CN109239332A - A kind of kit of auxiliary detection sub-health population cancer base antigen - Google Patents
A kind of kit of auxiliary detection sub-health population cancer base antigen Download PDFInfo
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/531—Production of immunochemical test materials
- G01N33/532—Production of labelled immunochemicals
- G01N33/535—Production of labelled immunochemicals with enzyme label or co-enzymes, co-factors, enzyme inhibitors or enzyme substrates
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/536—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase
- G01N33/537—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase with separation of immune complex from unbound antigen or antibody
- G01N33/538—Immunoassay; Biospecific binding assay; Materials therefor with immune complex formed in liquid phase with separation of immune complex from unbound antigen or antibody by sorbent column, particles or resin strip, i.e. sorbent materials
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
- G01N33/54326—Magnetic particles
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- 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
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/574—Immunoassay; Biospecific binding assay; Materials therefor for cancer
- G01N33/57484—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving compounds serving as markers for tumor, cancer, neoplasia, e.g. cellular determinants, receptors, heat shock/stress proteins, A-protein, oligosaccharides, metabolites
- G01N33/57492—Immunoassay; Biospecific binding assay; Materials therefor for cancer involving compounds serving as markers for tumor, cancer, neoplasia, e.g. cellular determinants, receptors, heat shock/stress proteins, A-protein, oligosaccharides, metabolites involving compounds localized on the membrane of tumor or cancer cells
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- 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
- G01N33/58—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances
- G01N33/581—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving labelled substances with enzyme label (including co-enzymes, co-factors, enzyme inhibitors or substrates)
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
- G01N2333/46—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans from vertebrates
- G01N2333/47—Assays involving proteins of known structure or function as defined in the subgroups
Abstract
The invention discloses the kits that a kind of auxiliary for belonging to technical field of biological detects sub-health population cancer base antigen.The kit includes carcinomebryonic antigen calibration object, cancer base antigen quality-control product, the Tris buffer of Cea Monoclonal Antibodies and bovine serum albumin(BSA) containing biotin labeling 1, the buffer of Cea Monoclonal Antibodies and bovine serum albumin(BSA) containing alkali phosphatase enzyme mark 2, the Tris buffer of magnetic particle and bovine serum albumin(BSA) containing marked by streptavidin 3, chemical substrate and cleaning solution.The kit high sensitivity, easy to operate, no radiocontamination, low in cost, reaction process is fast and reliable, improves sensitivity and linear measurement range, it realizes quantitative determination, provides a suitable theoretical judgment for the state of an illness of sub-health population common disease and broad-spectrum tumor screening.
Description
Technical field
The invention belongs to technical field of biological, and in particular to a kind of to assist detecting sub-health population cancer base antigen
Kit.
Background technique
Inferior health refers to a kind of state that human body is between health and disease.In sub-health state person, cannot reach
The standard of health shows as the symptom that the vigor in certain time reduces, function and adaptability decline, does not meet modern doctor
The clinic or subclinical inflammation standard for there are related disorders are learned, but the proposition of inferior health idea has highly important medical value
And clinical meaning, because it can promote the critical period of the prevention and treatment of disease to move forward.In recent years, due to environmental pollution, swash
Strong competition, people's lives and work rhythm are constantly accelerated, and the pressure from society and various aspects is continuously increased, along with suction
Cigarette excessive drinking, high heat high fat diet lack the bad habits such as manual labor, life is irregular, night life is excessive, inferior health
The ratio that state crowd accounts for population of China gradually increases.Inferior health has severely impacted people's lives, study and work
Make, becomes pervasive social problem, become the restraining factors of social development, harmfulness is considered century people by medical field
Class health formidable enemy.
Cancer base antigen (carcinoembryonic antigen, CEA) is a kind of with the decision of human embryos antigen-specific
The sugar antigen of cluster is present in the cancer cell surface that endoderm cell differentiates, and is the structural proteins of cell membrane.Cancer embryo
Antigen is important tumor associated antigen, it belongs to non-organ specificity tumor associated antigen, is primarily present in Rectum and colon cancer
In tissue and fetus intestinal mucosa, the Specific marker of early diagnosis colon cancer and the carcinoma of the rectum can be used as, through a large amount of clinical
Practice finds that the malignant tumour CEA value of not only gastrointestinal tract can increase, in the serum of breast cancer, lung cancer and other malignant tumours
In also have raising.CEA raising is common in colorectal cancer, cancer of pancreas, gastric cancer, breast cancer, medullary carcinoma of thyroid gland, liver cancer, lung cancer, ovum
Nest cancer, urological cancer etc., and smoke, the gestational period and cardiovascular disease, diabetes, enteron aisle diverticulitis, rectal polyp, colon
Inflammation, pancreatitis, cirrhosis, hepatitis, pulmonary disease etc., 15%~53% patients serum CEA can also be increased.The ginseng of general CEA
It examines range and is less than 5ng/Ml, when the constant height of CEA concentration or numerical value are more than normal 56 times, then explanation has lesion
Remaining or progress, also or prognosis mala, it is believed that have metastatic lesion.Therefore, it is swollen to be not only a kind of wide spectrum for carcinomebryonic antigen
Tumor markers also provide a suitable theoretical judgment to the state of an illness of some common diseases, be malignant tumour antidiastole,
State of illness monitoring, therapeutic evaluation etc., provide important clinical value.
The universal law of the life and health development of people is: life and health period --- sub-health state initial stage --- is sub- strong
The Kang Fazhan reversible refunding, --- the disease controllable phase --- disease develops the deterioration phase --- was dead.When people are in inferior health development
When the reversible refunding, the institutional framework of the cell of the internal body of people and various internal organs, all different degrees of gradually changes
, if be chronically under sub-health state, a possibility that canceration occurs for cell, is greatly increased, and the detection of cancer base antigen is
Our screenings for the tumour of sub-health population, assist it to judge inferior health degree, have great importance, therefore we
It needs to be timed the cancer base antigen of the sub-health state of people or periodically carry out testing and evaluation.Nowadays, carcinomebryonic antigen is normal
The detection method seen includes radio immunoassay and indirect immunofluorescence, and radioimmunoassay fluorescence method is due to higher cost
And need tip complex device to prevent radiocontamination, indirect immunofluorescence it is complicated for operation and can only carry out qualitative analysis and
Sensitivity is low, and quantitative analysis cannot be carried out for the carcinomebryonic antigen of sub-health population, it is therefore desirable to and it is a kind of easy to operate, at
This cheap, high sensitivity, no radiocontamination and the method for being able to carry out quantitative analysis.
Summary of the invention
For overcome the deficiencies in the prior art, the present invention provides a kind of high sensitivities, easy to operate, no radiocontamination,
Low-cost kit, invention are achieved through the following technical solutions.
The purpose of the present invention is to provide the kits and preparation method of auxiliary detection sub-health population cancer base antigen.
A kind of kit of auxiliary detection sub-health population cancer base antigen, the kit include:
1) carcinomebryonic antigen calibration object, the phosphate buffered saline solution containing carcinomebryonic antigen and bovine serum albumin(BSA), the cow's serum
The phosphate buffered saline solution of albumin includes following component, bovine serum albumin(BSA) 30-34g, potassium dihydrogen phosphate powder 0.2-
0.4g, disodium hydrogen phosphate powder 1-2g, sodium chloride powder 7-9g, potassium chloride powder 0.2-0.4g, the salt of concentration 36%~38%
Sour 0.1-1mL, the Proclin300 solution 0.1-0.2mL of concentration 98%, deionized water 700-900mL, the carcinomebryonic antigen
Calibration object is 6 horizontal liquid calibration objects, in 6 horizontal calibration objects the concentration containing carcinomebryonic antigen be respectively 0,5,
25,100,500 and 1000ng/mL, pH 7.2-7.4;
2) cancer base antigen quality-control product, the phosphate buffered saline solution containing one containing carcinomebryonic antigen and bovine serum albumin(BSA), the ox
Sero-abluminous phosphate buffered saline solution is that the phosphate-buffered salt of the bovine serum albumin(BSA) of the carcinomebryonic antigen calibration object is molten
Liquid, the carcinomebryonic antigen quality-control product include 2 horizontal liquid calibration objects, carcinomebryonic antigen in 2 horizontal liquid quality controls
Target value concentration be 10ng/mL and 100ng/mL, pH 7.2-7.4;
3) reagent 1, the Tris buffer of Cea Monoclonal Antibodies and bovine serum albumin(BSA) containing biotin labeling
No. 1, Tris buffer 1 of the bovine serum albumin(BSA) includes following component, bovine serum albumin(BSA) 3-5g, tromethamine
Powder 6-10g, sodium chloride powder 3-5g, the hydrochloric acid 0.1-1mL of concentration 36%~38%, the Proclin300 of concentration 98% are molten
Liquid 0.1-0.2mL, deionized water 700-900mL;
4) reagent 2, the buffering of Cea Monoclonal Antibodies and bovine serum albumin(BSA) containing alkali phosphatase enzyme mark
Liquid 2, buffer 2 of the bovine serum albumin(BSA) include following component, 4- hydroxyethyl piperazineethanesulfonic acid powder 4-6g, chlorine
Change sodium powder end 5-8g, bovine serum albumin(BSA) 3-5g, zinc chloride powder 0.05-0.1g, magnesium chloride powder 0.05-0.1g, concentration
36%~38% hydrochloric acid 0.1-1mL, Proclin300 the solution 0.1-0.2mL, deionized water 700- of concentration 98%
900mL;
5) Magneto separate reagent, the Tris buffer 3 of magnetic particle and bovine serum albumin(BSA) containing marked by streptavidin
Number, Tris buffer 3 of the bovine serum albumin(BSA) include following component, bovine serum albumin(BSA) 3-5g, cow's serum 30-
50g, tromethamine powder 6-10g, sodium chloride powder 5-8g, the hydrochloric acid 0.1-1mL of concentration 36%~38%, concentration 98%
Proclin300 solution 0.1-0.2mL, deionized water 700-900mL;
6) chemical substrate, the chemical substrate include following component, tromethamine powder 15-25g, ethylenediamine tetrem
Acid disodium powder 15-25g, sodium chloride powder 15-20g, dioxane compound 0.2-0.6g, lauryl sodium sulfate
0.05-0.1g, glycerine 35-45g, sodium sulfite 0.05-0.1g, the Proclin300 solution 0.1-0.2mL of concentration 98%,
Deionized water 700-900mL;
7) cleaning solution, the cleaning solution include following component, tromethamine powder 6-10g, sodium chloride powder 5-8g,
Polyoxyethylene 20 sorbitan monolaurate powder 3-5g, Triton X-100 3-5g, concentration 36%~38%
Hydrochloric acid 0.1-1mL, deionized water 700-900mL, the pH value of the cleaning solution are 7.5-8.0.
