CN109187450A - A kind of biomolecule concentration detection method based on quantum dot - Google Patents
A kind of biomolecule concentration detection method based on quantum dot Download PDFInfo
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- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6402—Atomic fluorescence; Laser induced fluorescence
Abstract
The biomolecule concentration detection method based on quantum dot that the invention discloses a kind of, the following steps are included: by the antibody coupling of quantum dot and the target biological molecules for needing to detect, make quantum dot-antibody in conjunction with target biological molecules with the solution to be measured mixed culture containing target biological molecules quantum dot-antibody stoste, by mixed solution by microchannel, control mixed solution flow makes a quantum dot-antibody or quantum dot-antibody-biomolecule pass through the microchannel；Measurement distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule by quantum dot-antibody or quantum dot-antibody-biomolecule time resolution single photon characteristic spectrum of the microchannel；Calculate the concentration of target biological molecules in solution to be measured.This method can detecte out the biomolecule of fM concentration, and detects speed and be largely increased.
Description
Technical field
The present invention relates to a kind of biomolecule concentration detection methods, more particularly to a kind of biomolecule based on quantum dot
Concentration detection method.
Background technique
The high speed development of society makes people's living standard be greatly improved, and living-hygienic, disease forecasting etc. are mentioned
Gone out new requirement, however it is existing biomolecule detection means are difficult to realize quickly to identify measurement, therefore such as drinking
Bacterial monitoring in water, bacterial drug resistance investigation, identification and quantization of cancer biomarkers object etc. are difficult in practical application
In broken through.The prior art be used for identify and quantify low concentration biomolecule method include:
Liquid chromatogram (Liquid chromatography) and mass spectrum (mass spectrometry) (LC-MS): LC exists
The component being physically separated in liquid solution；The structure that MS provides each component by measuring the mass-to-charge ratio of each group segregant is special
Property.Enzyme-linked immunosorbent assay (ELISA): can be used the immunoassay based on antibody, as ELISA comes in test sample to resist
Former presence.In order to quantify the antigen in solution, antigen is first by being directly adsorbed to surface or using be coated in surface
On " capture antibody " be fixed in the surface of solids (polystyrene microtiter plates).Then the specificity connecting with enzyme is added just
Grade antibody is to combine immobilized antigen.It is eventually adding the substrate of enzyme, color changes when with enzyme reaction.
Western blotting: protein lytic cell and tissue in detection and quantization usually by Western blotting into
Row, the trace mainly comprise the steps that sample preparation, gel/transfer/blocking, primary antibody combines and signal detection.
Flow cytometry, fluorescence in situ hybridization (FISH), polymerase chain reaction (PCR): the identification of microorganism is usual in water
Using molecules such as flow cytometry, magnetic separation technique, fluorescence in situ hybridization (FISH), polymerase chain reaction (PCR) and microarrays
Technology.
Also have and recognition detection is carried out using technology of quantum dots, quantum dot (quantum dot, be abbreviated as QD) has been able to work
Industry metaplasia produces, and can be coupled with antibody (being abbreviated as Ab) by surface modification quantum dot and form quantum dot-antibody (QD-Ab) coupling
Body is coupled by the Ab on QD-Ab zygosome with biomolecule to be detected targeting.Wherein irradiation of the quantum dot in outer exciting light
Under, emit fluorescence, to demarcate targeting biological molecules.In the prior art, pass through detection quantum dot fluorescence intensity or wavelength of fluorescence
Variation is to distinguish QD-Ab- biomolecule and QD-Ab zygosome, to realize the purpose of detection targeting biological molecules.But in theory
On, the equal very little of variation of fluorescence intensity and wavelength of the QD-Ab before and after being coupled with biomolecule.And it is answered in practical optical measurement
It is difficult to realize to the quantitative measurment of the minor change of fluorescence intensity and wavelength of fluorescence in.Meanwhile quantum dot synthesis is by many realities
Condition influence is tested, is difficult to find fluorescence intensity and the identical quantum dot of wavelength with the quantum dot of a batch growth, in
It is the quantum dot for biomolecule inspection per se with the inhomogeneities of fluorescence intensity and wavelength, this is to utilize quantum dot fluorescence
The variation of intensity and wavelength brings unsurmountable difficulty to detect the realization of the instrument of biomolecule.So being based on this method
The instrument of quanta point biological detection apply for a patent from 2000 or so and just have proposition, but be all not carried out such instrument till now
The marketization of device, this is from illustrating on the other hand through the fluorescence intensity and wave on measurement quantum point coupling after biomolecule
Long variation is difficult to carry out quantitative judge to QD-Ab- biomolecule.
