producing the drop with good bio-compatibility, and these drops still have when PCR is warming up to 95 °C it is good
Good stability.But the commercially produced product of these surfactants is not pure single substance, but with specific concentration
It has been dissolved in carbon fluorocarbon oil, can not have met that the concentration requirement to pure material and surfactant, especially PCR are anti-under specific condition
It should must avoid interference of the impurity to reaction.And the building-up process of pure fluorine surfactant it is comparatively laborious, it is necessary to multistep reaction and
In the strict purification of products stage, seriously hinder the extensive use and industrialization of this kind of surfactant.
Therefore, develop a kind of bio-compatibility and stability with high-purity, good fc-surfactant with
Important Research Significance and application value.
The content of the invention
It is an object of the invention to overcome the defects of fc-surfactant purity is low in the prior art, there is provided one kind is available
In high flux numeral droplet PCR fluorocarbon surfactant.Fluorocarbon surfactant purity provided by the invention is high, and with good
Good bio-compatibility and stability, the interfacial tension between water phase and an oil phase can be reduced, prevent the fusion between drop, obtained steady
Fixed drop.
For achieving the above object, the present invention adopts the following technical scheme that：
A kind of fluorocarbon surfactant available for high flux numeral droplet PCR, the surfactant pass through formula（Ⅰ）Shown sun
Ionic portions and formula（Ⅱ）Shown anionicsite obtains in water phase and an oil phase interface electrostatic interaction：
Wherein, x is that 2 ~ 15, y is that 15 ~ 45, z is that 2 ~ 15, n is 20 ~ 80, the mass ratio of the cationic moiety and anionicsite
For 1:1~16.
Formula provided by the invention（I）Shown cationic moiety and formula（II）Shown anionicsite has certain in itself
Surface-active, so when water-oil phase coexists, both hydrophilic and oleophilic head bases are respectively toward moving in two-phase.When anion portion
When the negative electrical charge divided encounters the positive charge of cationic moiety at water-oil interface, positive and negative charge passes through electrostatic force at two-phase interface
Interact, form fluorocarbon surfactant of the present invention.Fluorocarbon surfactant purity provided by the invention is high,
And there is good bio-compatibility and stability, it can reduce the interfacial tension between water phase and an oil phase, prevent melting between drop
Close, the drop stablized.
Preferably, the mass ratio of the cationic moiety and anionicsite is 1:2~10.It is further preferable that it is described sun from
The mass ratio of subdivision and anionicsite is 1:8.
Preferably, the x is that 2 ~ 12, y is that 20 ~ 40, z is that 2 ~ 10, n is 25 ~ 65.It is further preferable that the x is for 2, y
35, z 4, n 45.
A kind of preparation method of the above-mentioned fluorocarbon surfactant available for high flux numeral droplet PCR, methods described bag
Include following steps：
S1：By formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite is dissolved in water and carbon fluorocarbon oil respectively obtains aqueous phase
Fluid and oil phase fluid, the mass fraction of the aqueous phase fluid cationic part are 0.5 ~ 4%, in the oil phase fluid the moon from
The mass fraction of subdivision is 1% ~ 8%；
S2：The aqueous phase fluid obtained by S1 and oil phase fluid are injected into the corresponding stream in micro-fluidic microlayer model generation chip respectively
In road；
S3：The volume flow rate of oil phase fluid and aqueous phase fluid is adjusted, mixes and produces the fluorocarbon surfactant.
The building-up process of traditional perfluoropolyether surfactants is usually a two-step reaction, and yield is relatively low, and product
Purity hardly result in guarantee.Further, since solubility of the fluorochemical polyether in ordinary organic solvents is relatively low, and molecular weight is big,
The steric hindrance of active group carboxyl is relatively large, and in rear hydrocarbon reaction with amido, activity is low.And building-up process
In must carry out second step reaction again after first where acyl chlorides, and the stability of acyl chlorides is low, influences the purity of next step product.This hair
The preparation method of bright offer can be combined by electrostatic force combination zwitterion part, and the fluorocarbon surface for obtaining high-purity is lived
Property agent, preparation process are simple.
Preferably, biological reagent is also contained in the aqueous phase fluid, the dosage of biological reagent is 10 in the aqueous phase fluid
~500 nM。
The biological reagent that the present invention uses is the biological agent that in the market can be sold.