Preferably, in the kit of the auxiliary detection sub-health population cancer base antigen, the carcinomebryonic antigen calibration
Product, the carcinomebryonic antigen quality-control product, the reagent 1, the reagent 2, the Magneto separate reagent, the chemical substrate
Storage temperature is 2-8 DEG C, and the storage temperature of the cleaning solution is 10-30 DEG C of room temperature, and the kit avoids direct sunlight.
Preferably, biological in the reagent 1 in the kit of the auxiliary detection sub-health population cancer base antigen
The concentration of the Cea Monoclonal Antibodies of element label is 0.1-0.3 μ g/mL, pH value 7.0-7.5, alkali in the reagent 2
The concentration of the Cea Monoclonal Antibodies of acid phosphatase label is 0.02-0.1 μ g/mL, pH value 7.5-8.0, the magnetic point
Concentration from reagent is 0.5ng/mL, and pH value 7.9-8.1, the diameter of the magnetic particle is 0.5-3.5 μm, kernel four
Fe 3 O, surface are wrapped in the polymer containing carboxyl, and the concentration of dioxane compound is in the chemical substrate
0.5mg/mL, pH value 9.2-9.5.
Preferably, in the kit of the auxiliary detection sub-health population cancer base antigen, the bovine serum albumin(BSA)
Tris buffer 1, in Tris buffer 3 of the buffer of the bovine serum albumin(BSA) 2 and the bovine serum albumin(BSA)
Casein 3-5g can also be added.
Preferably, in the kit of the auxiliary detection sub-health population cancer base antigen, the bovine serum albumin(BSA)
Tris buffer 1, in Tris buffer 3 of the buffer of the bovine serum albumin(BSA) 2 and the bovine serum albumin(BSA)
N- hydroxysuccinimide powder 3-5g can also be added.
Preferably, in the kit of the auxiliary detection sub-health population cancer base antigen, the bovine serum albumin(BSA)
Tris buffer 1, in Tris buffer 3 of the buffer of the bovine serum albumin(BSA) 2 and the bovine serum albumin(BSA)
(Na can also be added2.3K0.7)(B6O10)(N O3) powder 3-5g.
The preparation method of the kit of above-mentioned auxiliary detection sub-health population cancer base antigen, carries out in accordance with the following steps:
(1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.2-0.4g, disodium hydrogen phosphate powder 1-2g, chlorine
Change sodium powder end 7-9g, potassium chloride powder 0.2-0.4g, the Proclin300 solution 0.1-0.2mL of concentration 98%, deionized water
700-900mL is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid 0.1-1mL of concentration 36%~38% is added, by pH
Value is adjusted to 7.2-7.4, and bovine serum albumin(BSA) 30-34g is added to the container, is stirred well to and is completely dissolved, with 0.3 μm of aperture
Filter filtering after bovine serum albumin(BSA) phosphate buffered saline solution, the phosphoric acid of cancer base antigen bovine serum albumin(BSA) is delayed
Rushing salting liquid and being diluted to each concentration point is 0,5,25,100,500 and 1000ng/mL;
(2) cancer base antigen quality-control product is prepared: cancer base antigen is dilute with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Releasing to each concentration point is 10ng/mL and 100ng/mL;
(3) reagent preparation 1: by tromethamine powder 6-10g, sodium chloride powder 3-5g, concentration 98%
Proclin300 solution 0.1-0.2mL, deionized water 700-900mL are added to the container, are stirred well to and are completely dissolved, and are added
The hydrochloric acid 0.1-1mL of concentration 36%~38%, is adjusted to 7.0-7.5 for pH value, and bovine serum albumin(BSA) 3-5g is added to the container,
It is stirred well to and is completely dissolved, Tris buffer 1 of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture, it is conventional
Method prepares Cea Monoclonal Antibodies --- the solution of biotin conjugate, with the Tris buffer 1 of bovine serum albumin(BSA)
Number containing Cea Monoclonal Antibodies --- the solution of biotin conjugate is diluted to 0.1-0.3 μ g/mL, obtains reagent 1
Number;
(4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 4-6g, sodium chloride powder 5-8g, chlorination zinc powder
Last 0.05-0.1g, magnesium chloride powder 0.05-0.1g, the Proclin300 solution 0.1-0.2mL of concentration 98%, deionized water
700-900mL is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid 0.1-1mL of concentration 36%~38% is added, by pH
Value is adjusted to 7.5-8.0, and bovine serum albumin(BSA) 3-5g is added to the container, is stirred well to and is completely dissolved, with 0.3 μm of aperture
Buffer 2 of bovine serum albumin(BSA) are obtained after filter filtering, conventional method prepares Cea Monoclonal Antibodies --- alkaline phosphorus
The solution of sour enzyme attachment will contain Cea Monoclonal Antibodies with buffer 2 of bovine serum albumin(BSA) --- alkaline phosphorus
The solution of sour enzyme attachment is diluted to 0.02-0.1 μ g/mL, obtains reagent 2;
(5) Magneto separate reagent is prepared: by tromethamine powder 6-10g, sodium chloride powder 5-8g, deionized water 700-
900mL is added to the container, and is stirred well to and is completely dissolved, by bovine serum albumin(BSA) 3-5g, cow's serum 30-50g, concentration 98%
Proclin300 solution 0.1-0.2mL be added to the container, be stirred well to and be completely dissolved, concentration 36%~38% is added
PH value is adjusted to 7.9-8.1 by hydrochloric acid 0.1-1mL, and the Tris that bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture is slow
Fliud flushing 3, conventional method prepares magnetic particle --- the solution of Streptavidin attachment, with the Tris of bovine serum albumin(BSA)
Buffer 3 will contain magnetic particle --- and the solution of Streptavidin attachment is diluted to 0.5ng/mL, obtains Magneto separate examination
Agent;
(6) chemical substrate is prepared: by tromethamine powder 15-25g, disodium ethylene diamine tetraacetate powder 15-25g, chlorine
Change sodium powder end 15-20g, lauryl sodium sulfate 0.05-0.1g, glycerine 35-45g, sodium sulfite 0.05-0.1g, concentration
98% Proclin300 solution 0.1-0.2mL, deionized water 700-900mL are added to the container, and are stirred well to completely molten
Solution is added the hydrochloric acid 0.1-1mL of concentration 36%~38%, the pH value of solution is adjusted to 9.2-9.5, with 0.3 μm of the filter in aperture
Dioxane compound 0.2-0.6g is added after filtering under light protected environment, being configured to concentration is 0.5mg/mL, obtains chemical bottom
Object;
(7) cleaning solution is prepared: by tromethamine powder 6-10g, sodium chloride powder 5-8g, polyoxyethylene sorbitan
Alcohol monolaurate powder 3-5g, Triton X-100 3-5g, deionized water 700-900mL are added to the container, sufficiently
Stirring is added the hydrochloric acid 0.1-1mL of concentration 36%~38%, the pH value of solution is adjusted to 7.5-8.0, uses hole to being completely dissolved
Cleaning solution is obtained after the filter filtering that 0.3 μm of diameter.
Casein 3-5g can be also added in the step (5) in the step (3), the step (4).
