CN106290920A - Exenatide detection kit - Google Patents
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- CN106290920A CN106290920A CN201610781418.XA CN201610781418A CN106290920A CN 106290920 A CN106290920 A CN 106290920A CN 201610781418 A CN201610781418 A CN 201610781418A CN 106290920 A CN106290920 A CN 106290920A
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- G—PHYSICS
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- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
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- G—PHYSICS
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Abstract
The invention discloses a kind of Exenatide detection kit, it includes that Exenatide monoclonal antibody, ELISA Plate, positive reference substance, negative controls, cleaning mixture, stop buffer, nitrite ion, described Exenatide monoclonal antibody include the Exenatide monoclonal antibody of Exenatide monoclonal antibody and the horseradish peroxidase-labeled being coated in ELISA Plate.Detection kit disclosed by the invention is when the concentration of detection Exenatide in test sample, and its sensitivity can reach 1ng/mL, highly sensitive, and testing result is accurate, and detection object can be people, animal serum, and Exenatide formulation, applied range.
Description
Technical field
The present invention relates to biology and field of medical examination, be specifically related to a kind of Exenatide detection kit.
Background technology
Type 2 diabetes mellitus original name is Adult Onset's patients with type Ⅰ DM, have another name called non-insulin-dependent diabetes mellitus, this patient from
Body produces insulin ability and does not completely lose, but is in internal insulin and relatively lacks, can be by medicine irritation body
The state of insulin secretion.The cause of disease and the pathogenesis of type 2 diabetes mellitus are the most indefinite, and its significant pathophysiological features is
The islets of langerhans that insulin resistant (i.e. the decline of insulin regulation and control glucose metabolism ability) is caused with islet beta cell function defect
Element secretion reduces or relatively reduces.
Secretin is a kind of hormone after ingesting by the secretion of little enteroendocrine cell reactivity, and it includes gastrointestinal tract L
The glucagon-like peptide 1 (Glucagon-likepeptide1, GLP-1) of Hemapoiesis, the dependence on the glucose of K Hemapoiesis
Property pancreotropic hormone polypeptide (Glucose-dependentinsulinotropicpolypeptide, GIP), vasoactive peptide
(Vasoactive intestinal peptide, VIP), cholecystokinin (Cholecystokinin, CCK) etc., wherein
GLP-1 has protective effect to beta Cell of islet, and ADA/EASD common recognition (2008) is listed in treating diabetes route map,
GLP-1 is used for treating the successive treatment of patient the most up to standard as metformin, especially for hypoglycemia must be avoided to occur as far as possible
Diabetics, this type of patient has cardiovascular diseases's history more or has high blood pressure disease risk.But, the GLP-that human body self produces
1 easily by internal DPP-IV (dipeptidyl peptidase IV) degraded, and its plasma half-life was less than 2 minutes, i.e. GLP-1 can be fast by human body
Speed inactivation, it has to last for intravenous drip or continuous subcutaneous infusion could produce curative effect, and this is to suffering from this chronic disease of diabetes
More inconvenience for sick patient, thus greatly limit the clinical practice of GLP-1.
Exenatide (Exenatide) be from uncommon draw isolated one huge lizard sialoprotein contain 39 amino acid whose
Polypeptide, its GLP-1 similarity with human body reaches more than 50%, in vitro in experiment can and known human body β cell on GLP-1
Receptor combines, and can resist the deactivation of DPP-IV.Exenatide be 2005 by U.S. food pharmaceuticals administration
1st GLP-1 analog of office (FDA) approval listing, 2009 by state food and drug administration (CFDA) batch
Mutatis mutandis in improving the glycemic control of type 2 diabetes mellitus patient, there is Exenatide injection (trade name: hundred secrete what China had listed
Reach), the most domestic have many pharmaceutical producing enterprises and research unit studying and declaring Exenatide injection, inject micro-
The relevant dosage form such as ball, enteric coatel tablets.Along with Exenatide research being goed deep into and the expansion of related drugs formulation development, Exenatide
Pharmacokinetic in blood is particularly important, pharmacokinetic to pass judgment on the reasonability of pharmaceutical dosage form, science,
And clinical administration dosage is all played vital directive function.
