CN108107222A - A kind of Soluble growth stimulates 2 protein immunization detection reagent of expressing gene and its preparation and detection method - Google Patents

A kind of Soluble growth stimulates 2 protein immunization detection reagent of expressing gene and its preparation and detection method Download PDF

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CN108107222A
CN108107222A CN201711403036.4A CN201711403036A CN108107222A CN 108107222 A CN108107222 A CN 108107222A CN 201711403036 A CN201711403036 A CN 201711403036A CN 108107222 A CN108107222 A CN 108107222A
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expressing gene
soluble growth
stimulates
albumen
preparation
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严芳芳
王亚盟
程月萍
李瑶
杜爱铭
徐兵
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Taiyuan Rui Sheng Biotechnology Co Ltd
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Taiyuan Rui Sheng Biotechnology Co Ltd
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    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6863Cytokines, i.e. immune system proteins modifying a biological response such as cell growth proliferation or differentiation, e.g. TNF, CNF, GM-CSF, lymphotoxin, MIF or their receptors
    • G01N33/6869Interleukin
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/5306Improving reaction conditions, e.g. reduction of non-specific binding, promotion of specific binding
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/53Immunoassay; Biospecific binding assay; Materials therefor
    • G01N33/531Production of immunochemical test materials
    • G01N33/532Production of labelled immunochemicals
    • G01N33/535Production of labelled immunochemicals with enzyme label or co-enzymes, co-factors, enzyme inhibitors or enzyme substrates

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  • Life Sciences & Earth Sciences (AREA)
  • Immunology (AREA)
  • Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Chemical & Material Sciences (AREA)
  • Urology & Nephrology (AREA)
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  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

The invention discloses a kind of Soluble growths to stimulate 2 protein immunization detection reagent of expressing gene and its preparation and detection method.Including:Enzyme mark Soluble growth stimulates 2 albumen of expressing gene, stimulates the indicator of 2 protein antibodies enzyme mark Soluble growth of expressing gene stimulation 2 albumen composition of expressing gene for detecting Soluble growth;Above-mentioned enzyme mark Soluble growth stimulates 2 albumen of expressing gene by Soluble growth 2 albumen of expressing gene and glucose dehydrogenase to be stimulated to be coupled.The Soluble growth of the present invention stimulates 2 protein immunization detection reagent of expressing gene that can accurately and quickly determine that Soluble growth stimulates 2 protein content of expressing gene in the samples such as blood of human body.Compared with existing detection reagent in the market, detection reagent of the present invention has many advantages, such as fast and easy, high sensitivity, high specificity, quantitative accurate, is conducive to clinical promote the use of.

Description

A kind of Soluble growth stimulates 2 protein immunization detection reagent of expressing gene and its preparation And detection method
Technical field
The present invention relates to field of biological detection, are specifically that a kind of Soluble growth stimulates the detection of 2 protein immunization of expressing gene Reagent and its preparation and detection method.
Background technology
Soluble growth stimulates 2 albumen of expressing gene (ST2), is one of IL-1 (interleukin 1) receptor family member, Two kinds of protein products of the gene expression:One kind is known as cross-module type ST2 (ST2L), one kind can be secreted into carefully with transmembrane structure It is extracellular, it is known as secreting type ST2 (sST2).Research shows that sST2 is the Decoy receptor of IL-33, it can be combined with IL-33, so as to IL-33 is blocked to be combined with ST2L, the cardiovascular protective effect for then weakening IL-33/ST2L signal paths is subject to excessively in cardiac muscle During drawing causes damage, a large amount of sST2 generations make the protection that cardiac muscle lacks enough IL-33, so as to accelerate myocardial remodelling And ventricular dysfunction, it ultimately results in mortality risk and increases.Research finds that ST2 can be by cardiac fibroblast and cardiac muscle cell's table It reaches, is a kind of myocardium protein that Biomechanical force induction generates.A variety of pathophysiological processes are participated in, there is immunological regulation work( Can, especially play a significant role in various inflammatory and allergic disease.In addition, the quantitative determination of ST2 contents contributes to Clinician does prognosis risk assessment for patients with heart failure well.