Summary of the invention
In view of the above-mentioned defects in the prior art, the present invention provides a kind of biomolecule concentration detection side based on quantum dot
Method solves the problems, such as QD-Ab- biomolecule and the identification of QD-Ab, realizes the fast quantification identification of low concentration biomolecule.
Technical solution of the present invention is as follows: a kind of biomolecule concentration detection method based on quantum dot, comprising the following steps:
The antibody coupling of S1, the target biological molecules for detecting quantum dot and needs, form quantum dot-antibody stoste；
S2, quantum dot-antibody stoste and the solution to be measured mixed culture containing target biological molecules are made into quantum dot-antibody
In conjunction with target biological molecules, obtain containing quantum dot-antibody and quantum dot-antibody-biomolecule mixed solution；
S3, the mixed solution for obtaining step 2 control mixed solution flow and make a quantum dot-by microchannel
Antibody or quantum dot-antibody-biomolecule pass through the microchannel；
S4, measurement pass through quantum dot-antibody of the microchannel or quantum dot-antibody-biomolecule time point
It distinguishes single photon characteristic spectrum, distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule；
Mixed solution flowing time when S5, statistics quantum dot-antibody-biomolecule amount to concentration effectively identify quantity,
Mixed liquor volume is obtained by mixed solution flowing time and mixed solution flow velocity, by quantum dot-antibody-biomolecule amount and
Mixed liquor volume calculates the concentration of target biological molecules in solution to be measured.
Further, the step S4 distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule is according to continuous light
The photon pulse sequence that the photon pulse sequence and excitation quantum dot-antibody-biomolecule for exciting quantum dot-antibody to generate generate
Column, the pulse width probability density for calculating fluorescent pulse distinguish.
Further, the step S4 distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule is according to continuous light
The photon pulse sequence that the photon pulse sequence and excitation quantum dot-antibody-biomolecule for exciting quantum dot-antibody to generate generate
Column calculate fluorescent pulse correlation spectrum and distinguish.
Further, the step S4 distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule is according to continuous light
The photon pulse sequence that the photon pulse sequence and excitation quantum dot-antibody-biomolecule for exciting quantum dot-antibody to generate generate
Column, the pulse spacing probability density for calculating fluorescent pulse distinguish.
Further, it is described calculate fluorescent pulse width probability density distinguish be to the photon pulse sequence into
Ms grades of the row pulse width probability density for sampling and calculating the fluorescent pulse that pulse width is 1ms or more distinguish.
Further, it is to carry out 10 to the photon pulse sequence that the calculating fluorescent pulse correlation spectrum, which distinguishes,-7S grades
It samples and is calculated by the following formula association
It is wherein gA(τ) relating value, IAIt (t) is time resolution fluorescence spectral light intensity, τ is lag time,<>tIt is to the time
It averages, lag time 10-7~100s。
Further, it is to the photon pulse sequence that the pulse spacing probability density for calculating fluorescent pulse, which distinguishes,
Ns grades of column progress, which samples and calculates pulse spacing probability density of the pulse spacing between 1~100ns, to be distinguished.