N- hydroxysuccinimide powder 3- can be also added in the step (5) in the step (3), the step (4)
5g。
(Na can be also added in the step (5) in the step (3), the step (4)2.3K0.7)(B6 O10)(N O3) powder
Last 3-5g.
Testing principle of the invention: by sample to be tested, No. 1 mixing of reagent is incubated, the biotin labeling in reagent 1
Then the magnetic particle for being coated with Streptavidin is added in conjunction with specimen needle carcinomebryonic antigen molecule in Cea Monoclonal Antibodies,
Immune complex is adsorbed to magnetic particle surface.No. 2 mixing temperature of reagent are added after taking out unbonded antibody and impurity in washing
It educates, the Cea Monoclonal Antibodies and the carcinomebryonic antigen in above-mentioned immune complex of the alkali phosphatase enzyme mark in reagent 2
Molecule combines.Chemical substrate, alkaline phosphatase catalytic chemistry substrate hair is added after taking out unbonded antibody and impurity in washing
Light measures relative luminous intensity.Luminous intensity in a certain range is directly proportional to carcinomebryonic antigen concentration, just by interpolation method
It can be from the content for reading carcinomebryonic antigen in sample to be tested on standard curve.
Beneficial effects of the present invention: the kit of auxiliary detection sub-health population cancer base antigen prepared by the present invention uses
The reaction pattern of double antibody sandwich method, high sensitivity is easy to operate, and no radiocontamination, low in cost, reaction process quickly may be used
It leans on, realizes high-throughput detection and quantitative determination, advise the state of an illness of disease for sub-health population and broad-spectrum tumor screening provides a conjunction
Suitable theoretical judgment.It the kit alignment product, quality-control product, reagent 1, reagent 2, Magneto separate reagent, chemical substrate and washes
Washing liquid etc. is the optimization formula under the reaction system, fully ensures that validity period and detection performance.
Specific embodiment
The present invention will be further described combined with specific embodiments below.
Embodiment 1
A kind of kit of auxiliary detection sub-health population cancer base antigen, consists of the following compositions: carcinomebryonic antigen calibration
Product, cancer base antigen quality-control product, reagent 1, reagent 2, Magneto separate reagent, chemical substrate and cleaning solution.
The preparation method of the kit of above-mentioned auxiliary detection sub-health population cancer base antigen, carries out in accordance with the following steps:
(1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.3g, disodium hydrogen phosphate powder 1.5g, sodium chloride
Powder 8g, potassium chloride powder 0.3g, the Proclin300 solution 0.15g of concentration 98%, deionized water 800g are added to the container,
It is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.3, by bovine serum albumin(BSA)
32g is added to the container, and is stirred well to and is completely dissolved, and the phosphoric acid of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture
Buffer salt solution, by the phosphate buffered saline solution of cancer base antigen bovine serum albumin(BSA) be diluted to each concentration point be 0,5,25,
100,500 and 1000ng/mL;
(2) cancer base antigen quality-control product is prepared: cancer base antigen is dilute with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Releasing to each concentration point is 10ng/mL and 100ng/mL;
(3) reagent preparation 1: by tromethamine powder 8g, sodium chloride powder 4g, the Proclin300 of concentration 98%
Solution 0.15g, deionized water 800g, is added to the container, and is stirred well to and is completely dissolved, and the salt of concentration 36%~38% is added
PH value is adjusted to 7.3 and is added to the container bovine serum albumin(BSA) 4g, is stirred well to and be completely dissolved by sour 0.5g, with 0.3 μ of aperture
Tris buffer 1 of bovine serum albumin(BSA) is obtained after the filter filtering of m, it is anti-that conventional method prepares carcinomebryonic antigen monoclonal
The solution of body --- biotin conjugate will be resisted with the Tris buffer 1 of bovine serum albumin(BSA) containing carcinomebryonic antigen monoclonal
The solution of body --- biotin conjugate is diluted to 0.2 μ g/mL, obtains reagent 1;
(4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 5g, sodium chloride powder 0.65g, zinc chloride powder
0.075g, container is added in magnesium chloride powder 0.075g, Proclin300 the solution 0.15g, deionized water 800g of concentration 98%
In, it is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.7, ox blood is pure
Albumen 4g is added to the container, and is stirred well to and is completely dissolved, and obtains bovine serum albumin(BSA) after being filtered with 0.3 μm of the filter in aperture
Buffer 2, Cea Monoclonal Antibodies --- the solution of alkaline phosphatase attachment uses cow's serum for conventional method preparation
Buffer 2 of albumin will contain Cea Monoclonal Antibodies --- and the solution of alkaline phosphatase attachment is diluted to
0.06 μ g/mL obtains reagent 2;
(5) it prepares Magneto separate reagent: tromethamine powder 8g, sodium chloride powder 6.5g, deionized water 800g is added
In container, it is stirred well to and is completely dissolved, the Proclin300 of bovine serum albumin(BSA) 4g, cow's serum 40g, concentration 98% is molten
Liquid 0.15g is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to
8.0, Tris buffer 3 of bovine serum albumin(BSA) are obtained after being filtered with 0.3 μm of the filter in aperture, conventional method is prepared magnetic micro-
Grain --- solution of Streptavidin attachment will contain magnetic particle with Tris buffer 3 of bovine serum albumin(BSA) ---
The solution of Streptavidin attachment is diluted to 0.5ng/mL, obtains Magneto separate reagent;
(6) chemical substrate is prepared: by tromethamine powder 20g, disodium ethylene diamine tetraacetate powder 20g, sodium chloride powder
Last 18g, lauryl sodium sulfate 0.075g, glycerine 40g, sodium sulfite 0.075g, the Proclin300 solution of concentration 98%
0.15g, deionized water 800g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
The pH value of solution is adjusted to 9.3 by 0.5g, and dioxane is added under light protected environment after being filtered with 0.3 μm of the filter in aperture
Object 0.4g is closed, being configured to concentration is 0.5mg/mL, obtains chemical substrate;
(7) cleaning solution is prepared: by tromethamine powder 8g, sodium chloride powder 6.5g, polyoxyethylene sorbitan list
Laurate powder 4g, Triton X-100 4g, deionized water 800g are added to the container, are stirred well to completely molten
Solution is added the hydrochloric acid 0.5g of concentration 36%~38%, the pH value of solution is adjusted to 7.7, after being filtered with 0.3 μm of the filter in aperture
Obtain cleaning solution.
Embodiment 2
A kind of kit of auxiliary detection sub-health population cancer base antigen, consists of the following compositions: carcinomebryonic antigen calibration
Product, cancer base antigen quality-control product, reagent 1, reagent 2, Magneto separate reagent, chemical substrate and cleaning solution.
The preparation method of the kit of above-mentioned auxiliary detection sub-health population cancer base antigen, carries out in accordance with the following steps:
(1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.2g, disodium hydrogen phosphate powder 1g, sodium chloride powder
Last 7g, potassium chloride powder 0.2g, the Proclin300 solution 0.1g of concentration 98%, deionized water 700g are added to the container, sufficiently
Stirring is added the hydrochloric acid 0.1g of concentration 36%~38%, pH value is adjusted to 7.4, by bovine serum albumin(BSA) 30g to being completely dissolved
It is added to the container, is stirred well to and is completely dissolved, the phosphoric acid that bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture is slow
Rush salting liquid, by the phosphate buffered saline solution of cancer base antigen bovine serum albumin(BSA) be diluted to each concentration point be 0,5,25,100,
500 and 1000ng/mL;
(2) cancer base antigen quality-control product is prepared: cancer base antigen is dilute with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Releasing to each concentration point is 10ng/mL and 100ng/mL;
(3) reagent preparation 1: by tromethamine powder 6g, sodium chloride powder 3g, the Proclin300 of concentration 98%
Solution 0.1g, deionized water 700g, is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
PH value is adjusted to 7.5 by 0.1g, and bovine serum albumin(BSA) 3g is added to the container, is stirred well to and is completely dissolved, with 0.3 μm of aperture
Filter filtering after Tris buffer 1 of bovine serum albumin(BSA), conventional method prepares Cea Monoclonal Antibodies ---
The solution of biotin conjugate will contain Cea Monoclonal Antibodies with Tris buffer 1 of bovine serum albumin(BSA) ---
The solution of biotin conjugate is diluted to 0.1 μ g/mL, obtains reagent 1;
(4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 4g, sodium chloride powder 5g, zinc chloride powder
0.05g, magnesium chloride powder 0.05g, the Proclin300 solution 0.1g of concentration 98%, deionized water 700g are added to the container,
It is stirred well to and is completely dissolved, the hydrochloric acid 0.1g of concentration 36%~38% is added, pH value is adjusted to 8.0, by bovine serum albumin(BSA)
3g is added to the container, and is stirred well to and is completely dissolved, and the buffering of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture
Liquid 2, Cea Monoclonal Antibodies --- the solution of alkaline phosphatase attachment uses bovine serum albumin for conventional method preparation
White buffer 2 will contain Cea Monoclonal Antibodies --- and the solution of alkaline phosphatase attachment is diluted to 0.02 μ g/
ML obtains reagent 2;
(5) it prepares Magneto separate reagent: tromethamine powder 6g, sodium chloride powder 5g, deionized water 700g being added and held
It in device, is stirred well to and is completely dissolved, by bovine serum albumin(BSA) 3g, cow's serum 30g, the Proclin300 solution of concentration 98%
0.1g is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid 0.1g of concentration 36%~38% is added, pH value is adjusted to
8.1, Tris buffer 3 of bovine serum albumin(BSA) are obtained after being filtered with 0.3 μm of the filter in aperture, conventional method is prepared magnetic micro-
Grain --- solution of Streptavidin attachment will contain magnetic particle with Tris buffer 3 of bovine serum albumin(BSA) ---
The solution of Streptavidin attachment is diluted to 0.5ng/mL, obtains Magneto separate reagent;
(6) chemical substrate is prepared: by tromethamine powder 15g, disodium ethylene diamine tetraacetate powder 15g, sodium chloride powder
Last 15g, lauryl sodium sulfate 0.05g, glycerine 35g, sodium sulfite 0.05g, the Proclin300 solution of concentration 98%
0.1g, deionized water 700g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid 0.1g of concentration 36%~38% is added,
The pH value of solution is adjusted to 9.5, dioxane compound is added under light protected environment after being filtered with 0.3 μm of the filter in aperture
0.2g, being configured to concentration is 0.5mg/mL, obtains chemical substrate;
(7) cleaning solution is prepared: by tromethamine powder 6g, sodium chloride powder 5g, polyoxyethylene sorbitan Dan Yue
Cinnamic acid ester powder 3g, Triton X-100 3g, deionized water 700g are added to the container, are stirred well to and are completely dissolved,
The hydrochloric acid 0.1g of concentration 36%~38% is added, the pH value of solution is adjusted to 8.0, must be washed after being filtered with 0.3 μm of the filter in aperture
Wash liquid.