Preferably, the biological reagent is protein, hotstart master mix complexs primer, probe, DNA moulds
One or several kinds in plate or biology enzyme.
All kinds of perfluoro solvents that market can be sold can be used as carbon fluorocarbon oil to be used in the present invention.
Preferably, carbon fluorocarbon oil is in FC-40, FC-43, FC-770, HFE-7500 or HFE-7100 in the oil phase fluid
One or several kinds.
Preferably, the injection of oil phase fluid and aqueous phase fluid is realized in S2 by the way of pressure injection.
The volume flow rate of oil phase fluid and aqueous phase fluid is that conventional flow conditions are that can obtain the drop of suitable size in S3.
Preferably, the volume flow rate of oil phase fluid and aqueous phase fluid is respectively 100 ~ 1200 μ L/min and 50 ~ 800 in S3
μ L/min, the volume flow rate ratio of the oil phase fluid and aqueous phase fluid is 1 ~ 5:1.It is further preferable that the oil phase fluid and water
The volume flow rate of phase fluid is respectively 600 μ L/min and 300 μ L/min.
The above-mentioned fluorocarbon surfactant available for high flux numeral droplet PCR is in digital pcr or high flux biology divides
Application in analysis is also within the scope of the present invention.
Compared with prior art, the present invention has the advantages that：
Fluorocarbon surfactant provided by the invention available for high flux numeral droplet PCR passes through formula（I）And formula（II）It is shown
Cationic moiety and anionicsite combine and obtain in water phase and an oil phase interface electrostatic interaction, purity is high, and with good
Bio-compatibility and stability, can reduce the interfacial tension between water phase and an oil phase, prevent the fusion between drop, obtain stabilization
Drop.In addition, the preparation method of the surfactant provided by the invention can obtain the fluorocarbon surfactant of high-purity, system
Standby process is simple.
Brief description of the drawings
Fig. 1 is the Microfluidic droplet generating process schematic diagram for the surfactant that embodiment 1 provides；
Fig. 2 is the electrostatic interaction mistake between the surfactant cationic part that embodiment 1 provides and anionicsite
Journey schematic diagram；
Fig. 3 is the infrared spectrum of surfactant and positive and negative ionic portions that the embodiment of the present invention 1 provides；
Fig. 4 is drop microscope figure of the surfactant in different phase of the offer of the embodiment of the present invention 1.
Embodiment
The present invention is expanded on further with reference to embodiment.These embodiments are merely to illustrate the present invention rather than limitation
The scope of the present invention.The experimental method of unreceipted actual conditions in lower example embodiment, generally according to this area normal condition or is pressed
The condition suggested according to manufacturer；Used raw material, reagent etc., unless otherwise specified, being can be from the business such as conventional market
The raw material and reagent that approach obtains.The change for any unsubstantiality that those skilled in the art is done on the basis of the present invention
And replace and belong to scope of the present invention.
Embodiment 1 can be used for high flux numeral droplet PCR fluorocarbon surfactant
The surfactant that the present embodiment provides is by formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite is in water oil
Two-phase interface electrostatic interaction is combined and obtained：
Wherein, x, y, z, n are 2,35,4,45 respectively, and the mass ratio of cationic moiety and anionicsite is 1:8, by walking as follows
Suddenly it is prepared：
（1）By formula（I）Shown cationic moiety is dissolved in deionization by 1% mass fraction, as aqueous phase fluid；By formula
（II）Shown anionicsite is dissolved in HFE-7500 by 8% mass fraction, as oil phase fluid.
（2）Aqueous phase fluid and oil phase fluid are injected into pair in Microfluidic droplet generation chip respectively with pressure injection pump
In the runner answered（Such as Fig. 1）.
（3）Regulation oil, the volume flow rate and velocity ratio of water two-phase, obtain the drop that size is homogeneous, stablizes, such as Fig. 1.Wherein
Oil, the volume flow rate of water two-phase are respectively 600 μ L/min, 300 μ L/min, velocity ratio 2:1.