Further, the photon pulse sequence and excitation quantum dot-that the continuous light excitation quantum dot-antibody of the basis generates
The photon pulse sequence that antibody-biomolecule generates is to excite quantum dot-antibody to produce in a time span according to continuous light
The photon pulse sequence that raw photon pulse sequence and excitation quantum dot-antibody-biomolecule generates in a time span,
The pulse train time span is 1ms~1s.
Preferably, the antibody coupling of the target biological molecules that quantum dot and needs are detected is to utilize ethyl -3- [3-
Dimethylaminopropyl] antibody and quantum dot is directly coupled by carbodiimides hydrochloride, and quantum dot surface therein is carboxylic
Base activation, antibody surface are amino-reactives, form amido bond after quantum dot and antibody coupling.
Preferably, the antibody coupling of the target biological molecules that quantum dot and needs are detected is to utilize succinimide
Base 4- [N- maleimidomehyl] cyclohexane -1- carboxylate coupling reaction, which carries out amino and the covalent reaction of sulfydryl, keeps sulfydryl living
The quantum dot of change and the antibody coupling of amino-reactive.
Further, comprising the following steps:
The antibody coupling of S1, the plurality of target biomolecule for detecting a variety of quantum dots with needs respectively, make same amount
Son point and same antibody coupling, form a variety of quantum dots-antibody stoste；
S2, a variety of quantum dots-antibody stoste and the solution to be measured containing plurality of target biomolecule are mixed the amount of making
Sub- point-antibody obtains in conjunction with target biological molecules containing a variety of quantum dot-antibody and quantum dot-antibody-biomolecule
Mixed solution；
S3, the mixed solution for obtaining step 2 control mixed solution flow and make a quantum dot-by microchannel
Antibody or quantum dot-antibody-biomolecule pass through the microchannel；
S4, measurement pass through quantum dot-antibody of the microchannel or quantum dot-antibody-biomolecule time point
It distinguishes single photon characteristic spectrum, distinguishes different quantum dot-antibody and different quantum dot-antibody-biomolecule；
When mixed solution flows when S5, statistics single quantum dot-antibody-biomolecule amount to concentration effectively identify quantity
Between, mixed liquor volume is obtained by mixed solution flowing time and mixed solution flow velocity, by this kind of quantum dot-antibody-biology point
Subnumber amount and mixed liquor volume calculate this kind of quantum dot-corresponding target biological molecules of antibody-biomolecule in solution to be measured
Concentration.
The advantages of technical solution provided by the present invention, is: distinguishing quantum dot-with time resolution single photon characteristic spectrum
Antibody and quantum dot-antibody-biomolecule can be avoided individual of the differentiation mode because of quantum dot of fluorescence intensity and wavelength of fluorescence
Identification error caused by difference, while also avoiding too small fluorescence intensity and being asked with what wavelength of fluorescence difference caused to be difficult to differentiate between
Topic.Quantum dot-antibody is distinguished with time resolution single photon characteristic spectrum and quantum dot-antibody-biomolecule can be extremely short
Discrimination differentiation is carried out in time, therefore is suitable for the measurement of micro biomolecule concentration liquid, and a few hours can detect that fM is dense
The biomolecule of degree, detection speed greatly improve.
Detailed description of the invention
Fig. 1 is the biomolecule concentration detection method flow diagram based on quantum dot.
Fig. 2 is continuous light activated quantum dot-antibody and quantum dot-antibody-biomolecule photon pulse sequence chart.
Fig. 3 is quantum dot-antibody and quantum dot-antibody-biomolecule photon pulse sequence pulse width probability
Density map.
Fig. 4 is quantum dot-antibody and quantum dot-antibody-biomolecule photon pulse sequence correlation spectrum.
Fig. 5 is quantum dot-antibody and quantum dot-antibody-biomolecule photon pulse sequence pulse width interval probability
Density map.