Embodiment 3
A kind of kit of auxiliary detection sub-health population cancer base antigen, consists of the following compositions: carcinomebryonic antigen calibration
Product, cancer base antigen quality-control product, reagent 1, reagent 2, Magneto separate reagent, chemical substrate and cleaning solution.
The preparation method of the kit of above-mentioned auxiliary detection sub-health population cancer base antigen, carries out in accordance with the following steps:
(1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.4g, disodium hydrogen phosphate powder 2g, sodium chloride powder
Last 9g, potassium chloride powder 0.4g, the Proclin300 solution 0.2g of concentration 98%, deionized water 900g are added to the container, sufficiently
Stirring is added the hydrochloric acid 1g of concentration 36%~38%, pH value is adjusted to 7.2, bovine serum albumin(BSA) 34g is added to being completely dissolved
Enter in container, be stirred well to and be completely dissolved, the phosphoric acid buffer of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture
Salting liquid, by the phosphate buffered saline solution of cancer base antigen bovine serum albumin(BSA) be diluted to each concentration point be 0,5,25,100,
500 and 1000ng/mL;
(2) cancer base antigen quality-control product is prepared: cancer base antigen is dilute with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Releasing to each concentration point is 10ng/mL and 100ng/mL;
(3) reagent preparation 1: by tromethamine powder 10g, sodium chloride powder 5g, the Proclin300 of concentration 98%
Solution 0.2g, deionized water 900g, is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
PH value is adjusted to 7.0 by 1g, and bovine serum albumin(BSA) 5g is added to the container, is stirred well to and is completely dissolved, with 0.3 μm of aperture
Tris buffer 1 of bovine serum albumin(BSA) is obtained after filter filtering, conventional method prepares Cea Monoclonal Antibodies --- it is raw
The solution of object element attachment will contain Cea Monoclonal Antibodies with Tris buffer 1 of bovine serum albumin(BSA) --- and it is raw
The solution of object element attachment is diluted to 0.3 μ g/mL, obtains reagent 1;
(4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 6g, sodium chloride powder 8g, zinc chloride powder
0.1g, magnesium chloride powder 0.1g, the Proclin300 solution 0.2g of concentration 98%, deionized water 900g are added to the container, sufficiently
Stirring is added the hydrochloric acid 1g of concentration 36%~38%, pH value is adjusted to 7.5, bovine serum albumin(BSA) 5g is added to being completely dissolved
It in container, is stirred well to and is completely dissolved, buffer 2 of bovine serum albumin(BSA) are obtained after being filtered with 0.3 μm of the filter in aperture,
Conventional method prepares Cea Monoclonal Antibodies --- the solution of alkaline phosphatase attachment, with delaying for bovine serum albumin(BSA)
Fliud flushing 2 will contain Cea Monoclonal Antibodies --- and the solution of alkaline phosphatase attachment is diluted to 0.1 μ g/mL, must try
Agent 2;
(5) it prepares Magneto separate reagent: tromethamine powder 10g, sodium chloride powder 8g, deionized water 900g is added
In container, it is stirred well to and is completely dissolved, the Proclin300 of bovine serum albumin(BSA) 5g, cow's serum 50g, concentration 98% is molten
Liquid 0.2g is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid 1g of concentration 36%~38% is added, pH value is adjusted to
7.9, Tris buffer 3 of bovine serum albumin(BSA) are obtained after being filtered with 0.3 μm of the filter in aperture, conventional method is prepared magnetic micro-
Grain --- solution of Streptavidin attachment will contain magnetic particle with Tris buffer 3 of bovine serum albumin(BSA) ---
The solution of Streptavidin attachment is diluted to 0.5ng/mL, obtains Magneto separate reagent;
(6) chemical substrate is prepared: by tromethamine powder 25g, disodium ethylene diamine tetraacetate powder 25g, sodium chloride powder
Last 20g, lauryl sodium sulfate 0.1g, glycerine 45g, sodium sulfite 0.1g, the Proclin300 solution of concentration 98%
0.2g, deionized water 900g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid 1g of concentration 36%~38% is added, will
The pH value of solution is adjusted to 9.2, and dioxane compound is added under light protected environment after being filtered with 0.3 μm of the filter in aperture
0.6g, being configured to concentration is 0.5mg/mL, obtains chemical substrate;
(7) cleaning solution is prepared: by tromethamine powder 10g, sodium chloride powder 8g, polyoxyethylene sorbitan list
Laurate powder 5g, Triton X-100 5g, deionized water 900g are added to the container, are stirred well to completely molten
Solution is added the hydrochloric acid 1g of concentration 36%~38%, the pH value of solution is adjusted to 7.5, after being filtered with 0.3 μm of the filter in aperture
Cleaning solution.
Embodiment 4
A kind of kit of auxiliary detection sub-health population cancer base antigen, consists of the following compositions: carcinomebryonic antigen calibration
Product, cancer base antigen quality-control product, reagent 1, reagent 2, Magneto separate reagent, chemical substrate and cleaning solution.
The preparation method of the kit of above-mentioned auxiliary detection sub-health population cancer base antigen, carries out in accordance with the following steps:
(1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.3g, disodium hydrogen phosphate powder 1.5g, sodium chloride
Powder 8g, potassium chloride powder 0.3g, the Proclin300 solution 0.15g of concentration 98%, deionized water 800g are added to the container,
It is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.3, by bovine serum albumin(BSA)
32g is added to the container, and is stirred well to and is completely dissolved, and the phosphoric acid of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture
Buffer salt solution, by the phosphate buffered saline solution of cancer base antigen bovine serum albumin(BSA) be diluted to each concentration point be 0,5,25,
100,500 and 1000ng/mL;
(2) cancer base antigen quality-control product is prepared: cancer base antigen is dilute with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Releasing to each concentration point is 10ng/mL and 100ng/mL;
(3) reagent preparation 1: by tromethamine powder 8g, sodium chloride powder 4g, the Proclin300 of concentration 98%
Solution 0.15g, deionized water 800g, is added to the container, and is stirred well to and is completely dissolved, and the salt of concentration 36%~38% is added
PH value is adjusted to 7.3 and is added to the container, is stirred well to and is completely dissolved bovine serum albumin(BSA) 4g, casein 4g by sour 0.5g,
Tris buffer 1 of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture, conventional method prepares carcinomebryonic antigen list
The solution of clonal antibody --- biotin conjugate will contain carcinomebryonic antigen list with the Tris buffer 1 of bovine serum albumin(BSA)
The solution of clonal antibody --- biotin conjugate is diluted to 0.2 μ g/mL, obtains reagent 1;
(4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 5g, sodium chloride powder 0.65g, zinc chloride powder
0.075g, container is added in magnesium chloride powder 0.075g, Proclin300 the solution 0.15g, deionized water 800g of concentration 98%
In, it is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.7, ox blood is pure
Albumen 4g, casein 4g are added to the container, are stirred well to and are completely dissolved, and obtain cow's serum after being filtered with 0.3 μm of the filter in aperture
The buffer of albumin 2, conventional method prepare Cea Monoclonal Antibodies --- the solution of alkaline phosphatase attachment,
Cea Monoclonal Antibodies will be contained with buffer 2 of bovine serum albumin(BSA) --- the solution of alkaline phosphatase attachment
It is diluted to 0.06 μ g/mL, obtains reagent 2;
(5) it prepares Magneto separate reagent: tromethamine powder 8g, sodium chloride powder 6.5g, deionized water 800g is added
In container, it is stirred well to and is completely dissolved, the Proclin300 of bovine serum albumin(BSA) 4g, cow's serum 40g, concentration 98% is molten
Liquid 0.15g, casein 4g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid 0.5g of concentration 36%~38% is added,
PH value is adjusted to 8.0, Tris buffer 3 of bovine serum albumin(BSA), conventional method are obtained after being filtered with 0.3 μm of the filter in aperture
Magnetic particle --- the solution of Streptavidin attachment is prepared, magnetic will be contained with Tris buffer 3 of bovine serum albumin(BSA)
Particle --- the solution of Streptavidin attachment is diluted to 0.5ng/mL to property, obtains Magneto separate reagent;
(6) chemical substrate is prepared: by tromethamine powder 20g, disodium ethylene diamine tetraacetate powder 20g, sodium chloride powder
Last 18g, lauryl sodium sulfate 0.075g, glycerine 40g, sodium sulfite 0.075g, the Proclin300 solution of concentration 98%
0.15g, deionized water 800g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
The pH value of solution is adjusted to 9.3 by 0.5g, and dioxane is added under light protected environment after being filtered with 0.3 μm of the filter in aperture
Object 0.4g is closed, being configured to concentration is 0.5mg/mL, obtains chemical substrate;
(7) cleaning solution is prepared: by tromethamine powder 8g, sodium chloride powder 6.5g, polyoxyethylene sorbitan list
Laurate powder 4g, Triton X-100 4g, deionized water 800g are added to the container, are stirred well to completely molten
Solution is added the hydrochloric acid 0.5g of concentration 36%~38%, the pH value of solution is adjusted to 7.7, after being filtered with 0.3 μm of the filter in aperture
Obtain cleaning solution.
Embodiment 5
A kind of kit of auxiliary detection sub-health population cancer base antigen, consists of the following compositions: carcinomebryonic antigen calibration
Product, cancer base antigen quality-control product, reagent 1, reagent 2, Magneto separate reagent, chemical substrate and cleaning solution.
The preparation method of the kit of above-mentioned auxiliary detection sub-health population cancer base antigen, carries out in accordance with the following steps:
(1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.3g, disodium hydrogen phosphate powder 1.5g, sodium chloride
Powder 8g, potassium chloride powder 0.3g, the Proclin300 solution 0.15g of concentration 98%, deionized water 800g are added to the container,
It is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.3, by bovine serum albumin(BSA)
32g is added to the container, and is stirred well to and is completely dissolved, and the phosphoric acid of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture
Buffer salt solution, by the phosphate buffered saline solution of cancer base antigen bovine serum albumin(BSA) be diluted to each concentration point be 0,5,25,
100,500 and 1000ng/mL;
(2) cancer base antigen quality-control product is prepared: cancer base antigen is dilute with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Releasing to each concentration point is 10ng/mL and 100ng/mL;
(3) reagent preparation 1: by tromethamine powder 8g, sodium chloride powder 4g, the Proclin300 of concentration 98%
Solution 0.15g, deionized water 800g, is added to the container, and is stirred well to and is completely dissolved, and the salt of concentration 36%~38% is added
PH value is adjusted to 7.3 and bovine serum albumin(BSA) 4g, N- hydroxysuccinimide powder 4g is added to the container, is sufficiently stirred by sour 0.5g
It mixes to being completely dissolved, Tris buffer 1 of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture, conventional method is matched
The solution of Cea Monoclonal Antibodies processed --- biotin conjugate will be contained with Tris buffer 1 of bovine serum albumin(BSA)
There are Cea Monoclonal Antibodies --- the solution of biotin conjugate is diluted to 0.2 μ g/mL, obtains reagent 1;
(4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 5g, sodium chloride powder 0.65g, zinc chloride powder
0.075g, container is added in magnesium chloride powder 0.075g, Proclin300 the solution 0.15g, deionized water 800g of concentration 98%
In, it is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.7, ox blood is pure
Albumen 4g, N- hydroxysuccinimide powder 4g is added to the container, and is stirred well to and is completely dissolved, with 0.3 μm of the filter in aperture
Buffer 2 of bovine serum albumin(BSA) are obtained after filtering, conventional method prepares Cea Monoclonal Antibodies --- alkaline phosphatase
The solution of attachment will contain Cea Monoclonal Antibodies with buffer 2 of bovine serum albumin(BSA) --- alkaline phosphatase
The solution of attachment is diluted to 0.06 μ g/mL, obtains reagent 2;
(5) it prepares Magneto separate reagent: tromethamine powder 8g, sodium chloride powder 6.5g, deionized water 800g is added
In container, it is stirred well to and is completely dissolved, the Proclin300 of bovine serum albumin(BSA) 4g, cow's serum 40g, concentration 98% is molten
Liquid 0.15g, N- hydroxysuccinimide powder 4g is added to the container, and is stirred well to and is completely dissolved, and addition concentration 36%~
38% hydrochloric acid 0.5g, is adjusted to 8.0 for pH value, and the Tris that bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture is buffered
Liquid 3, conventional method prepares magnetic particle --- the solution of Streptavidin attachment, slow with the Tris of bovine serum albumin(BSA)
Fliud flushing 3 will contain magnetic particle --- and the solution of Streptavidin attachment is diluted to 0.5ng/mL, obtains Magneto separate reagent;
(6) chemical substrate is prepared: by tromethamine powder 20g, disodium ethylene diamine tetraacetate powder 20g, sodium chloride powder
Last 18g, lauryl sodium sulfate 0.075g, glycerine 40g, sodium sulfite 0.075g, the Proclin300 solution of concentration 98%
0.15g, deionized water 800g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
The pH value of solution is adjusted to 9.3 by 0.5g, and dioxane is added under light protected environment after being filtered with 0.3 μm of the filter in aperture
Object 0.4g is closed, being configured to concentration is 0.5mg/mL, obtains chemical substrate;
(7) cleaning solution is prepared: by tromethamine powder 8g, sodium chloride powder 6.5g, polyoxyethylene sorbitan list
Laurate powder 4g, Triton X-100 4g, deionized water 800g are added to the container, are stirred well to completely molten
Solution is added the hydrochloric acid 0.5g of concentration 36%~38%, the pH value of solution is adjusted to 7.7, after being filtered with 0.3 μm of the filter in aperture
Obtain cleaning solution.
Embodiment 6
A kind of kit of auxiliary detection sub-health population cancer base antigen, consists of the following compositions: carcinomebryonic antigen calibration
Product, cancer base antigen quality-control product, reagent 1, reagent 2, Magneto separate reagent, chemical substrate and cleaning solution.
The preparation method of the kit of above-mentioned auxiliary detection sub-health population cancer base antigen, carries out in accordance with the following steps:
(1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.3g, disodium hydrogen phosphate powder 1.5g, sodium chloride
Powder 8g, potassium chloride powder 0.3g, the Proclin300 solution 0.15g of concentration 98%, deionized water 800g are added to the container,
It is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.3, by bovine serum albumin(BSA)
32g is added to the container, and is stirred well to and is completely dissolved, and the phosphoric acid of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture
Buffer salt solution, by the phosphate buffered saline solution of cancer base antigen bovine serum albumin(BSA) be diluted to each concentration point be 0,5,25,
100,500 and 1000ng/mL;
(2) cancer base antigen quality-control product is prepared: cancer base antigen is dilute with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Releasing to each concentration point is 10ng/mL and 100ng/mL;
(3) reagent preparation 1: by tromethamine powder 8g, sodium chloride powder 4g, the Proclin300 of concentration 98%
Solution 0.15g, deionized water 800g, is added to the container, and is stirred well to and is completely dissolved, and the salt of concentration 36%~38% is added
PH value is adjusted to 7.3 for bovine serum albumin(BSA) 4g, (Na by sour 0.5g2.3K0.7) (B6O10)(N O3) powder 4g is added to the container,
It is stirred well to and is completely dissolved, Tris buffer 1 of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture, it is conventional
Method prepares Cea Monoclonal Antibodies --- the solution of biotin conjugate, with the Tris buffer 1 of bovine serum albumin(BSA)
Number containing Cea Monoclonal Antibodies --- the solution of biotin conjugate is diluted to 0.2 μ g/mL, obtains reagent 1;
(4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 5g, sodium chloride powder 0.65g, zinc chloride powder
0.075g, container is added in magnesium chloride powder 0.075g, Proclin300 the solution 0.15g, deionized water 800g of concentration 98%
In, it is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.7, ox blood is pure
Albumen 4g, (Na2.3K0.7)(B6O10)(N O3) powder 4g is added to the container, is stirred well to and is completely dissolved, with 0.3 μm of aperture
Filter filtering after buffer 2 of bovine serum albumin(BSA), conventional method prepares Cea Monoclonal Antibodies --- alkalinity
The solution of phosphatase attachment will contain Cea Monoclonal Antibodies with buffer 2 of bovine serum albumin(BSA) --- alkalinity
The solution of phosphatase attachment is diluted to 0.06 μ g/mL, obtains reagent 2;
(5) it prepares Magneto separate reagent: tromethamine powder 8g, sodium chloride powder 6.5g, deionized water 800g is added
In container, it is stirred well to and is completely dissolved, the Proclin300 of bovine serum albumin(BSA) 4g, cow's serum 40g, concentration 98% is molten
Liquid 0.15g, (Na2.3K0.7)(B6O10)(N O3) powder 4g is added to the container, is stirred well to and is completely dissolved, concentration is added
36%~38% hydrochloric acid 0.5g, is adjusted to 8.0 for pH value, obtains bovine serum albumin(BSA) after being filtered with 0.3 μm of the filter in aperture
Tris buffer 3, magnetic particle --- the solution of Streptavidin attachment uses bovine serum albumin(BSA) for conventional method preparation
Tris buffer 3 will contain magnetic particle --- the solution of Streptavidin attachment is diluted to 0.5ng/mL, obtains magnetic point
From reagent;
(6) chemical substrate is prepared: by tromethamine powder 20g, disodium ethylene diamine tetraacetate powder 20g, sodium chloride powder
Last 18g, lauryl sodium sulfate 0.075g, glycerine 40g, sodium sulfite 0.075g, the Proclin300 solution of concentration 98%
0.15g, deionized water 800g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
The pH value of solution is adjusted to 9.3 by 0.5g, and dioxane is added under light protected environment after being filtered with 0.3 μm of the filter in aperture
Object 0.4g is closed, being configured to concentration is 0.5mg/mL, obtains chemical substrate;
(7) cleaning solution is prepared: by tromethamine powder 8g, sodium chloride powder 6.5g, polyoxyethylene sorbitan list
Laurate powder 4g, Triton X-100 4g, deionized water 800g are added to the container, are stirred well to completely molten
Solution is added the hydrochloric acid 0.5g of concentration 36%~38%, the pH value of solution is adjusted to 7.7, after being filtered with 0.3 μm of the filter in aperture
Obtain cleaning solution.
Embodiment 7
A kind of kit of auxiliary detection sub-health population cancer base antigen, consists of the following compositions: carcinomebryonic antigen calibration
Product, cancer base antigen quality-control product, reagent 1, reagent 2, Magneto separate reagent, chemical substrate and cleaning solution.
The preparation method of the kit of above-mentioned auxiliary detection sub-health population cancer base antigen, carries out in accordance with the following steps:
(1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.3g, disodium hydrogen phosphate powder 1.5g, sodium chloride
Powder 8g, potassium chloride powder 0.3g, the Proclin300 solution 0.15g of concentration 98%, deionized water 800g are added to the container,
It is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.3, by bovine serum albumin(BSA)
32g is added to the container, and is stirred well to and is completely dissolved, and the phosphoric acid of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture
Buffer salt solution, by the phosphate buffered saline solution of cancer base antigen bovine serum albumin(BSA) be diluted to each concentration point be 0,5,25,
100,500 and 1000ng/mL;
(2) cancer base antigen quality-control product is prepared: cancer base antigen is dilute with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Releasing to each concentration point is 10ng/mL and 100ng/mL;
(3) reagent preparation 1: by tromethamine powder 8g, the Proclin300 of sodium chloride powder 4g, concentration 98% are molten
Liquid 0.15g, deionized water 800g, is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
PH value is adjusted to 7.3 and is added to the container bovine serum albumin(BSA) 4g, Eudesmin powder 4g by 0.5g, is stirred well to completely molten
Solution obtains Tris buffer 1 of bovine serum albumin(BSA) after being filtered with 0.3 μm of the filter in aperture, conventional method prepares carcinomebryonic antigen
The solution of monoclonal antibody --- biotin conjugate will contain carcinomebryonic antigen with Tris buffer 1 of bovine serum albumin(BSA)
The solution of monoclonal antibody --- biotin conjugate is diluted to 0.2 μ g/mL, obtains reagent 1;
(4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 5g, sodium chloride powder 0.65g, zinc chloride powder
Container is added in 0.075g, magnesium chloride powder 0.075g, Proclin300 the solution 0.15g, deionized water 800g of concentration 98%
In, it is stirred well to and is completely dissolved, the hydrochloric acid 0.5g of concentration 36%~38% is added, pH value is adjusted to 7.7, ox blood is pure
Albumen 4g, Eudesmin powder 4g are added to the container, are stirred well to and are completely dissolved, and obtain ox after being filtered with 0.3 μm of the filter in aperture
Sero-abluminous buffer 2, conventional method prepare Cea Monoclonal Antibodies --- alkaline phosphatase attachment it is molten
Liquid will contain Cea Monoclonal Antibodies with buffer 2 of bovine serum albumin(BSA) --- alkaline phosphatase attachment it is molten
Liquid is diluted to 0.06 μ g/mL, obtains reagent 2;
(5) it prepares Magneto separate reagent: tromethamine powder 8g, sodium chloride powder 6.5g, deionized water 800g is added
In container, it is stirred well to and is completely dissolved, the Proclin300 of bovine serum albumin(BSA) 4g, cow's serum 40g, concentration 98% is molten
Liquid 0.15g, Eudesmin powder 4g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
PH value is adjusted to 8.0 by 0.5g, and Tris buffer 3 of bovine serum albumin(BSA) are obtained after being filtered with 0.3 μm of the filter in aperture, conventional
Method prepares magnetic particle --- the solution of Streptavidin attachment, will be contained with Tris buffer 3 of bovine serum albumin(BSA)
Be magnetic particle --- and the solution of Streptavidin attachment is diluted to 0.5ng/mL, obtains Magneto separate reagent;
(6) chemical substrate is prepared: by tromethamine powder 20g, disodium ethylene diamine tetraacetate powder 20g, sodium chloride powder
Last 18g, lauryl sodium sulfate 0.075g, glycerine 40g, sodium sulfite 0.075g, the Proclin300 solution of concentration 98%
0.15g, deionized water 800g are added to the container, are stirred well to and are completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
The pH value of solution is adjusted to 9.3 by 0.5g, and dioxane is added under light protected environment after being filtered with 0.3 μm of the filter in aperture
Object 0.4g is closed, being configured to concentration is 0.5mg/mL, obtains chemical substrate;
(7) cleaning solution is prepared: by tromethamine powder 8g, sodium chloride powder 6.5g, polyoxyethylene sorbitan list
Laurate powder 4g, Triton X-100 4g, deionized water 800g are added to the container, are stirred well to completely molten
Solution is added the hydrochloric acid 0.5g of concentration 36%~38%, the pH value of solution is adjusted to 7.7, after being filtered with 0.3 μm of the filter in aperture
Obtain cleaning solution.
Experimental example 1:
1 batch is respectively taken according to the kit prepared in embodiment 1-7, measures the serum of basic, normal, high concentration respectively, 50 holes are flat
Row measurement, obtains variation within batch coefficient, separately respectively takes 3 batches according to the kit prepared in embodiment 1-7, every a batch kit difference
The serum of basic, normal, high concentration is measured, 50 holes are measured in parallel, interassay coefficient of variation is obtained, as a result referring to table 1:
The comparison of the variation within batch coefficient and interassay coefficient of variation of each experimental example of table 1
Note: * represents P < 0.05 compared with example 1 group
By table 1, it can be concluded that, the variation within batch coefficient of the measurement serum-concentration 3.5ng/mL of embodiment 1-3 is suitable, does not have
The variation within batch coefficient of significant difference, the measurement serum-concentration 3.5ng/mL of embodiment 4-7 is substantially less than embodiment 1.Embodiment
The variation within batch coefficient of 1-3, the measurement serum-concentration 30.6ng/mL of embodiment 5 are suitable, are not significantly different, embodiment 4, real
The variation within batch coefficient for applying the measurement serum-concentration 30.6ng/mL of a 6-7 is substantially less than embodiment 1.The measurement of embodiment 1-3
The variation within batch coefficient of serum-concentration 50.3g/mL is suitable, is not significantly different, the measurement serum-concentration of embodiment 4-7
The variation within batch coefficient of 50.3ng/mL is substantially less than embodiment 1.The measurement serum-concentration 3.5ng/mL of embodiment 1-3 and
The interassay coefficient of variation of 50.3ng/mL is suitable, is not significantly different, the measurement serum-concentration 3.5ng/mL's of embodiment 4-7
Interassay coefficient of variation is substantially less than embodiment 1.The variation within batch coefficient phase of the measurement serum-concentration 30.6ng/mL of embodiment 1-4
When being not significantly different, the variation within batch coefficient of the measurement serum-concentration 30.6ng/mL of embodiment 5-7 is substantially less than embodiment
1。
Experimental example 2:
Respectively take 1 batch according to the kit prepared in embodiment 1-7, use the zero-dose calibration object of calibration object STD-A point as
Sample is detected, and replication 50 times, obtains the relative luminous intensity RLU value of 50 measurement result calibration object STD-A points,
Its average value (M) and standard deviation (SD) are calculated, obtains relative luminous intensity RLU corresponding to the M+2SD of calibration object STD-A point
Value, uses the calibration object of calibration object STD-B point to be detected as sample, replication 50 times, obtains 50 measurement result calibrations
The relative luminous intensity RLU value of product STD-B point simultaneously calculates average value M ', according to the zero-dose calibration object of STD-A point and calibration
The mean value result of concentration and relative luminous intensity RLU value between product STD-B point carries out two o'clock regression fit and obtains first power
RLU value corresponding to the M+2SD of calibration object STD-A point is brought into above-mentioned equation, finds out corresponding concentration value by journey, as empty
White limit, as a result referring to table 2:
The comparison of the blank limit of each experimental example of table 2
Note: * represents P < 0.05 compared with example 1 group
By table 2, it can be concluded that, the blank limit of embodiment 1-3 quite, is not significantly different, the blank limit of embodiment 4-7
Substantially less than embodiment 1 illustrates that the sensitivity of embodiment 4-7 is higher.
Experimental example 3:
Accuracy refers to that measured value obtains degree close to true value, is usually indicated with the rate of recovery, according to preparing in embodiment 1-7
Kit respectively take 1 batch, separately take clinical definite value serum, concentration 8.38ng/mL adds certain density reality thereto respectively
A standard items for 1-7 kit are applied, and measure the concentration value of sample after 100 mixing, calculate measurement average value and the rate of recovery,
As a result referring to table 3:
The rate of recovery of each experimental example of table 3
Note: * represents P < 0.05 compared with example 1 group
By table 3, it can be concluded that, the rate of recovery of embodiment 1-3 is suitable, is not significantly different, and the rate of recovery of embodiment 4-7 is aobvious
It writes and is higher than embodiment 1, illustrate that the accuracy of embodiment 4-7 is higher.
Experimental example 4:
1 batch is respectively taken according to the kit prepared in embodiment 1-7, respectively taking a g carcinomebryonic antigen is 0 sample, and first tire is added
Albumin A FP antigen concentration is 100ng/mL, is detected using the kit prepared in embodiment 1-7, and cancer embryo in sample is detected
Content, measurement 100 times and calculates relative light unit RLU and concentration, as a result referring to table 4:
The Evaluation on specificity of each experimental example of table 4
Note: * represents P < 0.05 compared with example 1 group
By table 4, it can be concluded that, the measurement result mean value of embodiment 1-3 is suitable, is not significantly different, the survey of embodiment 4-7
Determine result mean value and is substantially less than embodiment 1.
Experimental example 5:
1 batch is respectively taken according to the kit prepared in embodiment 1-7, it will be close to the height of the range of linearity upper limit (1000ng/mL)
Value sample is diluted at least six kinds of concentration according to a certain percentage, and wherein the sample of low value concentration must be close under the range of linearity
Limit repeats to detect 3 times, calculates its average value to the sample standard deviation of each concentration, by result average value and dilution ratio minimum two
Multiplication carries out straight line fitting, and calculates linearly dependent coefficient r, as a result referring to table 5:
It is evaluated after the dilution of each experimental example of table 5
It can be concluded that, after the dilution of embodiment 1-7 quite, it is not significantly different by table 5, illustrates that the range of linearity is good.
Experimental example 6:
1 batch is respectively taken according to the kit prepared in embodiment 1-7, carries out 4 DEG C respectively to kit 12 months and 37 DEG C 7
It accelerated stability experiment, the results showed that the variation of kit standard product luminous intensity criticizes interior and betweenrun precision, is accurate
The indicator deviations such as degree are little, and within normal range (NR), kit validity period was up to 12 months.
Claims (10)
1. a kind of kit of auxiliary detection sub-health population cancer base antigen, which is characterized in that the kit includes:
1) carcinomebryonic antigen calibration object, the phosphate buffered saline solution containing carcinomebryonic antigen and bovine serum albumin(BSA), the bovine serum albumin
White phosphate buffered saline solution includes following component: bovine serum albumin(BSA) 30-34g, potassium dihydrogen phosphate powder 0.2-0.4g, phosphoric acid
Disodium hydrogen powder 1-2g, sodium chloride powder 7-9g, potassium chloride powder 0.2-0.4g, the hydrochloric acid 0.1-1mL of concentration 36%~38%,
Proclin300 the solution 0.1-0.2mL, deionized water 700-900mL of concentration 98%;The carcinomebryonic antigen calibration object is 6 water
Flat liquid calibration object, the concentration of carcinomebryonic antigen is respectively 0,5,25,100,500 and in 6 horizontal calibration objects
1000ng/mL, pH 7.2-7.4;
2) cancer base antigen quality-control product, the phosphate buffered saline solution containing carcinomebryonic antigen and bovine serum albumin(BSA), the bovine serum albumin
White phosphate buffered saline solution is the phosphate buffered saline solution of the bovine serum albumin(BSA) of the carcinomebryonic antigen calibration object, the cancer embryo
Antigen quality-control product is 2 horizontal liquid calibration objects, and the target value concentration of carcinomebryonic antigen is in 2 horizontal liquid quality controls
10ng/mL and 100ng/mL, pH 7.2-7.4;
3) reagent 1, the Tris buffer of Cea Monoclonal Antibodies and bovine serum albumin(BSA) containing biotin labeling 1,
Tris buffer 1 of the bovine serum albumin(BSA) includes following component, bovine serum albumin(BSA) 3-5g, tromethamine powder 6-
10g, sodium chloride powder 3-5g, the hydrochloric acid 0.1-1mL of concentration 36%~38%, the Proclin300 solution 0.1- of concentration 98%
0.2mL, deionized water 700-900mL;
4) reagent 2, the buffer 2 of Cea Monoclonal Antibodies and bovine serum albumin(BSA) containing alkali phosphatase enzyme mark
Number, buffer 2 of the bovine serum albumin(BSA) include following component, 4- hydroxyethyl piperazineethanesulfonic acid powder 4-6g, sodium chloride
Powder 5-8g, bovine serum albumin(BSA) 3-5g, zinc chloride powder 0.05-0.1g, magnesium chloride powder 0.05-0.1g, concentration 36%~
38% hydrochloric acid 0.1-1mL, Proclin300 the solution 0.1-0.2mL, deionized water 700-900mL of concentration 98%;
5) Magneto separate reagent, the Tris buffer of magnetic particle and bovine serum albumin(BSA) containing marked by streptavidin 3, institute
Tris buffer 3 for stating bovine serum albumin(BSA) include following component, bovine serum albumin(BSA) 3-5g, cow's serum 30-50g, amino
Butantriol powder 6-10g, sodium chloride powder 5-8g, the hydrochloric acid 0.1-1mL of concentration 36%~38%, concentration 98%
Proclin300 solution 0.1-0.2mL, deionized water 700-900mL;
6) chemical substrate, the chemical substrate include following component, tromethamine powder 15-25g, disodium ethylene diamine tetraacetate
Powder 15-25g, sodium chloride powder 15-20g, dioxane compound 0.2-0.6g, lauryl sodium sulfate 0.05-
0.1g, glycerine 35-45g, sodium sulfite 0.05-0.1g, the Proclin300 solution 0.1-0.2mL of concentration 98%, deionization
Water 700-900mL;
7) cleaning solution, the cleaning solution include following component, tromethamine powder 6-10g, sodium chloride powder 5-8g, polyoxy second
Alkene sorbitan mono-laurate powder 3-5g, Triton X-100 3-5g, the hydrochloric acid of concentration 36%~38%
0.1-1mL, deionized water 700-900mL, the pH value of the cleaning solution are 7.5-8.0.
2. the kit of auxiliary detection sub-health population cancer base antigen according to claim 1, which is characterized in that the cancer
Embryonal antigen calibration object, the carcinomebryonic antigen quality-control product, the reagent 1, the reagent 2, the Magneto separate reagent, describedization
The storage temperature for learning substrate is 2-8 DEG C, and the storage temperature of the cleaning solution is 10-30 DEG C of room temperature, and the kit avoids sunlight
Direct projection.
3. the kit of auxiliary detection sub-health population cancer base antigen according to claim 1, which is characterized in that the examination
The concentration of the Cea Monoclonal Antibodies of biotin labeling is 0.1-0.3 μ g/mL, pH value 7.0-7.5 in agent 1, described
The concentration of the Cea Monoclonal Antibodies of No. 2 alkaline phosphatases of reagent label is 0.02-0.1 μ g/mL, pH value 7.5-
8.0, the concentration of the Magneto separate reagent is 0.5ng/mL, and pH value 7.9-8.1, the diameter of the magnetic particle is 0.5-3.5 μ
M, kernel are ferroso-ferric oxide, and surface is wrapped in the polymer containing carboxyl, dioxane chemical combination in the chemical substrate
The concentration of object is 0.5mg/mL, pH value 9.2-9.5.