Electrostatic interaction mistake between the surfactant cationic part of the present embodiment offer and anionicsite
Journey schematic diagram is as shown in Figure 2：Formula（I）Shown cationic moiety and formula（II）Shown anionicsite has certain in itself
Surface-active, so when water-oil phase coexists, both hydrophilic and oleophilic head bases are respectively toward moving in two-phase.Work as anionicsite
Negative electrical charge when the positive charge of cationic moiety is encountered at water-oil interface, positive and negative charge is sent out at two-phase interface by electrostatic force
Raw interaction, forms fluorocarbon surfactant, and reduces the interfacial tension between two-phase, prevents melting between drop
Close, the drop stablized.
In order to contrast, we have synthesized anionicsite and cationic moiety covalent bond also by the mode of chemical synthesis
Material.Fig. 3 is the INFRARED SPECTRUM of surfactant provided by the invention, anionicsite and covalently bound surfactant
Figure, as seen from the figure, C=O peak position is in 1773 cm in the carboxylate radical COOH of raw material anionicsite-1Place.Pass through covalent bond
Surfactant amido link C=O peak position in 1684 cm-1Place.And the surfactant obtained by electrostatical binding
Carboxylate radical C=O peak position be displaced to 1715 cm-1Place, the peak position of primary amine is in 1645 cm-1Place, illustrates cation portion
Divide and a kind of new surfactant is formd by electrostatic interaction with anionicsite.
Stability test
This test to certain temperature, verifies the heat endurance of micro-fluidic gained drop by temperature programming.Temperature Programmed Processes can
Think and 95 °C be directly warming up to from room temperature or is raised to 55 °C from room temperature, 55 °C maintain 30 s after, after from 55 °C
72 °C are risen to, and maintains 30 s, then is warming up to 95 °C, and 60 s are maintained under 95 °C, above whole process repeats 35 times.
The drop that has just been collected into micro- sem observation, without the drop that placed 11 days of any processing and by program
Drop and its size after heating, judge its heat endurance.Related microscopy images figure such as Fig. 4.Understand, place 11 days and
Through above-mentioned thermal cycle processing after drop with it is initial when the basic indifference of drop, i.e., the present embodiment provide table inside activating agent tool
There is preferable heat endurance.
Embodiment 2 can be used for high flux numeral droplet PCR fluorocarbon surfactant
The surfactant that the present embodiment provides is by formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite is in water oil
Two-phase interface electrostatic force is combined and obtained：
Wherein, x, y, z, n are 2,20,10,65 respectively, and the mass ratio of cationic moiety and anionicsite is 1:1, by walking as follows
Suddenly it is prepared：
（1）By formula（I）Shown cationic moiety is dissolved in deionization by 1.5% mass fraction, as aqueous phase fluid；By formula
（II）Shown anionicsite is dissolved in HFE-7500 by 1.5% mass fraction, as oil phase fluid.
（2）Aqueous phase fluid and oil phase fluid will be injected into respectively in Microfluidic droplet generation chip with pressure injection pump
In corresponding runner.
（3）Regulation oil, the volume flow rate and velocity ratio of water two-phase, obtain the drop that size is homogeneous, stablizes.Wherein oil, water
The volume flow rate of two-phase is respectively 600 μ L/min, 500 μ L/min, velocity ratio 1.2:1.
Similarly, zwitterion part is interacted at two-phase interface by electrostatic force, forms fluoro-carbon surface active
Agent, and the interfacial tension between two-phase is reduced, the fusion between drop is prevented, the drop stablized.
Embodiment 3 can be used for high flux numeral droplet PCR fluorocarbon surfactant
The surfactant that the present embodiment provides is by formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite is in water oil
Two-phase interface electrostatic force is combined and obtained：
Wherein, x, y, z, n are 12,40,2,25 respectively, and the mass ratio of cationic moiety and anionicsite is 1:16, by as follows
Step is prepared：
（1）By formula（I）Shown cationic moiety is dissolved in deionization by 0.5% mass fraction, as aqueous phase fluid；By formula
（II）Shown anionicsite is dissolved in HFE-7500 by 8% mass fraction, as oil phase fluid.
（2）Aqueous phase fluid and oil phase fluid will be injected into respectively in Microfluidic droplet generation chip with pressure injection pump
In corresponding runner.