The common method of detection ST2 is mainly enzyme-linked immunosorbent assay at present(ELISA).But ELISA method is quantitative Accuracy is poor, detection device requirement is high, of high cost, the operating time is long, the degree of automation is low, and disturbing factor is more, and repeatability is not It is good.Therefore Enzyme-multiplied immune technique detection ST2 is not suitable for clinical quick diagnosis.
The method that the present invention uses is for homogeneous enzyme immunoassay detection method, advantage:Easy to operate, quick, high sensitivity, standard Really property is good, is suitable for automating, and is widely used, and with automatic clinical chemistry analyzer to small-molecule substance and macromolecular substances all It can high-throughput quick measure.
The content of the invention
It is an object of the invention to solve, ST2 detection process is complicated for operation in the prior art and accuracy of measurement is low Problem, the present invention provides a kind of quick, high sensitivity, the accurate ST2 homogeneous enzyme immunoassay for detecting ST2 contents in sample to be tested Detection reagent and preparation method thereof.
To achieve the above object, the present invention provides following technical solution:
A kind of ST2 immunologic function test reagents and its preparation and detection method, it is characterised in that:Enzyme mark ST2, for detect ST2 antibody- The indicator of enzyme mark ST2 compounds;Above-mentioned enzyme mark ST2 is coupled by ST2 and glucose dehydrogenase.
As further embodiment of the present invention, the indicator is selected from enzymatic reagent, including:The bottom of enzyme mark conjugate and enzyme Object;Above-mentioned enzyme mark conjugate includes glucose dehydrogenase-ST2 conjugates;The substrate of above-mentioned enzyme is glucose.
As further embodiment of the present invention, described a kind of ST2 immunologic function test reagents and preparation method thereof, feature exists In including the following steps:
(1)The preparation of glucose dehydrogenase-ST2 conjugates:Prepare glucose dehydrogenase(GDH)Solution, the activation of ST2, by GDH It is coupled with ST2, purifies the enzyme-labelled antigen of coupling;
(2)The preparation of ST2 homogeneous enzyme immunoassay detection reagents:
The preparation of reagent 1:It is mixed by ST2 antibody and homogeneous zymolyte;
The preparation of reagent 2:It is mixed by glucose dehydrogenase-antigen conjugates with phosphate buffer.
As further embodiment of the present invention, a kind of preparation method of ST2 immunologic function test reagents, feature exists In the step(1)Detailed process is:
1)Glucose dehydrogenase(GDH)The preparation of solution:
A. the GDH that 5-20 mg specifications are 100-300 KU is weighed, room-temperature dissolution is in 6-10 mL PBS solutions, pH 7.0- 10.0;
2)The activation of ST2
Following chemicals is added to stirring and dissolving in beaker:10-30 mg Soluble growths stimulate 2 albumen of expressing gene, 10- 40 mg 1- ethyls -3- (- 3- dimethylaminopropyls) carbodiimides(EDC), 2-10 mg N- hydroxy thiosuccinimides (NHS), it is dissolved in 2- (N- morpholines) ethanesulfonic acid(MES)In solution, stirring and dissolving, reacts 15-60 minutes at room temperature;
3)The coupling of GDH and ST2
A. the ST2 solution of above-mentioned activation is added dropwise in the GDH solution in above-mentioned stirring;
B 2-8 DEG C are stirred overnight;
4)Purify the enzyme-labelled antigen of coupling
The enzyme-labelled antigen being coupled by G-25 gel chromatographies column purification, obtains glucose dehydrogenase-ST2 conjugates, at 2-8 DEG C Storage.
As further embodiment of the present invention, a kind of preparation method of ST2 immunologic function test reagents, feature exists In step(2)Detailed process it is as follows:
The preparation of reagent 1:By nicotinamide adenine dinucleotide NAD, the 0.5-3 g glucose 0.5-2L of 2-5 g oxidation state Homogeneous zymolyte is made in phosphate buffer dissolving;ST2 antibody is added in above-mentioned homogeneous zymolyte, antibody and homogeneous zymolyte Volume ratio be 1:100~1:10000;
The preparation of reagent 2:Glucose dehydrogenase-ST2 the conjugates of preparation are added in phosphate buffer, above-mentioned conjugate with The volume ratio of phosphate buffer is 1:100~1:10000.