4. the kit of auxiliary detection sub-health population cancer base antigen according to claim 1, which is characterized in that the ox
Sero-abluminous Tris buffer 1, the Tris of the buffer of the bovine serum albumin(BSA) 2 and the bovine serum albumin(BSA)
Casein 3-5g can also be added in buffer 3.
5. the kit of auxiliary detection sub-health population cancer base antigen according to claim 1, which is characterized in that the ox
Sero-abluminous Tris buffer 1, the buffer of the bovine serum albumin(BSA) 2, the Tris of the bovine serum albumin(BSA)
N- hydroxysuccinimide powder 3-5g can also be added in buffer 3.
6. the kit of auxiliary detection sub-health population cancer base antigen according to claim 1, which is characterized in that the ox
Sero-abluminous Tris buffer 1, the buffer of the bovine serum albumin(BSA) 2, the Tris of the bovine serum albumin(BSA)
(Na can also be added in buffer 32.3K0.7)(B6O10)(NO3) powder 3-5g.
7. the preparation method of the kit of auxiliary detection sub-health population cancer base antigen according to claim 1, feature
It is, carries out in accordance with the following steps:
1) carcinomebryonic antigen calibration object is prepared: by potassium dihydrogen phosphate powder 0.2-0.4g, disodium hydrogen phosphate powder 1-2g, sodium chloride powder
Last 7-9g, potassium chloride powder 0.2-0.4g, Proclin300 the solution 0.1-0.2mL, deionized water 700-900mL of concentration 98%
It is added to the container, is stirred well to and is completely dissolved, the hydrochloric acid 0.1-1mL of concentration 36%~38% is added, pH value is adjusted to 7.2-
7.4, bovine serum albumin(BSA) 30-34g is added to the container, is stirred well to and is completely dissolved, after being filtered with 0.3 μm of the filter in aperture
The phosphate buffered saline solution for obtaining bovine serum albumin(BSA), the phosphate buffered saline solution of cancer base antigen bovine serum albumin(BSA) is diluted to
Each concentration point is 0,5,25,100,500 and 1000ng/mL;
2) it prepares cancer base antigen quality-control product: cancer base antigen is diluted to respectively with the phosphate buffered saline solution of above-mentioned bovine serum albumin(BSA)
Concentration point is 10ng/mL and 100ng/mL;
3) reagent preparation 1: by tromethamine powder 6-10g, the Proclin300 of sodium chloride powder 3-5g, concentration 98% are molten
Liquid 0.1-0.2mL, deionized water 700-900mL, is added to the container, and is stirred well to and is completely dissolved, and addition concentration 36%~
38% hydrochloric acid 0.1-1mL, is adjusted to 7.0-7.5 for pH value, and bovine serum albumin(BSA) 3-5g is added to the container, has been stirred well to
Fully dissolved obtains Tris buffer 1 of bovine serum albumin(BSA) after being filtered with 0.3 μm of the filter in aperture, conventional method prepares cancer embryo
The solution of antigen monoclonal antibody --- biotin conjugate will contain cancer embryo with the Tris buffer 1 of bovine serum albumin(BSA)
The solution of antigen monoclonal antibody --- biotin conjugate is diluted to 0.1-0.3 μ g/mL, obtains reagent 1;
4) reagent preparation 2: by 4- hydroxyethyl piperazineethanesulfonic acid powder 4-6g, sodium chloride powder 5-8g, zinc chloride powder 0.05-
0.1g, magnesium chloride powder 0.05-0.1g, Proclin300 the solution 0.1-0.2mL, deionized water 700-900mL of concentration 98%
It is added to the container, is stirred well to and is completely dissolved, the hydrochloric acid 0.1-1mL of concentration 36%~38% is added, pH value is adjusted to 7.5-
8.0, bovine serum albumin(BSA) 3-5g is added to the container, is stirred well to and is completely dissolved, after being filtered with 0.3 μm of the filter in aperture
The buffer of bovine serum albumin(BSA) 2, conventional method prepare Cea Monoclonal Antibodies --- alkaline phosphatase attachment
Solution, will be molten containing Cea Monoclonal Antibodies-alkaline phosphatase attachment with the buffer 2 of bovine serum albumin(BSA)
Liquid is diluted to 0.02-0.1 μ g/mL, obtains reagent 2;
5) it prepares Magneto separate reagent: tromethamine powder 6-10g, sodium chloride powder 5-8g, deionized water 700-900mL is added
Enter in container, is stirred well to and is completely dissolved, by bovine serum albumin(BSA) 3-5g, cow's serum 30-50g, concentration 98%
Proclin300 solution 0.1-0.2mL is added to the container, and is stirred well to and is completely dissolved, and the hydrochloric acid of concentration 36%~38% is added
PH value is adjusted to 7.9-8.1 by 0.1-1mL, and the Tris buffer 3 of bovine serum albumin(BSA) is obtained after being filtered with 0.3 μm of the filter in aperture
Number, conventional method prepares magnetic particle --- the solution of Streptavidin attachment, with the Tris buffer 3 of bovine serum albumin(BSA)
Number containing magnetic particle --- the solution of Streptavidin attachment is diluted to 0.5ng/mL, obtains Magneto separate reagent;
6) chemical substrate is prepared: by tromethamine powder 15-25g, disodium ethylene diamine tetraacetate powder 15-25g, sodium chloride powder
Last 15-20g, lauryl sodium sulfate 0.05-0.1g, glycerine 35-45g, sodium sulfite 0.05-0.1g, concentration 98%
Proclin300 solution 0.1-0.2mL, deionized water 700-900mL are added to the container, are stirred well to and are completely dissolved, and are added dense
The hydrochloric acid 0.1-1mL of degree 36%~38%, is adjusted to 9.2-9.5 for the pH value of solution, is protected from light after being filtered with 0.3 μm of the filter in aperture
Dioxane compound 0.2-0.6g is added under environment, being configured to concentration is 0.5mg/mL, obtains chemical substrate;
7) cleaning solution is prepared: by tromethamine powder 6-10g, sodium chloride powder 5-8g, polyoxyethylene sorbitan Dan Yue
Cinnamic acid ester powder 3-5g, Triton X-100 3-5g, deionized water 700-900mL are added to the container, are stirred well to
It is completely dissolved, the hydrochloric acid 0.1-1mL of concentration 36%~38% is added, the pH value of solution is adjusted to 7.5-8.0, with 0.3 μm of aperture
Filter filtering after cleaning solution.
8. the preparation method of the kit of auxiliary detection sub-health population cancer base antigen according to claim 7, feature
It is, the step (3), the step (4), casein 3-5g or N- maloyl can be also added in the step (5)
Imines powder 3-5g.
9. the preparation method of the kit of auxiliary detection sub-health population cancer base antigen according to claim 7, feature
It is, the step (3), the step (4), (Na can be also added in the step (5)2.3K0.7)(B6O10)(NO3) powder 3-
5g。
10. the detection method of the kit of auxiliary detection sub-health population cancer base antigen according to claim 1, feature
It is, this method comprises the following steps:
1) plus 10 μ L cancer base antigen calibration objects, quality-control product or sample to be tested are into detection pipe;
2) the cancer base antigenic agents 1 of 50 μ L are added into detection pipe described in step 1), after mixing, 37 ± 0.5 DEG C incubate 10 points
Clock;
3) 50 μ L Magneto separate reagents are added into detection pipe described in step 2), after mixing, 37 ± 0.5 DEG C are incubated 5 minutes, carry out magnetic
Separate supernatant;
4) cleaning solution of 500 μ L is added into detection pipe described in step 3), after mixing, carries out Magneto separate and removes supernatant;
5) it repeats twice of step 4);
6) 50 μ L reagents 2 are added into detection pipe described in step 5), after mixing, 37 ± 0.5 DEG C are incubated 7 minutes, carry out magnetic point
It leaves away supernatant;
7) cleaning solution of 500 μ L is added into detection pipe described in step 6), after mixing, carries out Magneto separate and removes supernatant;
8) it repeats twice of step 4);
9) 200 μ L chemical substrates are added into detection pipe described in step 8), luminous intensity is detected after mixing.
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Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
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CN110426520A (en) * | 2019-06-18 | 2019-11-08 | 上海彧成生物科技有限公司 | A kind of magnetic particle storing liquid and its preparation |
CN113358868A (en) * | 2021-05-24 | 2021-09-07 | 迪瑞医疗科技股份有限公司 | Sperm acrosome enzyme chemiluminescence immunoassay kit and preparation method thereof |
-
2018
- 2018-09-13 CN CN201811066799.9A patent/CN109239332A/en not_active Withdrawn
Cited By (2)
Publication number | Priority date | Publication date | Assignee | Title |
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CN110426520A (en) * | 2019-06-18 | 2019-11-08 | 上海彧成生物科技有限公司 | A kind of magnetic particle storing liquid and its preparation |
CN113358868A (en) * | 2021-05-24 | 2021-09-07 | 迪瑞医疗科技股份有限公司 | Sperm acrosome enzyme chemiluminescence immunoassay kit and preparation method thereof |
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