（3）Regulation oil, the volume flow rate and velocity ratio of water two-phase, obtain the drop that size is homogeneous, stablizes.Wherein oil, water
The volume flow rate of two-phase is respectively 800 μ L/min, 400 μ L/min, velocity ratio 2:1.
Similarly, zwitterion part is interacted at two-phase interface by electrostatic force, forms fluoro-carbon surface active
Agent, and the interfacial tension between two-phase is reduced, the fusion between drop is prevented, the drop stablized.
Embodiment 4 can be used for high flux numeral droplet PCR fluorocarbon surfactant
The surfactant that the present embodiment provides is by formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite is in water oil
Two-phase interface electrostatic force is combined and obtained：
Wherein, x, y, z, n are 2,15,15,80 respectively, and the mass ratio of cationic moiety and anionicsite is 1:2, by walking as follows
Suddenly it is prepared：
（1）By formula（I）Shown cationic moiety and bovine serum albumin respectively by 4% and 10 nM mass fraction be dissolved in deionization
In, as aqueous phase fluid；By formula（II）Shown anionicsite is dissolved in FC-40 by 8% mass fraction, as oil phase stream
Body.
（2）Aqueous phase fluid and oil phase fluid will be injected into respectively in Microfluidic droplet generation chip with pressure injection pump
In corresponding runner.
（3）Regulation oil, the volume flow rate and velocity ratio of water two-phase, obtain the drop that size is homogeneous, stablizes.Wherein oil, water
The volume flow rate of two-phase is respectively 1250 μ L/min, 250 μ L/min, velocity ratio 5:1.
Similarly, zwitterion part is interacted at two-phase interface by electrostatic force, forms fluoro-carbon surface active
Agent, and the interfacial tension between two-phase is reduced, the fusion between drop is prevented, the drop stablized.
Embodiment 5 can be used for high flux numeral droplet PCR fluorocarbon surfactant
The surfactant that the present embodiment provides is by formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite is in water oil
Two-phase interface electrostatic force is combined and obtained：
Wherein, x, y, z, n are 15,45,2,20 respectively, and the mass ratio of cationic moiety and anionicsite is 1:10, by as follows
Step is prepared：
（1）By formula（I）The DNA of shown cationic moiety and ROX- mark respectively by 0.5% and 500 nM concentration be dissolved in from
In son, as aqueous phase fluid；By formula（II）Shown anionicsite is dissolved in FC-43 by 5% mass fraction, as oil phase
Fluid.
（2）Aqueous phase fluid and oil phase fluid will be injected into respectively in Microfluidic droplet generation chip with pressure injection pump
In corresponding runner.
（3）Regulation oil, the volume flow rate and velocity ratio of water two-phase, obtain the drop that size is homogeneous, stablizes.Wherein oil, water
The volume flow rate of two-phase is respectively 120 μ L/min, 80 μ L/min, velocity ratio 3:2.
Similarly, zwitterion part is interacted at two-phase interface by electrostatic force, forms fluoro-carbon surface active
Agent, and the interfacial tension between two-phase is reduced, the fusion between drop is prevented, the drop stablized.
Claims (10)
1. a kind of fluorocarbon surfactant available for high flux numeral droplet PCR, it is characterised in that the surfactant leads to
Cross formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite combines in water phase and an oil phase interface electrostatic interaction
Arrive：
Wherein, x is that 2 ~ 15, y is that 15 ~ 45, z is that 2 ~ 15, n is 20 ~ 80, the mass ratio of the cationic moiety and anionicsite
For 1:1~16.
2. it can be used for high flux numeral droplet PCR fluorocarbon surfactant according to claim 1, it is characterised in that institute
The mass ratio for stating cationic moiety and anionicsite is 1:2~10.
3. it can be used for high flux numeral droplet PCR fluorocarbon surfactant according to claim 2, it is characterised in that institute
The mass ratio for stating cationic moiety and anionicsite is 1:8.
4. it can be used for high flux numeral droplet PCR fluorocarbon surfactant according to claim 1, it is characterised in that institute
It is that 2 ~ 12, y is that 20 ~ 40, z is that 2 ~ 10, n is 25 ~ 65 to state x.
5. it can be used for high flux numeral droplet PCR fluorocarbon surfactant according to claim 4, it is characterised in that institute
It is 2, y 35, z 4, n 45 to state x.