As further embodiment of the present invention, the detection method of the ST2 immunologic function test reagents, which is characterized in that bag Include following steps:
1)Sample to be tested is contacted with ST2 antibody;
2)According to the combination situation of enzyme mark ST2 in sample to be tested and ST2 antibody, contained using ST2 in indicator judgement sample Amount;The sample to be tested is serum, blood plasma, saliva or urine.
The principle of the present invention is that antigen is combined into enzyme-labelled antigen with enzyme, retains the bioactivity of antigen and enzyme, when enzyme mark resists After original is combined with antibody, zymoprotein and antibody close contact on antigen molecule make the activated centre of enzyme be affected, the work of enzyme Property is suppressed.Antigen, enzyme-labelled antigen during measure in sample are combined with antibody competition, and the antigenic content in sample is higher, Its OD value is higher after adding substrate.
The advantage of the invention is that:The ST2 immunologic function test reagents of the present invention can accurately and quickly determine blood of human body etc. ST2 contents in sample.Compared with existing detection reagent in the market, detection reagent of the present invention have fast and easy, high sensitivity, High specificity quantifies the advantages that accurate, is conducive to clinical promote the use of.
Description of the drawings
Fig. 1 is ST2 homogeneous enzyme immunoassay reaction calibration graph.
Fig. 2 is ST2 homogeneous enzyme immunoassay range of linearity figures.
Specific embodiment
The present invention provides a kind of ST2 immunologic function test reagents and its preparation and detection method, to make the object of the invention, technology Scheme and effect are clearer, clear and definite, and the present invention is described in detail below.
The present invention provides a kind of ST2 immunologic function test reagents and its preparation and detection methods.Including:Enzyme mark ST2, for examining Survey the indicator of ST2 antibody-enzyme mark ST2 compounds;Above-mentioned enzyme mark ST2 is coupled by ST2 and glucose dehydrogenase.
Signified " ST2 " refers not only to complete ST2 molecules in the present invention, also includes retaining intact antigen specific binding The ST2 segments or derivative of ability.
A kind of ST2 homogeneous enzyme immunoassay detection reagent, including:Enzyme mark ST2, for detecting ST2 antibody-enzyme mark ST2 compounds Indicator.Indicator is selected from enzymatic reagent, radio isotope reagent, fluorometric reagent and chemical illuminating reagent.Preferably, Indicator is enzymatic reagent, including:The substrate of enzyme mark conjugate and enzyme.Wherein, enzyme mark conjugate includes glucose dehydrogenase-anti- Former conjugate can be obtained by chemical synthesis process.
The application method of above-mentioned ST2 immunologic function test reagents, comprises the following steps:
1)Sample to be tested is contacted with ST2 antibody;
2)According to the combination situation of enzyme mark ST2 in sample to be tested and ST2 antibody, contained using ST2 in indicator judgement sample Amount;The sample to be tested is serum, blood plasma, saliva or urine etc..Preferably, sample to be tested is serum or blood plasma.
Below by specific embodiment, the present invention is described in detail.
Embodiment one:The preparation of glucose dehydrogenase-antigen conjugates
1)Glucose dehydrogenase(GDH)The preparation of solution:
A. the GDH that 10 mg specifications are 100 KU is weighed, room-temperature dissolution is in the solution of 8 mL 50mM PBS, pH=7.4;
2)The activation of ST2
Following chemicals is added to stirring and dissolving in beaker:25 mg ST2,20mg 1- ethyls -3- (- 3- dimethylaminopropyls) Carbodiimide, 5mg N- hydroxy thiosuccinimides are dissolved in 2 mL 2- (N- morpholines) ethanesulfonic acid, stir at room temperature Dissolving is mixed, is reacted 30 minutes;
3)The coupling of GDH and ST2
A. the ST2 solution of above-mentioned activation is added dropwise in the GDH solution in above-mentioned stirring;
B 2-8 DEG C are stirred overnight;
4)Purify the enzyme-labelled antigen of coupling
The enzyme-labelled antigen being coupled by G-25 gel chromatographies column purification, obtains glucose dehydrogenase-ST2 conjugates, at 2-8 DEG C Storage.