Fig. 6 is green quantum dot-antibody-biomolecule and red quantum dot-antibody-bio-molecule ratio is 7.5:2.5
When photon pulse sequence correlation spectrum.
Fig. 7 is green quantum dot-antibody-biomolecule and red quantum dot-antibody-bio-molecule ratio is 2.5:7.5
When photon pulse sequence correlation spectrum.
Specific embodiment
Below with reference to embodiment, the invention will be further described, it should be understood that these embodiments be merely to illustrate the present invention and
It is not used in and limits the scope of the invention, after the present invention has been read, those skilled in the art are to various equal similar shapes of the invention
The modification of formula is fallen in the application range as defined in the appended claims.
Incorporated by reference to shown in Fig. 1, embodiment 1
Biomolecule concentration detection method based on quantum dot involved in the present embodiment, comprising the following steps:
S1, it is detected by different crosslinking agent and needs according to water-soluble quantum dot QD surface modification (carboxyl, sulfydryl)
The antibody coupling of target biological molecules:
It is using EDC (ethyl -3- [3- dimethylaminopropyl] carbodiimides hydrochloride) that antibody and quantum dot is straight
Coupling is connect, quantum dot surface therein is activated carboxylic, and antibody surface is amino-reactive, amido bond is formed after the two coupling,
Form quantum dot-antibody stoste.
S2, quantum dot-antibody stoste and the solution to be measured containing target biological molecules are stirred 20 minutes, mixed culture makes
Quantum dot-antibody obtains mixed containing quantum dot-antibody and quantum dot-antibody-biomolecule in conjunction with target biological molecules
Close solution；
S3, the mixed solution for obtaining step 2 control mixed solution flow and make a quantum dot-by microchannel
Antibody or quantum dot-antibody-biomolecule pass through microchannel；
Quantum dot-antibody or quantum dot-antibody-biomolecule time resolution list that S4, measurement pass through microchannel
Photon characteristic spectrum, the photon pulse sequence for exciting quantum dot-antibody to generate at ms grades according to continuous light and excitation quantum dot-are anti-
The photon pulse sequence that body-biomolecule is generated at ms grades, as shown in Fig. 2, sampling time length is 5.2ms.Ms grades are carried out to adopt
Sample and the pulse width probability density for calculating the fluorescent pulse that pulse width is 1ms or more, obtain as shown in Figure 3 as a result, by phase
With width probability density, the different of corresponding pulse width carry out area to quantum dot-antibody or quantum dot-antibody-biomolecule
Point.
Mixed solution flowing time when S5, statistics quantum dot-antibody-biomolecule amount to concentration effectively identify quantity,
General concentration effectively identifies that quantity is 600 times, obtains mixed liquor volume by mixed solution flowing time and mixed solution flow velocity,
Target biological molecules in solution to be measured are calculated divided by mixed liquor volume by the quantum dot-antibody-biomolecule amount counted
Concentration.
Embodiment 2
Biomolecule concentration detection method based on quantum dot involved in the present embodiment, comprising the following steps:
S1, it is detected by different crosslinking agent and needs according to water-soluble quantum dot QD surface modification (carboxyl, sulfydryl)
The antibody coupling of target biological molecules:
Using SMCC, (succinimido 4- [N- maleimidomehyl] cyclohexane -1- carboxylate Ou Lian is reacted mercapto
The quantum dot of base activation and the antibody coupling of amino-reactive, carry out the covalent reaction coupled antibody and quantum dot of amino and sulfydryl
At quantum dot-antibody stoste.