A kind of 6. preparation side of any fluorocarbon surfactant available for high flux numeral droplet PCR of claim 1 ~ 3
Method, it is characterised in that methods described comprises the following steps：
S1：By formula（Ⅰ）Shown cationic moiety and formula（Ⅱ）Shown anionicsite is dissolved in water and carbon fluorocarbon oil respectively obtains aqueous phase
Fluid and oil phase fluid, the mass fraction of the aqueous phase fluid cationic part are 0.5 ~ 4%, in the oil phase fluid the moon from
The mass fraction of subdivision is 1 ~ 8%；
S2：The aqueous phase fluid obtained by S1 and oil phase fluid are injected into the corresponding stream in micro-fluidic microlayer model generation chip respectively
In road；
S3：The volume flow rate of oil phase fluid and aqueous phase fluid is adjusted, mixes and produces the fluorocarbon surfactant.
7. can be used for the preparation method of high flux numeral droplet PCR fluorocarbon surfactant according to claim 4, it is special
Sign is, also contains biological reagent in the aqueous phase fluid, and the dosage of biological reagent is 10 ~ 500 nM in the aqueous phase fluid.
8. can be used for the preparation method of high flux numeral droplet PCR fluorocarbon surfactant according to claim 4, it is special
Sign is that carbon fluorocarbon oil is one kind in FC-40, FC-43, FC-770, HFE-7500 or HFE-7100 in the oil phase fluid.
9. can be used for the preparation method of high flux numeral droplet PCR fluorocarbon surfactant according to claim 4, it is special
Sign is, the volume flow rate of oil phase fluid and aqueous phase fluid is respectively 100 ~ 1200 μ L/min and 50 ~ 800 μ L/min in S3,
The volume flow rate ratio of the oil phase fluid and aqueous phase fluid is 1 ~ 5:1.
10. any fluorocarbon surfactant available for high flux numeral droplet PCR of claim 1 ~ 5 is in digital pcr
Or the application in high flux bioanalysis.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711015652.2A CN107754723B (en) | 2017-10-26 | 2017-10-26 | A kind of fluorocarbon surfactant and its preparation method and application can be used for high-throughput digital droplet PCR |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201711015652.2A CN107754723B (en) | 2017-10-26 | 2017-10-26 | A kind of fluorocarbon surfactant and its preparation method and application can be used for high-throughput digital droplet PCR |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN107754723A true CN107754723A (en) | 2018-03-06 |
| CN107754723B CN107754723B (en) | 2019-01-01 |
Family
ID=61271878
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201711015652.2A Active CN107754723B (en) | 2017-10-26 | 2017-10-26 | A kind of fluorocarbon surfactant and its preparation method and application can be used for high-throughput digital droplet PCR |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN107754723B (en) |
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110095386A (en) * | 2019-05-15 | 2019-08-06 | 中国石油大学(北京) | A kind of interaction prediction method and device between being adsorbed with the drop of surfactant |
| WO2019230566A1 (en) * | 2018-05-31 | 2019-12-05 | キヤノン株式会社 | Analysis system |
| CN112495316A (en) * | 2020-10-20 | 2021-03-16 | 大连理工大学 | Method for preparing micro-nano gel microspheres based on metastable emulsion |
| CN113559783A (en) * | 2021-07-16 | 2021-10-29 | 南京理工大学 | Fluorocarbon surfactant and preparation method and application thereof |
| CN114539536A (en) * | 2022-02-18 | 2022-05-27 | 华南师范大学 | Preparation method and application of perfluoropolyether surfactant containing amide bonds |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060210997A1 (en) * | 2005-03-16 | 2006-09-21 | Joel Myerson | Composition and method for array hybridization |
| CN106622017A (en) * | 2016-12-30 | 2017-05-10 | 广东顺德工业设计研究院(广东顺德创新设计研究院) | Fluorine surfactant and preparation method and application thereof |
| CN106823986A (en) * | 2017-01-20 | 2017-06-13 | 广东顺德工业设计研究院(广东顺德创新设计研究院) | Three-stage fluorine surfactant and its preparation method and application |
-
2017
- 2017-10-26 CN CN201711015652.2A patent/CN107754723B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060210997A1 (en) * | 2005-03-16 | 2006-09-21 | Joel Myerson | Composition and method for array hybridization |