Embodiment two:The preparation of ST2 homogeneous enzyme immunoassay detection reagents
ST2 homogeneous enzyme immunoassay detection reagents, including:Enzyme mark ST2, tried for detecting the instruction of ST2 antibody-enzyme mark ST2 compounds Agent.Indicator is selected from enzymatic reagent, radio isotope reagent, fluorometric reagent and chemical illuminating reagent.Preferably, indicator For enzymatic reagent, including:The substrate of enzyme mark conjugate and enzyme.Wherein, enzyme mark conjugate includes glucose dehydrogenase-antigen coupling Object can be obtained by chemical synthesis process.
ST2 homogeneous enzyme immunoassay detection reagent before the use, in order to avoid the enzyme mark conjugate in indicator and enzyme Substrate reacts, and the substrate of enzyme mark conjugate and enzyme is separated, therefore ST2 homogeneous enzyme immunoassay detection reagent includes two The reagent that kind is provided separately, it is specific as follows:
1. the preparation of reagent 1:By 3.588 g(10 mM)Nicotinamide adenine dinucleotide NAD, 1.802 g of oxidation state (10mM)Homogeneous zymolyte is made with the phosphate buffer dissolving of 50 mM, pH 8.0 of 1L in glucose;ST2 antibody is added to It states in homogeneous zymolyte, the volume ratio of antibody and homogeneous zymolyte is 1:100~1:10000, it is specific in the present embodiment to compare Example is 1:600.
2. the preparation of reagent 2:Glucose dehydrogenase-ST2 the conjugates of preparation are added to the phosphoric acid of 50 mM, pH 8.0 In salt buffer, the volume ratio of above-mentioned conjugate and phosphate buffer is 1:100~1:10000, in the present embodiment specifically Ratio be 1:1600.
The application method of above-mentioned ST2 homogeneous enzyme immunoassay detection reagent, comprises the following steps:
1)Sample to be tested is contacted with ST2 antibody;
2)According to the combination situation of enzyme mark ST2 in sample to be tested and ST2 antibody, contained using ST2 in indicator judgement sample Amount;
Specifically, sample to be tested is added in reagent 1 during detection, the ST2 in sample to be tested occurs with the ST2 antibody in reagent 1 Specific binding, generates anti-ST2 antibody-ST2 compounds;Reagent 2 is added, at this time glucose dehydrogenase-the ST2 in reagent 2 Conjugate is mixed with the substrate of the enzyme in reagent 1, contacted, and enzymatic reaction occurs, and forms detection ST2 antibody-enzyme mark ST2 compounds Indicator, indicator judges ST2 in sample to be tested according to the combination situation of ST2 in sample to be tested and above-mentioned ST2 antibody Content.
Due to the ST2 competitive binding ST2 antibody in glucose dehydrogenase-antigen conjugates and sample to be tested, so, it treats The amount of ST2 is more in test sample sheet, and the amount of the glucose dehydrogenase-antigen conjugates to dissociate in homogeneous enzyme solutions is more, and enzymatic is anti- Should be faster, cause OD340 Rise.
Above-mentioned sample to be tested is physiology sample, such as serum, blood plasma, urine, saliva etc., as a preferred solution, Above-mentioned sample to be tested is serum or blood plasma.
Embodiment three:ST2 homogeneous enzyme immunoassay detection reagent reacts calibration curve.
1)ST2 calibration objects are prepared:Commercially available people ST2 recombinant proteins are dissolved in the solution of similar human serum matrix(NaCl 0.9%, BSA0.2%, NaN30.1%, Tris-HCl pH 7.4)In, the calibration object of various concentration is made.With Shenzhen person of outstanding talent ground China Bio tech ltd's ST2 calibration objects are opened up as primary standard, the calibration object of various concentration is examined respectively using its ST2 kit It surveys 10 times, average is obtained, obtains the concentration of ST2 calibration objects:10,20,37.5,75,150,300 ng/mL.
2)Biochemical Analyzer detects:By taking Hitachi 7170 operates as an example:Measure wavelength is 340 nm, takes various concentration respectively Calibration object solution(15μL), add in ST2 R1Reagent(160μL), mixing adds ST2 R2Reagent(40μL), after mixing, measure The OD of different time points340Light absorption value calculates reaction rate during different calibration object concentration, needs constantly to adjust in actual mechanical process The volume ratio of whole reagent 1 and reagent 2, while survey luminous point is adjusted, comparatively ideal reaction normal graph is finally drawn, per Guan Chong Repetition measurement is 3 times fixed, and using the average value of 3 absorbance difference Δ A measured of each calibration pipe as ordinate, corresponding calibration object concentration is Abscissa draws " concentration-absorbance difference " calibration curve(See Fig. 1).
Test serum or plasma sample are taken, is measured in the same method the absorbance difference of sample, substitutes into calibration curve, you can calculate The content of ST2 in sample to be tested.If the concentration of ST2 exceeds calibration curve scope in serum or blood plasma, sample need to be carried out dilute Detect to ensure the accuracy of testing result after releasing again.
This detection reagent is applicable not only to Hitachi 7170, applies also for semi-automatic, the full-automatic life of other brands and model Change analyzer, design parameter can be adjusted according to instrument.
Example IV:The range of linearity determines
With the ST2 high concentration samples close to the range of linearity upper limit(320 ng/mL), it is pressed 1/2,1/4,1/8 with physiological saline, 1/16,1/32,1/64 dilution, is configured to 6 diluted concentrations altogether(xi)Solution, with the Biochemical Analyzer detection method survey Fixed each diluted sample concentration.Each diluted concentration is tested 3 times, and the average of each diluted concentration testing result is obtained respectively(yi). With diluted concentration(xi)For independent variable, to measure average(yi)Equation of linear regression is obtained for dependent variable, according to formula(1)It calculates The correlation coefficient r of linear regression, the results show regression equation are y=1.0213x-0.2237, and correlation coefficient r=0.9998 shows Reagent of the present invention good relationship in the 5 ng/mL320 ng/mL ranges of linearity(See Fig. 2).
Since the detection process of the present invention is completed by instrument is full-automatic, so to the of less demanding of testing staff, easily In realizing and promote the use of.
It should be noted that obviously the invention is not restricted to the details of above-mentioned exemplary embodiment, the scope of the present invention is by institute Attached claim rather than above description limit, it is intended that will fall within the meaning and scope of the equivalent requirements of the claims All changes are included in the scope of patent protection of the present invention.
In addition, above-described is only the preferred embodiments of the invention, for the technical staff in this technology neck city, Without departing from the principle of the present invention, several modifications and adaptations can also be done, these improved adjustment also should be regarded as this The protection domain of invention.

Claims (7)

1. a kind of Soluble growth stimulates 2 protein immunization detection reagent of expressing gene and its preparation and detection method, feature to exist In:Enzyme mark Soluble growth stimulates 2 albumen of expressing gene, stimulates 2 protein antibodies of expressing gene-enzyme for detecting Soluble growth Marking Soluble growth stimulates the indicator of 2 albumen composition of expressing gene.
2. Soluble growth according to claim 1 stimulates the immunologic function test reagent of 2 albumen of expressing gene, feature exists In:The enzyme mark Soluble growth stimulates 2 albumen of expressing gene by Soluble growth 2 albumen of expressing gene and glucose to be stimulated to take off Hydrogen enzyme is coupled.
3. Soluble growth according to claim 1 stimulates 2 protein immunization detection reagent of expressing gene, it is characterised in that: The indicator is selected from enzymatic reagent, including:The substrate of enzyme mark conjugate and enzyme;Above-mentioned enzyme mark conjugate includes glucose dehydrogenation Enzyme-Soluble growth stimulates 2 protein conjugate of expressing gene;The substrate of above-mentioned enzyme is glucose.
4. a kind of Soluble growth stimulates 2 protein immunization detection reagent of expressing gene and preparation method thereof, which is characterized in that including Following steps:
(1)Glucose dehydrogenase-Soluble growth stimulates the preparation of 2 protein conjugate of expressing gene:Prepare glucose dehydrogenase (GDH)Solution, Soluble growth stimulate the activation of 2 albumen of expressing gene, and GDH and Soluble growth are stimulated 2 egg of expressing gene White coupling purifies the enzyme-labelled antigen of coupling;
(2)Soluble growth stimulates the preparation of 2 albumen homogeneous enzyme immunoassay detection reagent of expressing gene:
The preparation of reagent 1:By Soluble growth 2 protein antibodies of expressing gene and homogeneous zymolyte is stimulated to mix;
The preparation of reagent 2:2 protein conjugate of expressing gene and phosphate-buffered are stimulated by glucose dehydrogenase-Soluble growth Liquid mixes.
5. a kind of Soluble growth according to claim 5 stimulates the preparation side of 2 protein immunization detection reagent of expressing gene Method, which is characterized in that the step(1)Detailed process is:
1)Glucose dehydrogenase(GDH)The preparation of solution:
A. the GDH that 5-20 mg specifications are 100-300 KU is weighed, room-temperature dissolution is in 6-10 mL PBS solutions, pH 7.0- 10.0;
2)Soluble growth stimulates the activation of 2 albumen of expressing gene
Following chemicals is added to stirring and dissolving in beaker:10-30 mg Soluble growths stimulate 2 albumen of expressing gene, 10- 40 mg 1- ethyls -3- (- 3- dimethylaminopropyls) carbodiimides(EDC), 2-10 mg N- hydroxy thiosuccinimides (NHS), it is dissolved in 2- (N- morpholines) ethanesulfonic acid(MES)In solution, stirring and dissolving, reacts 15-60 minutes at room temperature;
3)GDH stimulates the coupling of 2 albumen of expressing gene with Soluble growth
A. the GDH for 2 protein solution of expressing gene being stimulated to be added dropwise in above-mentioned stirring the Soluble growth of above-mentioned activation is molten In liquid;
B. it is stirred overnight for 2-8 DEG C;
4)Purify the enzyme-labelled antigen of coupling
The enzyme-labelled antigen being coupled by G-25 gel chromatographies column purification, obtaining glucose dehydrogenase-Soluble growth stimulates expression 2 protein conjugate of gene, stores at 2-8 DEG C.
6. a kind of Soluble growth according to claim 5 stimulates the preparation side of 2 protein immunization detection reagent of expressing gene Method, which is characterized in that step(2)Detailed process it is as follows:
The preparation of reagent 1:By nicotinamide adenine dinucleotide NAD, the 0.5-3 g glucose 0.5-2L of 2-5 g oxidation state Homogeneous zymolyte is made in phosphate buffer dissolving;Soluble growth stimulation 2 protein antibodies of expressing gene are added to above-mentioned homogeneous In zymolyte, the volume ratio of antibody and homogeneous zymolyte is 1:100~1:10000;
The preparation of reagent 2:2 protein conjugate of expressing gene is stimulated to be added to phosphorus the glucose dehydrogenase of preparation-Soluble growth In phthalate buffer, the volume ratio of above-mentioned conjugate and phosphate buffer is 1:100~1:10000.
7. stimulate 2 protein immunization detection reagent of expressing gene using the Soluble growth described in 4 any one of Claims 1-4 Detection method, which is characterized in that comprise the following steps:
1)With Soluble growth 2 protein antibodies of expressing gene is stimulated to contact sample to be tested;
2)According to Soluble growth in sample to be tested 2 albumen of expressing gene is stimulated to stimulate 2 albumen of expressing gene with Soluble growth The combination situation of antibody stimulates the content of 2 albumen of expressing gene using Soluble growth in indicator judgement sample;It is described to treat Test sample sheet is serum, blood plasma, saliva or urine.
CN201711403036.4A 2017-12-22 2017-12-22 A kind of Soluble growth stimulates 2 protein immunization detection reagent of expressing gene and its preparation and detection method Pending CN108107222A (en)

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Application publication date: 20180601