S2, quantum dot-antibody stoste and the solution to be measured containing target biological molecules are stirred 20 minutes, mixed culture makes
Quantum dot-antibody obtains mixed containing quantum dot-antibody and quantum dot-antibody-biomolecule in conjunction with target biological molecules
Close solution；
S3, the mixed solution for obtaining step 2 control mixed solution flow and make a quantum dot-by microchannel
Antibody or quantum dot-antibody-biomolecule pass through microchannel；
Quantum dot-antibody or quantum dot-antibody-biomolecule time resolution list that S4, measurement pass through microchannel
Photon characteristic spectrum excites quantum dot-antibody 10 according to continuous light-7The photon pulse sequence and excitation quantum dot-of s grades of generations
Antibody-biomolecule is 10-7The photon pulse sequence of s grades of generations,.Carry out 10-7S grades sample and are calculated by the following formula pass
Connection
It is wherein gA(τ) relating value, IAIt (t) is time resolution fluorescence spectral light intensity, τ is lag time,<>tIt is to the time
It averages, lag time 10-7~100S is obtained as shown in Figure 4 as a result, by the difference of correlation peaks and half-peak breadth to quantum
Point-antibody or quantum dot-antibody-biomolecule distinguish.
Mixed solution flowing time when S5, statistics quantum dot-antibody-biomolecule amount to concentration effectively identify quantity,
General concentration effectively identifies that quantity is 600 times, obtains mixed liquor volume by mixed solution flowing time and mixed solution flow velocity,
Target biological molecules in solution to be measured are calculated divided by mixed liquor volume by the quantum dot-antibody-biomolecule amount counted
Concentration.
Embodiment 3
Biomolecule concentration detection method based on quantum dot involved in the present embodiment, comprising the following steps:
S1, it is detected by different crosslinking agent and needs according to water-soluble quantum dot QD surface modification (carboxyl, sulfydryl)
The antibody coupling of target biological molecules:
Using SMCC, (succinimido 4- [N- maleimidomehyl] cyclohexane -1- carboxylate Ou Lian is reacted mercapto
The quantum dot of base activation and the antibody coupling of amino-reactive, carry out the covalent reaction coupled antibody and quantum dot of amino and sulfydryl
At quantum dot-antibody stoste.
S2, quantum dot-antibody stoste and the solution to be measured containing target biological molecules are stirred 20 minutes, mixed culture makes
Quantum dot-antibody obtains mixed containing quantum dot-antibody and quantum dot-antibody-biomolecule in conjunction with target biological molecules
Close solution；
S3, the mixed solution for obtaining step 2 control mixed solution flow and make a quantum dot-by microchannel
Antibody or quantum dot-antibody-biomolecule pass through microchannel；
Quantum dot-antibody or quantum dot-antibody-biomolecule time resolution list that S4, measurement pass through microchannel
Photon characteristic spectrum, the photon pulse sequence for exciting quantum dot-antibody to generate at ns grades according to continuous light and excitation quantum dot-are anti-
The photon pulse sequence that body-biomolecule is generated in ns.0.1ns grades are carried out to sample and calculate the pulse spacing between 1~100ns
Pulse spacing probability density obtains as shown in Figure 5 as a result, the difference for the half-peak breadth composed by pulse spacing probability density is to quantum
Point-antibody or quantum dot-antibody-biomolecule distinguish.
Mixed solution flowing time when S5, statistics quantum dot-antibody-biomolecule amount to concentration effectively identify quantity,
General concentration effectively identifies that quantity is 600 times, obtains mixed liquor volume by mixed solution flowing time and mixed solution flow velocity,
Target biological molecules in solution to be measured are calculated divided by mixed liquor volume by the quantum dot-antibody-biomolecule amount counted
Concentration.