| CN106622017A (en) * | 2016-12-30 | 2017-05-10 | 广东顺德工业设计研究院(广东顺德创新设计研究院) | Fluorine surfactant and preparation method and application thereof |
| CN106823986A (en) * | 2017-01-20 | 2017-06-13 | 广东顺德工业设计研究院(广东顺德创新设计研究院) | Three-stage fluorine surfactant and its preparation method and application |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2019230566A1 (en) * | 2018-05-31 | 2019-12-05 | キヤノン株式会社 | Analysis system |
| CN110095386A (en) * | 2019-05-15 | 2019-08-06 | 中国石油大学(北京) | A kind of interaction prediction method and device between being adsorbed with the drop of surfactant |
| CN112495316A (en) * | 2020-10-20 | 2021-03-16 | 大连理工大学 | Method for preparing micro-nano gel microspheres based on metastable emulsion |
| CN112495316B (en) * | 2020-10-20 | 2021-11-19 | 大连理工大学 | Method for preparing micro-nano gel microspheres based on metastable emulsion |
| CN113559783A (en) * | 2021-07-16 | 2021-10-29 | 南京理工大学 | Fluorocarbon surfactant and preparation method and application thereof |
| CN113559783B (en) * | 2021-07-16 | 2022-09-13 | 南京理工大学 | Fluorocarbon surfactant and preparation method and application thereof |
| CN114539536A (en) * | 2022-02-18 | 2022-05-27 | 华南师范大学 | Preparation method and application of perfluoropolyether surfactant containing amide bonds |
Also Published As
| Publication number | Publication date |
|---|---|
| CN107754723B (en) | 2019-01-01 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN107754723A (en) | A kind of fluorocarbon surfactant available for high flux numeral droplet PCR and its preparation method and application | |
| US9012390B2 (en) | Fluorocarbon emulsion stabilizing surfactants | |
| Litschel et al. | Protein reconstitution inside giant unilamellar vesicles | |
| Pawar et al. | Fabrication, assembly, and application of patchy particles | |
| Song et al. | All-aqueous multiphase microfluidics | |
| CN101076417B (en) | Method for preparing metal nanometer structure | |
| CN107930542A (en) | One-step method continuously prepares the microflow control technique of calcium alginate microgel | |
| CN108441541A (en) | Composition, the method and system measured for polymerase chain reaction | |
| Pereira et al. | Aqueous two-phase systems | |
| CN109310955A (en) | The equal Porous materials of electrification for electrostatic separation | |
| Miyazaki et al. | Preparation of functionalized nanostructures on microchannel surface and their use for enzyme microreactors | |
| Inaba et al. | Light-induced propulsion of a giant liposome driven by peptide nanofibre growth | |
| Chang et al. | Bifunctional janus spheres with chemically orthogonal patches | |
| Quevedo et al. | Electrokinetic characterization of synthetic protein nanoparticles | |
| Lin et al. | Stabilizing aqueous three-dimensional printed constructs using chitosan-cellulose nanocrystal assemblies | |
| US7465381B2 (en) | Electrokinetic molecular separation in nanoscale fluidic channels | |
| Chen et al. | Temperature and pH responsive Janus silica nanoplates prepared by the Sol–Gel process and postmodification | |
| Toyota et al. | Chemically artificial rovers based on self-propelled droplets in micrometer-scale environment | |
| Khan et al. | Molecule (s) of Interest: I. Ionic Liquids–Gateway to Newer Nanotechnology Applications: Advanced Nanobiotechnical Uses’, Current Status, Emerging Trends, Challenges, and Prospects | |
| Street | Highly selective separations in biotechnology | |
| CN113351110A (en) | Temperature-sensitive fluorocarbon surfactant and preparation method and application thereof | |
| US20110003155A1 (en) | Polyurethane nanometer powder of which surface is modified with phosphoric acid and its preparation method | |
| Phillips et al. | Separations in poly (dimethylsiloxane) microchips coated with supported bilayer membranes | |
| Valentini et al. | Microorganism nutrition processes as a general route for the preparation of bionic nanocomposites based on intractable polymers | |
| JP2008185530A (en) | Gel composition for separation and recovery, and separation/recovery method using the same |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |