CN106124783B - Coated nanometer magnetic bead of modified cuorin and preparation method thereof - Google Patents
Coated nanometer magnetic bead of modified cuorin and preparation method thereof Download PDFInfo
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- CN106124783B CN106124783B CN201610512711.6A CN201610512711A CN106124783B CN 106124783 B CN106124783 B CN 106124783B CN 201610512711 A CN201610512711 A CN 201610512711A CN 106124783 B CN106124783 B CN 106124783B
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Abstract
The invention discloses a kind of modified coated nanometer magnetic bead of cuorin and preparation method thereof, the modified coated nanometer magnetic bead of cuorin includes：Modified cuorin, biotin derivative and Streptavidin MagneSphere；Modified cuorin is thus connected to together with biotin derivative by CO NH structures；Streptavidin MagneSphere is the nanometer magnetic bead for combining Streptavidin, and biotin derivative is connected with Streptavidin.Modified cuorin is directly securely firmly connect with biotin derivative by this coated nanometer magnetic bead of modified cuorin by chemical bond, and biotin derivative is connected with the Streptavidin on Streptavidin MagneSphere surface.This coated nanometer magnetic bead of modified cuorin is used directly for the detection of anti-phospholipid antibody, and the detection product prepared relative to the method that conventional physical adsorbs cuorin, has higher stability.
Description
Technical field
The present invention relates to vitro detection field more particularly to a kind of coated nanometer magnetic bead of modified cuorin and its preparation sides
Method.
Background technology
Anticardiolipin antibodies are a kind of antibody that can be reacted with a variety of antigenic substances containing structure of phospholipid, and antigen is
Participate in the negatively charged phospholipid composition of various kinds of cell film composition.Clinically be primarily present in antiphospholipid syndrome, it is various itself
Immunity disease and syphilis patient's body.
Anti- phospholipid syndrome (APS) includes itself a variety of property immunity disease, is more common in young man, men and women's incidence rate is about
It is 2：8.Patient may occur in which one or more performances, can involve multiple systems, organ, mainly have：Vein and Arterial thrombosis,
Thrombopenia, habitual abortion, cardiomyopathy, heart disease, brain and kidney infraction, pulmonary hypertension.Pernicious APS can behave as short
The extensive thrombosis of progressive in phase causes multiple organ failure even dead.Can secondary to systemic loupus erythematosus or
Other autoimmunity diseases, but (primary antiphospholipid syndrome) also can individually occur.
Also anti-phospholipid antibody can be generated in syphilitic's body, syphilis is the property caused by spirochete bacteria Tyreponema pallidum
It spreads the disease.It is reported that 5 million people's syphilizations are had more than every year, including 30,000 babies by heredo infection.
Syphilis can be latent in patient's body and hides many years, and can lead to various clinical manifestations.Wherein hidden in syphilis
The patient of phase has no clinical symptoms, and the secondary stage does not treat even lifelong in patient about 2/3rds.The infected latent phase not
With infectiousness, however child's may be infected congenital syphilis of the Mothers in latent phase.
Anti-phospholipid antibody detection experiment is widely used in the non-treponema experiment of phosphatide syndrome diagnosis and syphilis.
The detection has the advantages that cheap, quick and is conveniently executed to great amount of samples.Further, since the concentration of anti-phospholipid antibody can be with
The treatment of syphilis is successful and continuously decreases, and the specific antibody treponema antibody of syphilis can last for several years even throughout one's life
It is all high.Therefore anti-phospholipid antibody detection experiment is considered as monitoring the finer selection of syphilis treatment.
The main method of traditional detection anti-phospholipid antibody is to be applied to the mode of cuorin physical absorption specifically
Above solid such as ELISA Plate, the cuorin being attached to above ELISA Plate and the anti-phospholipid antibody knot in sample to be detected are then utilized
It closes, realizes the capture to the anti-phospholipid antibody in sample.It is developed the color by the anti-phospholipid antibody to capture and measures luminous concentration, just
The anti-phospholipid antibody concentration in sample can indirectly be read.
Due to being easy from solid panel under the influence of the external conditions such as solvent, heating with the fixed cuorin of physical adsorption way
It is upper to dissociate and fall off therefore poor by the stability for examining product prepared by this method.
Invention content
Based on this, it is necessary to provide a kind of preferably modified cuorin coating of the stability for detecting anticardiolipin antibodies
Nanometer magnetic bead and preparation method thereof.
A kind of coated nanometer magnetic bead of modified cuorin, including：Modified cuorin, biotin derivative and Streptavidin
Magnetic bead；
The modified cuorin is that the hydrophobic fat acid side chain of cuorin is aoxidized to obtain, and the modified cuorin contains
The structural formula of the biotin derivative is：Wherein ,-the R- is to contain 4
The saturated alkyl chain of a~20 carbon atoms or the polyglycol chain containing 2~10 carbon atoms, in the modified cuorin
'sWith-the NH in the biotin derivative2Generation-CO-NH- structures are to by the modified cuorin and described
Biotin derivative connects together；
The Streptavidin MagneSphere is the nanometer magnetic bead for combining Streptavidin, the biotin derivative and described
Streptavidin connects.
In one embodiment ,-the R- is the saturated alkyl chain containing 4~10 carbon atoms.
In one embodiment, there are one the modified cuorin contains
In one embodiment, the structural formula of the modified cuorin is as follows：
A kind of preparation method of the above-mentioned coated nanometer magnetic bead of modification cuorin, includes the following steps：
It is fully reacted under the conditions of by cuorin and peroxide existing for the first solvent, obtains modified cuorin, wherein
The hydrophobic fat acid side chain of the modified cuorin is aoxidized, and the modified cuorin contains
It is fully reacted under the conditions of by the modified cuorin and biotin derivative existing for the second solvent, obtains biology
The modification cuorin that plain derivative combines, wherein the structural formula of the biotin derivative is：
Wherein ,-the R- is the saturated alkyl chain containing 4~20 carbon atoms or the polyethylene glycol containing 2~10 carbon atoms
Chain, in the modified cuorinWith-the NH in the biotin derivative2Generation-CO-NH- structures will be thus will
The modified cuorin is connected together with the biotin derivative；And
Modification cuorin and Streptavidin MagneSphere that the biotin derivative combines are mixed and fully reacted, is obtained
The coated nanometer magnetic bead of modified cuorin, wherein the Streptavidin MagneSphere is the nanometer magnetic bead for combining Streptavidin,
The biotin derivative is connected with the Streptavidin.
In one embodiment, it is described by cuorin and peroxide existing for the first solvent under the conditions of fully react
In operation, the peroxide is benzoyl hydroperoxide, metachloroperbenzoic acid, Peracetic acid or Perpropionic Acid, the cuorin
Molar ratio with the peroxide is 1：1~8.
In one embodiment, it is described by cuorin and peroxide existing for the first solvent under the conditions of fully react
In operation, first solvent is dichloromethane, chloroform, chloroform, benzene or toluene.
In one embodiment, described by the modified cuorin and the biotin derivative condition existing for the second solvent
In the operation fully reacted down, second solvent is DMSO, DMF, tetrahydrofuran or pH are 6.5~8.5 phosphate-buffered
Liquid.
In one embodiment, described by the modified cuorin and the biotin derivative condition existing for the second solvent
In the operation fully reacted down, the molar ratio of the modified cuorin and the biotin derivative is 1：1.5~20.
In one embodiment, the modification cuorin and Streptavidin MagneSphere that the biotin derivative is combined
In mixing and the operation fully reacted, a concentration of 0.1mg/mL~1mg/ for the modification cuorin that the biotin derivative combines
ML, a concentration of 5mg/mL~15mg/mL of the Streptavidin MagneSphere.
This coated nanometer magnetic bead of modified cuorin is directly securely spread out modified cuorin by chemical bond with biotin
Biological firm connection, biotin derivative are connected with the Streptavidin on Streptavidin MagneSphere surface, are inhaled relative to physics
Attached mode, more easily controls the modification cuorin amount of magnetic bead surfaces, and can be changed by the length of adjusting-R-, realization
The control of the distance between property cuorin amount and nanometer magnetic bead, preferably remains modified cuorin amount and is combined with anti-phospholipid antibody
Space.This coated nanometer magnetic bead of modified cuorin is used directly for the detection of anti-phospholipid antibody, and relative to biography
Detection product prepared by the method for physical absorption cuorin of uniting, has higher stability.
Description of the drawings
Fig. 1 is the flow chart of the preparation method of the coated nanometer magnetic bead of modification cuorin of an embodiment.
Specific implementation mode
In order to make the foregoing objectives, features and advantages of the present invention clearer and more comprehensible, below in conjunction with the accompanying drawings and specific real
Example is applied to be described in detail the specific implementation mode of the present invention.Elaborate in the following description many details in order to
Fully understand the present invention.But the invention can be embodied in many other ways as described herein, art technology
Personnel can do similar improvement without violating the connotation of the present invention, therefore the present invention is not by following public specific implementation
Limitation.
The coated nanometer magnetic bead of modification cuorin of one embodiment, including：Modified cuorin, biotin derivative and chain
Mould avidin magnetic bead.
Cuorin is the Ester being made of 3 glycerine, 2 phosphoric acid and 4 long-chain unsaturated alkyls, which contains
There are 2 hydrophilic centers and 4 hydrophobic side chains.The structural formula of cuorin is as follows：
Modified cuorin is that the hydrophobic fat acid side chain of cuorin is aoxidized to obtain, and modified cuorin contains
Since cuorin has 4 side chains, 4 side chains can be aoxidized simultaneously when cuorin is modified, therefore the modification heart phosphorus
Fat may include multipleSpecifically, which can include 1~8
Preferably, there are one modified cuorin contains
Particularly preferred, the structural formula of modified cuorin is as follows：
The structural formula of biotin derivative is：
Wherein ,-R- is the saturated alkyl chain containing 4~20 carbon atoms or the poly- second containing 2~10 carbon atoms
Glycol chains.
In modified cuorinWith-the NH in biotin derivative2Generation-CO-NH- structures will be to be modified
Cuorin is connected together with biotin derivative.
Preferably ,-R- is the saturated alkyl chain containing 4~10 carbon atoms.
Particularly preferred ,-R- is-(CH2)6-。
Since the size of Cardiolipin molecules is very small, the space wretched insufficiency that can be transformed, to hydrophilic phosphate center
The compatibility of cuorin and anti-phospholipid antibody can be reduced by carrying out modification, or even antigen active is made to disappear.
This coated nanometer magnetic bead of modified cuorin remains heart phosphorus by being transformed to cuorin hydrophobic side chains
The modification cuorin at fat hydrophily phosphate center.
This coated nanometer magnetic bead of modified cuorin is directly securely spread out modified cuorin by chemical bond with biotin
Biological firm connection, biotin derivative are connected with the Streptavidin on Streptavidin MagneSphere surface, are inhaled relative to physics
Attached mode, more easily controls the modification cuorin amount of magnetic bead surfaces, and can be changed by the length of adjusting-R-, realization
The control of the distance between property cuorin amount and nanometer magnetic bead, preferably remains modified cuorin amount and is combined with anti-phospholipid antibody
Space.This coated nanometer magnetic bead of modified cuorin is used directly for the detection of anti-phospholipid antibody, and relative to biography
Detection product prepared by the method for physical absorption cuorin of uniting, has higher stability.
Further, since each Streptavidin can combine closely with 4 biotins, this modified coated nanometer of cuorin
Magnetic bead makes detected signal be exaggerated 4 times by biotin-Streptavidin enlarge-effect, and it is anti-to greatly improve anti-phosphatide
The detection sensitivity of body.
The preparation method of the above-mentioned modified coated nanometer magnetic bead of cuorin as shown in Figure 1, includes the following steps：
S10, by cuorin and peroxide existing for the first solvent under the conditions of fully react, obtain modified cuorin.
By cuorin and peroxide reactions, the fatty acid side chain of cuorin is aoxidized.
Peroxide is benzoyl hydroperoxide, metachloroperbenzoic acid, Peracetic acid or Perpropionic Acid.
The molar ratio of cuorin and peroxide is 1：1~8.
First solvent is dichloromethane, chloroform, chloroform, benzene or toluene.
S10 further includes the operation purified to intermediate product, and purifying can be after ethyl acetate extracts by liquid phase system
Standby chromatogram purification.The modification cuorin of purity about 80% can be obtained after purification.
In S10, reaction temperature is 60 DEG C~100 DEG C.
Since cuorin has 4 side chains, 4 side chains can be aoxidized simultaneously when cuorin is modified, therefore the modification heart phosphorus
Fat may include multipleSpecifically, modified cuorin can include 1~8
Preferably, there are one modified cuorin contains
Particularly preferred, the structural formula of modified cuorin is as follows：
S20, by the obtained modification cuorins of S10 and biotin derivative existing for the second solvent under the conditions of it is fully anti-
It answers, obtains the modification cuorin of biotin derivative combination.
The structural formula of biotin derivative is：
Wherein ,-R- is the saturated alkyl chain containing 4~20 carbon atoms or the poly- second containing 2~10 carbon atoms
Glycol chains.
In modified cuorinWith-the NH in biotin derivative2Generation-CO-NH- structures will be to be modified
Cuorin is connected together with biotin derivative.
Preferably ,-R- is the saturated alkyl chain containing 4~10 carbon atoms.
Particularly preferred ,-R- is-(CH2)6-。
Biotin derivative can be directly commercially available.
In S20, the second solvent is DMSO, DMF, tetrahydrofuran or pH are 6.5~8.5 phosphate buffer.
In S20, the molar ratio of modified cuorin and biotin derivative is 1：1.5~20.
S30, the modification cuorin for the biotin derivative combination that S20 is obtained and Streptavidin MagneSphere mixing are simultaneously abundant
Reaction obtains the coated nanometer magnetic bead of modified cuorin.
Streptavidin MagneSphere is the nanometer magnetic bead for combining Streptavidin, and biotin derivative and Streptavidin connect
It connects.
Streptavidin MagneSphere can be directly to buy, for example, being purchased from MagnaBind companies (article No. 21344).
The modification cuorin and Streptavidin MagneSphere that biotin derivative is combined mix and in the operations fully reacted,
Biotin derivative combine modification cuorin a concentration of 0.1mg/mL~1mg/mL, Streptavidin MagneSphere it is a concentration of
5mg/mL~15mg/mL.
It is secured will to be directly modified cuorin by chemical bond for the preparation method of this coated nanometer magnetic bead of modified cuorin
It is firmly connect with biotin derivative, biotin derivative is connected with the Streptavidin on Streptavidin MagneSphere surface, phase
For the mode of physical absorption, the modification cuorin amount of magnetic bead surfaces is more easily controlled, and the length of adjusting-R- can be passed through
Degree realizes the control of the distance between modified cuorin amount and nanometer magnetic bead, preferably remains modified cuorin amount and anti-phosphorus
The space that fat antibody combines.The coated nanometer magnetic bead of modified cuorin obtained is used directly for the detection of anti-phospholipid antibody,
And the detection product prepared relative to the method that conventional physical adsorbs cuorin, has higher stability.
Further, since each Streptavidin can combine closely with 4 biotins, modified cuorin obtained is coated to be received
Rice magnetic bead makes detected signal be exaggerated 4 times, greatly improves anti-phosphatide by biotin-Streptavidin enlarge-effect
The detection sensitivity of antibody.
It is specific embodiment below.
Embodiment 1：The preparation of modified cuorin
0.5mmol cuorins are dissolved in 1mL dry toluenes, the metachloroperbenzoic acid of 1.5mmol is added, in stirring by
110 DEG C are gradually heated to, and maintains the thermotonus 72 hours.After reaction solution is cooled to room temperature, pour into 25mL ice water,
20mL ethyl acetate extraction solution is used again 3 times, combined ethyl acetate.Combined ethyl acetate is molten molten with 10mL saturated sodium-chlorides
After liquid washs 1 time, anhydrous sodium sulfate is added and dries 12 hours.Ethyl acetate, a small amount of first of remaining solid are evaporated under liquid negative pressure
Alcohol dissolves, and filters out insoluble matter, and solution is purified with preparative liquid chromatography, and it is to be modified to obtain about 210mg colourless oil liquids
Cuorin, MS (ESI+,m/z):1479.83131。
Embodiment 2：The preparation for the modification cuorin that biotin derivative combines
Stirring is lower to sequentially add 200mg ring ethyoxyls modification cuorin, 110mg terminal amino groups into 4mL anhydrous DMF solutions
The biotin derivative of change is stirred to react 10 hours at room temperature.Reaction solution, residual solid 5mL ethyl acetate are evaporated under decompression
Dissolving filters insoluble matter, and the dissolving of the ethyl acetate of gained is evaporated, what obtained white solid i.e. biotin derivative combined
Modified cuorin.
In the present embodiment, the structural formula of biotin derivative is：- R- is-(CH2)6-。
The modification cuorin that biotin derivative manufactured in the present embodiment combines needs not move through especially purifying and can be used to
Subsequent use.
Embodiment 3：It is coated with the preparation of the Streptavidin MagneSphere of modified cuorin
The phosphate buffer for taking the modification cuorin that the biotin derivative that 100 μ L contain 0.2mg/mL combines, is added
100 μ L contain the Streptavidin MagneSphere of 10mg/mL, are incubated 10 minutes for 37 DEG C after mixing.1mL 30mM Tris are used after Magneto separate
Buffer solution redissolves, and redissolves Magneto separate again after the mixed liquor concussion of gained, and the solid of gained 5mL 30mM Tris are buffered
Liquid redissolves, and obtains the Streptavidin MagneSphere solution for being coated with modified cuorin of a concentration of 0.02mg/mL.
Embodiment 4：The avidin magnetic bead for being coated with modified cuorin is reacted with anti-phosphatide sample
The avidin magnetic bead solution for being coated with modified cuorin prepared in 200 μ L Examples 1 to 3 is taken respectively, and 5 μ L are added
Anti-phospholipid antibody serum sample is simultaneously incubated 30 minutes in 37 DEG C.The Streptavidin magnetic of modified cuorin will be coated with after Magneto separate
Pearl is redissolved to 200 μ L, is added the secondary antibody of horseradish peroxidase-labeled, 37 DEG C be incubated 30 minutes after clean successively, the bottoms TMB be added
Thing liquid is simultaneously incubated 10 minutes, is added 100 μ L terminate liquids, is read OD values in 10 minutes in microplate reader, obtain the luminous letter of sample
Number value.
Three cuorin positive serum samples and three cuorin negative serum samples are taken respectively, and with traditional physics
Absorption method is control, compares the OD values measured, obtains the following table 1.
Table 1：Examples 1 to 3 and control group (physisorphtion) measure the OD values of different samples
As can be seen from Table 1, avidin magnetic bead and the physical absorption for being coated with modified cuorin that prepared by Examples 1 to 3
Luminous signal of the magnetic bead (control group) of method absorption when testing anti-phospholipid antibody sample compares, coating prepared by Examples 1 to 3
There is the avidin magnetic bead of the modified cuorin luminous signal value when measuring negative sample is significantly low compared with control group (to reduce 1 times~4
Times), while the avidin magnetic bead for being coated with modified cuorin of Examples 1 to 3 preparation luminous signal when measuring positive sample
Have compared with control group and greatly improves (improving 3~10 times).
Thus illustrate, prepared by Examples 1 to 3 is coated with the avidin magnetic bead of modified cuorin in the anti-phosphatide sample of measurement
When, relative to the magnetic bead of traditional physisorphtion, detection sensitivity, which has, to be obviously improved.
Several embodiments of the invention above described embodiment only expresses, the description thereof is more specific and detailed, but simultaneously
Cannot the limitation to the scope of the claims of the present invention therefore be interpreted as.It should be pointed out that for those of ordinary skill in the art
For, without departing from the inventive concept of the premise, various modifications and improvements can be made, these belong to the guarantor of the present invention
Protect range.Therefore, the protection domain of patent of the present invention should be determined by the appended claims.
Claims (5)
1. a kind of coated nanometer magnetic bead of modified cuorin, which is characterized in that including：Modified cuorin, biotin derivative and
Streptavidin MagneSphere；
The modified cuorin is that the hydrophobic fat acid side chain of cuorin is aoxidized to obtain, there are one the modified cuorin containsThe structural formula of the modified cuorin is as follows：
The structural formula of the biotin derivative is：Wherein ,-the R- is-(CH2)6, institute
It states in modified cuorinWith-the NH in the biotin derivative2Generation-CO-NH- structures by described to change
Property cuorin is connected together with the biotin derivative；
The Streptavidin MagneSphere is the nanometer magnetic bead for combining Streptavidin, the biotin derivative and the strepto-
Avidin connects.
2. a kind of preparation method of the modified coated nanometer magnetic bead of cuorin described in claim 1, which is characterized in that including such as
Lower step：
It is fully reacted under the conditions of by cuorin and peroxide existing for the first solvent, obtains modified cuorin, wherein described
The hydrophobic fat acid side chain of modified cuorin is aoxidized, and the modified cuorin containsThe peroxide is
Benzoyl hydroperoxide, metachloroperbenzoic acid, Peracetic acid or Perpropionic Acid, first solvent be dichloromethane, chloroform,
Chloroform, benzene or toluene；
It is fully reacted under the conditions of by the modified cuorin and biotin derivative existing for the second solvent, obtains biotin and spread out
The modification cuorin of bioconjugation, wherein the structural formula of the biotin derivative is：Its
In ,-the R- is-(CH2)6, in the modified cuorinWith-the NH in the biotin derivative2Generate-
To connect together the modified cuorin with the biotin derivative, second solvent is CO-NH- structures
The phosphate buffer that DMSO, DMF, tetrahydrofuran or pH are 6.5~8.5；And
Modification cuorin and Streptavidin MagneSphere that the biotin derivative combines are mixed and fully reacted, is modified
The coated nanometer magnetic bead of cuorin, wherein the Streptavidin MagneSphere is the nanometer magnetic bead for combining Streptavidin, described
Biotin derivative is connected with the Streptavidin.
3. the preparation method of the coated nanometer magnetic bead of modified cuorin according to claim 2, which is characterized in that described to incite somebody to action
In the operation fully reacted under the conditions of cuorin and peroxide are existing for the first solvent, the cuorin and the peroxidating
The molar ratio of object is 1：1~8.
4. the preparation method of the coated nanometer magnetic bead of modified cuorin according to claim 2, which is characterized in that described to incite somebody to action
In the operation fully reacted under the conditions of the modified cuorin and biotin derivative are existing for the second solvent, the modified heart
The molar ratio of phosphatide and the biotin derivative is 1：1.5~20.
5. the preparation method of the coated nanometer magnetic bead of modified cuorin according to claim 2, which is characterized in that described to incite somebody to action
In the modification cuorin of the biotin derivative combination and Streptavidin MagneSphere mixing and the operation fully reacted, the life
Object element derivative combine modification cuorin a concentration of 0.1mg/mL~1mg/mL, the Streptavidin MagneSphere it is a concentration of
5mg/mL~15mg/mL.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610512711.6A CN106124783B (en) | 2016-06-30 | 2016-06-30 | Coated nanometer magnetic bead of modified cuorin and preparation method thereof |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610512711.6A CN106124783B (en) | 2016-06-30 | 2016-06-30 | Coated nanometer magnetic bead of modified cuorin and preparation method thereof |
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| Publication Number | Publication Date |
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| CN106124783A CN106124783A (en) | 2016-11-16 |
| CN106124783B true CN106124783B (en) | 2018-10-02 |
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| CN1714095A (en) * | 2002-10-16 | 2005-12-28 | 新药物公司 | Cardiolipin molecules and method of synthesis |
| CN101360997A (en) * | 2005-11-18 | 2009-02-04 | 美国政府健康及人类服务部，疾病控制和预防中心 | Modified cardiolipin and its application |
| CN102575244A (en) * | 2009-09-17 | 2012-07-11 | Jsr株式会社 | Dissociation method and dissociation agent of avidin and biotin derivative |
| CN102565405A (en) * | 2011-08-24 | 2012-07-11 | 苏州长光华医生物试剂有限公司 | Method for immunological detection by combining acridinium ester labeling technology with general magnetic particles |
| CN103059288A (en) * | 2012-12-18 | 2013-04-24 | 山东大学 | Biotin derivative of phenylephrine and preparation method and application thereof |
| CN103193976A (en) * | 2013-04-12 | 2013-07-10 | 上海艾韦特医药科技有限公司 | Synthesis and application of cardiolipin |
| CN103298949A (en) * | 2010-10-29 | 2013-09-11 | 生命科技公司 | Biotin derivatives |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN1714095A (en) * | 2002-10-16 | 2005-12-28 | 新药物公司 | Cardiolipin molecules and method of synthesis |
| CN101360997A (en) * | 2005-11-18 | 2009-02-04 | 美国政府健康及人类服务部，疾病控制和预防中心 | Modified cardiolipin and its application |
| CN102575244A (en) * | 2009-09-17 | 2012-07-11 | Jsr株式会社 | Dissociation method and dissociation agent of avidin and biotin derivative |
| CN103298949A (en) * | 2010-10-29 | 2013-09-11 | 生命科技公司 | Biotin derivatives |
| CN102565405A (en) * | 2011-08-24 | 2012-07-11 | 苏州长光华医生物试剂有限公司 | Method for immunological detection by combining acridinium ester labeling technology with general magnetic particles |
| CN103059288A (en) * | 2012-12-18 | 2013-04-24 | 山东大学 | Biotin derivative of phenylephrine and preparation method and application thereof |
| CN103193976A (en) * | 2013-04-12 | 2013-07-10 | 上海艾韦特医药科技有限公司 | Synthesis and application of cardiolipin |
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