For Exenatide medicine generation research, Chinese patent disclose entitled " a kind of Exenatide pharmacokinetics
Method of testing (Application No. CN 201010237160.X) " patent of invention, scheme disclosed in this patent is by human plasma sample
Redissolve to suitable concn after Solid-Phase Extraction column extracting is concentrated freeze-dried, then through HPLC-UV detection device to Ai Saina
Peptide detects, and in the method, Exenatide concentration is linear in 5～160 μ g/mL trial stretches, and detection limit is
1.5ng.It addition, LINCO Reagent Company of the existing document report U.S. produces for detecting Exenatide concentration in human plasma
ELISA kit；Wang Lin carries in " restructuring Exenatide-human serum albumin fusion proteins Preclinical metabolism and pharmacokinetics study "
And a kind of commercial goods Exenatide detection kit, it is as coated antibody by mouse-anti Exenatide monoclonal antibody,
Chicken AHS's albumin polyclonal antibody is detection antibody (two resist), the detectable concentration scope of this ELISA method be 2～
200ng·mL-1, minimum quantitatively it is limited to 2ng mL-1。
Enzyme-linked immunosorbent assay (Enzyme linked immunosorbent assay, ELISA) has quick, simple
Just, highly sensitive, the feature of high specificity, be one of widely used important method in current drug development process.Therefore base
Develop the Exenatide detection kit of high sensitivity, high specific and stability in enzyme-linked immunosorbent assay for measuring, use
The detection of Exenatide in the serum such as people, rabbit, rat, mice, pig, cattle, this is to Exenatide pharmacy and Pharmaceutical study meeting
Play an important role.
Summary of the invention
It is an object of the invention to provide and a kind of detect that elaboration is high, the Exenatide detection kit of good stability.
For achieving the above object, the technical solution used in the present invention is: a kind of Exenatide detection kit, its feature exists
In: include Exenatide monoclonal antibody, ELISA Plate, positive reference substance, negative controls, cleaning mixture, stop buffer, nitrite ion,
Described Exenatide monoclonal antibody includes Exenatide monoclonal antibody and the horseradish peroxidase being coated in ELISA Plate
The Exenatide monoclonal antibody of enzyme labelling.
Concrete scheme is, the described Exenatide monoclonal antibody being coated in ELISA Plate is Exenatide monoclonal
Antibody 3D7, the Exenatide monoclonal antibody of described horseradish peroxidase-labeled is Exenatide monoclonal antibody 5F3, Chinese mugwort
Fill in that peptide monoclonal antibody 3D7、5F3Preparation process as follows:
B) with the Balb/C in Exenatide immune a collection of 6-8 week, take immune mouse spleen cell and make cell suspension；
B) splenocyte suspension step a obtained mixes with myeloma cell SP2/0, and under the effect of fusion agent PEG
Carry out cell fusion；
C) with ELISA method filter out from the cell of above-mentioned fusion secretion Exenatide monoclonal antibody hybridoma thin
Born of the same parents, use limiting dilution assay that hybridoma carries out cloning, thus obtain five strains and have stably excreting Exenatide Dan Ke
The hybridoma cell strain of grand antibody ability；
D) by hybridoma cell strain 5F3、1C9、5C3、4H10And 3D7It is injected separately in different female Mus bodies, collects ascites, will
Antibody purification in ascites, obtains Exenatide monoclonal antibody 5F3、1C9、5C3、4H10And 3D7, then judge Ai Sai through screening
That peptide monoclonal antibody 3D7、5F3Respectively as be coated in ELISA Plate and the Exenatide list of horseradish peroxidase-labeled
Clonal antibody；
Exenatide monoclonal antibody 3D7、5F3The step of screening is: take five groups of Exenatide monoclonals after partial purification
Antibody 5F3、1C9、5C3、4H10And 3D7, use horseradish peroxidase to be marked respectively, add equal-volume glycerol, frozen, so
By chessboard method, five groups of Exenatide monoclonal antibodies of labelling horseradish peroxidase are screened afterwards, obtain two group echos peppery
The Exenatide monoclonal antibody of root peroxidase, the Exenatide monoclonal antibody of this two group echos horseradish peroxidase
Source be 3D7、5F3, the present invention is at screening 3D7、5F3During, it is five groups of Exenatide monoclonals by obtaining
Antibody takes part and screens, and it specifically carries out horseradish peroxidase-labeled respectively, then uses chessboard method screening, its
Purpose is two anti-(also the getting final product the Exenatide monoclonal antibody of labelling horseradish peroxidase) that screening obtains that stability is good.
What limiting dilution assay, antibody purification and the chessboard method etc. related in said method did not elaborated is conventional method, this
Skilled person is appreciated that and is capable of.It should be noted that present invention employs said method finally screen judgement
Exenatide monoclonal antibody 3D obtained7、5F3Being coated and horseradish peroxidase of ELISA Plate can be efficiently used for respectively
Labelling, so make the detection kit that obtains when the concentration of detection Exenatide in test sample, its sensitivity up to
To 1ng/mL, highly sensitive, testing result is accurate.
As further preferred version: the method for the female Mus of Balb/C in Exenatide immunity a collection of 6-8 week in step a)
For: the 1st week, take the Exenatide that 0.5ml concentration is 1mg/ml and fully mix emulsifying with isopyknic Freund's complete adjuvant, respectively
The each female Mus of immunity；2nd, 3,4,5 weeks all with the Exenatide that 0.5ml concentration is 1mg/ml and isopyknic incomplete Freund's adjuvant
Fully mix emulsifying, respectively each female Mus of immunity；Within 6th week, take blood from the afterbody of female Mus, survey serum titer by ELISA method, by titre
The highest female Mus 1ml concentration is the immunity of 1mg/ml Exenatide, and the splenocyte taking this female Mus after three days makes cell suspension.
Described cleaning mixture is to be the PBS solution of 7.4 containing 0.05% tween 20, pH；Described stop buffer is that concentration is
The sulfuric acid solution of 2mol/L；Described positive reference substance, negative controls are respectively the Exenatide positive serum of human body, Ai Sai
That peptide negative serum；In step d), the purification process of antibody is octanoic acid-ammonium sulfate precipitation method or polyethylene glycol precipitation；Described
Nitrite ion be TMB nitrite ion, use TMB nitrite ion time, be marked at Exenatide monoclonal antibody 5F3On Radix Cochleariae officinalis peroxidating
Thing enzyme can react with TMB nitrite ion, and after adding stop buffer, solution is reflected ELISA system by colourless yellowing, the yellow depth
Middle antigen-antibody combine number, thus judge antibody number, so can by microplate reader survey read light absorption value, and then judge
The content of Exenatide in testing sample.Concrete, described test kit also includes the confining liquid for coated elisa plate, specifically seals
Close liquid and can use conventional commercial.
Accompanying drawing explanation
Fig. 1 is the canonical plotting of the Exenatide made；
Fig. 2 is that subcutaneous injection hundred secretes the curve chart of Exenatide concentration change in the blood sample of the SD rat reached；
Fig. 3 is the Exenatide detection of different preparation.