Troponin I is that the cardiopathic crucial heart of clinical diagnosis and protein level of skeletal muscle detect 5-10 and receive as example
The Troponin I (ca24kDa) of grams per liter, this level are used to indicate the increase of myocardial damage probability.1pM is added in every 1nl blood sample
QD antibody, then be incubated for 20 minutes, respectively measure 1nL and 1000nL mixed liquor, it is as a result as follows:
Using the method for the present invention, various biomolecules can also be measured simultaneously, using different quantum dot (the present embodiment
For red and green quantum dot) antibody coupling with different biological molecules, quantum dot-antibody stoste is divided with target organism is contained
The solution to be measured of son stirs 20 minutes, and mixed culture makes quantum dot-antibody in conjunction with target biological molecules, obtains containing quantum
Point-antibody and quantum dot-antibody-biomolecule mixed solution；Control mixed solution flow simultaneously makes a quantum dot-antibody
Or quantum dot-antibody-biomolecule passes through the microchannel；Quantum dot-the antibody or amount that measurement passes through microchannel
Sub- point-antibody-biomolecule time resolution single photon characteristic spectrum, the light for exciting quantum dot-antibody to generate according to continuous light
The photon pulse sequence that series of sub-pulses and excitation quantum dot-antibody-biomolecule generate, calculates the association of photon pulse sequence
Spectrum.Sub- point-antibody-the life of two amounts when red quantum dot and green quantum dot different proportion (7.5:2.5 and 2.5:7.5) mix
The correlation spectrum of the photon pulse sequence of object molecule is as shown in Figure 6 and Figure 7, and green quantum dot is small and light, and diffusion coefficient is big, emits light
Lag time on the correlation spectrum of series of sub-pulses is shorter than red quantum dot.Just by comparing photon lag time associated diagram
The type and quantity of biomolecule corresponding to different quantum dots can be distinguished.
Claims (10)
1. a kind of biomolecule concentration detection method based on quantum dot, which comprises the following steps:
The antibody coupling of S1, the target biological molecules for detecting quantum dot and needs, form quantum dot-antibody stoste；
S2, quantum dot-antibody stoste and the solution to be measured mixed culture containing target biological molecules are made into quantum dot-antibody and mesh
It marks biomolecule to combine, obtain containing quantum dot-antibody and quantum dot-antibody-biomolecule mixed solution；
S3, the mixed solution for obtaining step 2 control mixed solution flow and make a quantum dot-antibody by microchannel
Or quantum dot-antibody-biomolecule passes through the microchannel；
Quantum dot-antibody or quantum dot-antibody-biomolecule time resolution list that S4, measurement pass through the microchannel
Photon characteristic spectrum distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule；
Mixed solution flowing time when S5, statistics quantum dot-antibody-biomolecule amount to concentration effectively identify quantity, by mixing
It closes solution flowing time and mixed solution flow velocity obtains mixed liquor volume, by quantum dot-antibody-biomolecule amount and mixing
Liquor capacity calculates the concentration of target biological molecules in solution to be measured.
2. the biomolecule concentration detection method according to claim 1 based on quantum dot, which is characterized in that the step
S4 distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule is the photon for exciting quantum dot-antibody to generate according to continuous light
The photon pulse sequence that pulse train and excitation quantum dot-antibody-biomolecule generate, the pulse width for calculating fluorescent pulse are general
Rate density distinguishes.
3. the biomolecule concentration detection method according to claim 1 based on quantum dot, which is characterized in that the step
S4 distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule is the photon for exciting quantum dot-antibody to generate according to continuous light
The photon pulse sequence that pulse train and excitation quantum dot-antibody-biomolecule generate, calculates fluorescent pulse correlation spectrum and carries out area
Point.
4. the biomolecule concentration detection method according to claim 1 based on quantum dot, which is characterized in that the step
S4 distinguishes quantum dot-antibody and quantum dot-antibody-biomolecule is the photon for exciting quantum dot-antibody to generate according to continuous light
The photon pulse sequence that pulse train and excitation quantum dot-antibody-biomolecule generate, the pulse spacing for calculating fluorescent pulse are general
Rate density distinguishes.
5. the biomolecule concentration detection method according to claim 2 based on quantum dot, which is characterized in that the calculating
It is to carry out ms grades to the photon pulse sequence to sample and calculate pulse width that the width probability density of fluorescent pulse, which distinguishes,
Pulse width probability density for the fluorescent pulse of 1ms or more distinguishes.
6. the biomolecule concentration detection method according to claim 3 based on quantum dot, which is characterized in that the calculating
It is to carry out 10 to the photon pulse sequence that fluorescent pulse correlation spectrum, which distinguishes,-7S grades sample and are calculated by the following formula pass
Connection
It is wherein gA(τ) relating value, IAIt (t) is time resolution fluorescence spectral light intensity, τ is lag time,<>tIt is that flat is asked to the time
Mean value, lag time 10-7~100s。
7. the biomolecule concentration detection method according to claim 4 based on quantum dot, which is characterized in that the calculating
It is to carry out 0.1ns grades to the photon pulse sequence to sample and calculate arteries and veins that the pulse spacing probability density of fluorescent pulse, which distinguishes,
The pulse spacing probability density being divided between 1~100ns between punching distinguishes.
8. the biomolecule concentration detection method based on quantum dot according to any one of claim 2,3 and 4, special
Sign is that the continuous light of basis excites quantum dot-antibody generation photon pulse sequence and excitation quantum dot-antibody-biology
The photon pulse sequence that molecule generates is the photon for exciting quantum dot-antibody to generate in a time span according to continuous light
The photon pulse sequence that pulse train and excitation quantum dot-antibody-biomolecule generate in a time span, the pulse
Sequence time length is 1ms~1s.
9. the biomolecule concentration detection method according to claim 1 based on quantum dot, which is characterized in that described to measure
The antibody coupling for the target biological molecules that son point is detected with needs is sub- using ethyl -3- [3- dimethylaminopropyl] carbonization two
Antibody and quantum dot are directly coupled by amine salt acidulants, and quantum dot surface therein is activated carboxylic, and antibody surface is that amino is living
Change, forms amido bond after quantum dot and antibody coupling；Either utilize succinimido 4- [N- maleimidomehyl] ring
Ethane -1- carboxylate coupling reaction, which carries out amino and the covalent reaction of sulfydryl, makes the anti-of sulfhydryl activated quantum dot and amino-reactive
Body coupling.
10. a kind of biomolecule concentration detection method based on quantum dot, which comprises the following steps:
The antibody coupling of S1, the plurality of target biomolecule for detecting a variety of quantum dots with needs respectively, make same quantum dot
With same antibody coupling, a variety of quantum dots-antibody stoste is formed；
S2, a variety of quantum dots-antibody stoste and the solution to be measured mixed culture containing plurality of target biomolecule are made into quantum dot-
Antibody obtains in conjunction with target biological molecules containing a variety of quantum dot-antibody and quantum dot-antibody-biomolecule mixing
Solution；
S3, the mixed solution for obtaining step 2 control mixed solution flow and make a quantum dot-antibody by microchannel
Or quantum dot-antibody-biomolecule passes through the microchannel；
Quantum dot-antibody or quantum dot-antibody-biomolecule time resolution list that S4, measurement pass through the microchannel
Photon characteristic spectrum distinguishes different quantum dot-antibody and different quantum dot-antibody-biomolecule；
Mixed solution flowing time when S5, statistics single quantum dot-antibody-biomolecule amount to concentration effectively identify quantity,
Mixed liquor volume is obtained by mixed solution flowing time and mixed solution flow velocity, by this kind of quantum dot-antibody-biomolecule number
It measures and calculates the dense of the corresponding target biological molecules of this kind of quantum dot-antibody-biomolecule in solution to be measured with mixed liquor volume
Degree.
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| CN110579454A (en) * | 2019-08-19 | 2019-12-17 | 陈云 | fluorescent group identification method and device based on electric field modulation fluorescence correlation spectroscopy |
| CN115167798A (en) * | 2022-07-26 | 2022-10-11 | 惠科股份有限公司 | Quantum dot optical simulation method, device and system and computer readable storage medium |
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