Detailed description of the invention
Hereinafter preparation, use step and the using effect of Exenatide detection kit is described further:
Embodiment 1: the preparation of Exenatide detection kit
1) conventional equipment and the preparation of reagent:
96 hole ELISA Plate, Exenatide positive serum, Exenatide negative serum, cleaning mixture, stop buffer, Radix Cochleariae officinalis peroxidating
Thing enzyme and TMB nitrite ion (it is made up of nitrite ion A, nitrite ion B), wherein Exenatide positive serum and Exenatide are cloudy
The coincidence rate of property serum reaches 100%.
2) preparation of Exenatide monoclonal antibody:
A) the 1st week, take the Exenatide that 0.5ml concentration is 1mg/ml and fully mix breast with isopyknic Freund's complete adjuvant
Change, respectively each female Mus of immunity；2nd, 3,4,5 weeks are not all with the Exenatide that 0.5ml concentration is 1mg/ml with isopyknic Freund
Freund's complete adjuvant fully mixes emulsifying, respectively each female Mus of immunity；Within 6th week, take blood from the afterbody of female Mus, survey serum by ELISA method and drip
Degree, is the immunity of 1mg/ml Exenatide by female Mus 1ml concentration the highest for titre, and the splenocyte taking this female Mus after three days is made carefully
Born of the same parents' suspension；
B) cell suspension obtained by step a) mixes with myeloma cell SP 2/0, and under the effect of fusion agent PEG
Carrying out cell fusion, described myeloma cell SP 2/0 is provided by Chinese Academy of Sciences's Shanghai cell bank；
C) with ELISA method filter out from the cell of above-mentioned fusion secretion Exenatide monoclonal antibody hybridoma thin
Born of the same parents, use limiting dilution assay that hybridoma carries out cloning, thus obtain five strains and have stably excreting Exenatide Dan Ke
The hybridoma cell strain of grand antibody ability, is respectively designated as hybridoma cell strain 5F3, hybridoma cell strain 1C9, hybridoma
Strain 5C3, hybridoma cell strain 4H10, hybridoma cell strain 3D7；
D) by hybridoma cell strain 5F3、1C9、5C3、4H10And 3D7It is injected separately in different female Mus bodies, collects ascites, adopt
With octanoic acid-ammonium sulfate precipitation method by the antibody purification in ascites, obtain Exenatide monoclonal antibody 5F3、1C9、5C3、4H10With
3D7, the purity of these Exenatide monoclonal antibodies is all up to more than 95%, and screening afterwards judges Exenatide monoclonal antibody
3D7、5F3Respectively as be coated in ELISA Plate and the Exenatide monoclonal antibody of horseradish peroxidase-labeled；
Exenatide monoclonal antibody 3D7、5F3The step of screening is: take five groups of Exenatide monoclonals after partial purification
Antibody 5F3、1C9、5C3、4H10And 3D7, use horseradish peroxidase to be marked respectively, add equal-volume glycerol, frozen, so
By chessboard method, five groups of Exenatide monoclonal antibodies of labelling horseradish peroxidase are screened afterwards, obtain two group echos peppery
The Exenatide monoclonal antibody of root peroxidase, the Exenatide monoclonal antibody of this two group echos horseradish peroxidase
Source be 3D7、5F3, experiment proves that, Exenatide monoclonal antibody 3D7、5F3Cell strain cultivate continuously 6 months with
On, mice interior generation more than 4 times, liquid nitrogen preserves after half a year and recovers, secretory antibody ability is constant, it can be said that bright both
The stability of secretory antibody is the best.
The step of horseradish peroxidase-labeled each Exenatide monoclonal antibody is: horseradish peroxidase is dissolved in steaming
Distilled water, adds the sodium periodate solution newly joined, is placed in refrigerator 30 minutes, takes out and add ethylene glycol, and room temperature is placed 30 minutes, adds
Entering in Exenatide monoclonal antibody 5F3 after purification, 1C9,5C3,4H10 and 3D7, mixing, then at the carbonic acid of PH 9.5
Dialysed overnight in saline solution, adds sodium borohydride afterwards and is placed in refrigerator 2 hours, purify with ammonium sulfate precipitation, centrifugal, precipitate
Molten for PH 7.4PBS dialyse, be eventually adding equal-volume glycerol, frozen.
Exenatide monoclonal antibody 3D7、5F3The selection of antibody working concentration: with the 3D of 10 μ g/ml7For coated antibody,
With HRP-5F3 (i.e. Exenatide monoclonal antibody 5F of horseradish peroxidase-labeled3) for detecting antibody, detectable concentration is respectively
10 times, 20 times, 50 times, 100 times, 200 times, 500 times of dilutions, add known variable concentrations Exenatide protein standard substance, choosing
Select blank group OD value less, and the closest the working as optimum detection of linear relationship between OD value and standard protein concentration
Concentration, the working concentration 100 of HRP-5F3 is diluted to optimal compatibility again.
3) 96 hole ELISA Plate it are coated:
(purchased from Thermo company, model is SuperBlocking Buffer to use SuperBlock confining liquid
InPBS, #37515) by Exenatide monoclonal antibody 3D7It is diluted to 10～100 μ g/ml, adds 96 according to the amount in 100 μ l/ holes
In the ELISA Plate of hole, stand overnight under conditions of 2～8 DEG C, wash with cleaning mixture, dry, Exenatide Dan Ke must be coated with
The 96 hole ELISA Plate of grand antibody 3D7.
4) preparation of the Exenatide monoclonal antibody of horseradish peroxidase-labeled: horseradish peroxidase is dissolved in steaming
Distilled water, adds the sodium periodate newly joined, is placed in the refrigerator that temperature is DEG C 30 minutes, takes out, add ethylene glycol, and ambient temperatare is put
30 minutes, add 5F after purification3, mixing, then dialysed overnight in the carbonate solution of PH9.5, add boron hydrogen afterwards
Changing sodium, in the refrigerator being placed in 2 hours, take out, purify with ammonium sulfate precipitation, centrifugal, precipitate is dissolved in PH 7.4PBS dialysis, adds
Enter equal-volume glycerol, frozen, obtain Exenatide monoclonal antibody 5F of horseradish peroxidase-labeled3。
Embodiment 2: the use of Exenatide detection kit
1) sample-adding:
It is being coated with Exenatide monoclonal antibody 3D796 hole ELISA Plate on divide Positive control wells, negative control hole,
Testing sample hole and blank well totally four groups of detection holes, add Exenatide positive serum, in negative control hole in Positive control wells
Adding Exenatide negative serum, add test serum in testing sample hole, the addition of three kinds of samples is 100 μ l/ holes, so
After add a cover in ELISA Plate or overlay film, 2h under the conditions of ELISA Plate is placed in 37 DEG C, discard liquid, with cleaning mixture wash, dry,
Washing, the step dried at least circulate and are repeated 3 times；
2) Exenatide monoclonal antibody 5F of horseradish peroxidase-labeled3:
Each detection hole adds Exenatide monoclonal antibody 5F of 100 μ l horseradish peroxidase-labeled3, at 37 DEG C
Under the conditions of place after 2h and discard liquid, each detection hole adds 350 μ l cleaning mixture and soaks 3～5min, dry or clap lightly
Dry, the action wash, dried at least circulates and is repeated 3 times；
3) nitrite ion is added:
By the nitrite ion A of TMB nitrite ion, the mixing of nitrite ion B equal-volume, in each detection hole, then add 50 μ l mixing
Liquid, lucifuge under the conditions of ELISA Plate is placed in 37 DEG C, colour developing；
4) add stop buffer: according to step 3) in nitrite ion addition sequence successively in each detection hole add stop buffer,
Terminate reaction, terminate the liquid showing as detecting in hole of reaction by blue fast transition yellowly；
5) under the wavelength condition of 490nm, detect the optical density OD value in each detection hole by microplate reader, then change through program
Calculate and i.e. obtain the concentration of Exenatide in testing sample.Use canonical plotting such as Fig. 1 that the OD value of Positive control wells is made at any time,
The Exenatide concentration of the Positive control wells in test kit is respectively 0.25,0.5,1,2,4,8,16,32ng/ml, can from Fig. 1
To find out in the range of this, standard curve linear good, lowest detectable limit can reach 1ng/ml.
Embodiment 3: the detection of Exenatide in animal serum:
Select test grade SD rat (about 6 week old), subcutaneous injection Exenatide injection (trade name: hundred secrete reach), adopt
Collect the 0th, 5,15,30,60,120,180, the blood sample of 240min, then use the examination that method disclosed in the embodiment of the present invention 2 makes
Agent box and ELISA method analyze the concentration of Exenatide in blood sample, and use microplate reader to be measured, and set up metabolic chart, knot
The most as shown in Figure 2: it can be seen that SD rat skin lower injection hundred is secreted after reaching, the Exenatide in its blood is left at 15min
The right side reaches peak value, becomes for Monitoring lower-cut after 3h.Use the test kit that the present invention relates to, can effectively detect in rat blood and end
Fill in the concentration of that peptide, set up complete medicine for concentration curve.
It should be noted that during actual application, detection kit disclosed by the invention can be used for people, rabbit, rat, mice,
The detection of Exenatide, applied range in the multiple serum such as pig, cattle.
Embodiment 4: the detection of Exenatide in different preparations:
By Exenatide injection (trade name: hundred secrete and reach；Specification: 0.25mg/ml；Manufacturer: U.S. Baxter
Pharmaceutical Solutions LLC) be diluted to 0.25 respectively with sample diluting liquid, 0.5,1,2,4,8,16,32ng/
ml；Exenatide lipidosome gel (An Ke biotech firm is from triturate, specification: 100 μ g/ml) is taken 1ml and adds 1ml injection
Water, room temperature stirring at low speed 4h, take middle fining end demultiplexing sample diluting liquid be diluted to 0.25 respectively, 0.5,1,2,4,8,16,
32ng/ml；With the present invention make detection kit both preparations are detected, result as shown in Figure 3: in Fig. 3
Standards refers to the positive controls of test kit, and Sample 1 refers to Exenatide injection group, and Sample 2 refers to
Being Exenatide lipidosome gel group, Sample 3 refers to negative control group, it can be seen that with positive controls phase
Ratio, preparation group is linearly good, and detection limit, up to 1ng/ml, illustrates that the Exenatide concentration in preparation is directly adopted after dilution processes
Detect with test kit of the present invention, do not limited by serum, it can be seen that the detection kit range of application of the present invention is not only limited to
Serum.Wherein, Fig. 1,3 all employing software softmax pro directly derive after carrying out computational analysis.
Claims (9)
1. an Exenatide detection kit, it is characterised in that: include that Exenatide monoclonal antibody, ELISA Plate, the positive are right
According to product, negative controls, cleaning mixture, stop buffer, nitrite ion, described Exenatide monoclonal antibody includes being coated on ELISA Plate
On Exenatide monoclonal antibody and the Exenatide monoclonal antibody of horseradish peroxidase-labeled.
Exenatide detection kit the most according to claim 1, it is characterised in that: described is coated in ELISA Plate
Exenatide monoclonal antibody is Exenatide monoclonal antibody 3D7, the Exenatide of described horseradish peroxidase-labeled
Monoclonal antibody is Exenatide monoclonal antibody 5F3, Exenatide monoclonal antibody 3D7、5F3Preparation process as follows:
A) with the Balb/C in Exenatide immune a collection of 6-8 week, take immune mouse spleen cell and make cell suspension；
B) splenocyte suspension step a obtained mixes with myeloma cell SP2/0, and carries out under the effect of fusion agent PEG
Cell merges；
C) from the cell of above-mentioned fusion, filter out the hybridoma of secretion Exenatide monoclonal antibody by ELISA method, adopt
With limiting dilution assay, hybridoma carried out cloning, thus obtain five strains and there is stably excreting Exenatide monoclonal antibody
The hybridoma cell strain of ability；
D) by hybridoma cell strain 5F3、1C9、5C3、4H10And 3D7It is injected separately in different female Mus bodies, collects ascites, by ascites
In antibody purification, obtain Exenatide monoclonal antibody 5F3、1C9、5C3、4H10And 3D7, then judge Exenatide through screening
Monoclonal antibody 3D7、5F3Respectively as be coated in ELISA Plate and the Exenatide monoclonal of horseradish peroxidase-labeled
Antibody；
Exenatide monoclonal antibody 3D7、5F3The step of screening is: take five groups of Exenatide monoclonal antibodies after partial purification
5F3、1C9、5C3、4H10And 3D7, use horseradish peroxidase to be marked respectively, add equal-volume glycerol, frozen, then use
Five groups of Exenatide monoclonal antibodies of labelling horseradish peroxidase are screened by chessboard method, obtain two group echo Radix Cochleariae officinalis mistakes
The Exenatide monoclonal antibody of oxide enzyme, coming of the Exenatide monoclonal antibody of this two group echos horseradish peroxidase
Source is 3D7、5F3。
Exenatide detection kit the most according to claim 2, it is characterised in that: Exenatide immunity one in step a)
The method of the female Mus of Balb/C criticizing 6-8 week is: the 1st week, takes Exenatide and isopyknic Freund that 0.5ml concentration is 1mg/ml
Freund's complete adjuvant fully mixes emulsifying, respectively each female Mus of immunity；2nd, 3,4,5 weeks is all the Ai Saina of 1mg/ml by 0.5ml concentration
Peptide and isopyknic incomplete Freund's adjuvant fully mix emulsifying, respectively each female Mus of immunity；Within 6th week, take blood from the afterbody of female Mus,
Survey serum titer by ELISA method, be the immunity of 1mg/ml Exenatide by female Mus 1ml concentration the highest for titre, after three days, take this
The splenocyte of female Mus makes cell suspension.
4. according to the Exenatide detection kit described in Claims 2 or 3, it is characterised in that: described cleaning mixture is to contain
0.05% tween 20, pH are the PBS solution of 7.4.
Exenatide detection kit the most according to claim 4, it is characterised in that: described stop buffer is that concentration is
The sulfuric acid solution of 2mol/L.
Exenatide detection kit the most according to claim 5, it is characterised in that: described positive reference substance, feminine gender
Reference substance is respectively the Exenatide positive serum of human body, Exenatide negative serum.
Exenatide detection kit the most according to claim 6, it is characterised in that: described nitrite ion is TMB colour developing
Liquid.
Exenatide detection kit the most according to claim 7, it is characterised in that: described test kit also includes for wrapping
By the confining liquid of ELISA Plate.
Exenatide detection kit the most according to claim 8, it is characterised in that: the purification process of antibody in step d)
For octanoic acid-ammonium sulfate precipitation method or polyethylene glycol precipitation.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108196062A (en) * | 2017-12-20 | 2018-06-22 | 上海美迪西生物医药股份有限公司 | A kind of emulative chemiluminescence determination dissociates the method for Exenatide |
| CN112684177A (en) * | 2020-12-17 | 2021-04-20 | 北京维德维康生物技术有限公司 | Dairy multi-factor rapid detection kit and detection method thereof |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN108196062A (en) * | 2017-12-20 | 2018-06-22 | 上海美迪西生物医药股份有限公司 | A kind of emulative chemiluminescence determination dissociates the method for Exenatide |
| CN112684177A (en) * | 2020-12-17 | 2021-04-20 | 北京维德维康生物技术有限公司 | Dairy multi-factor rapid detection kit and detection method thereof |
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