CN104777303A - Basophilic granulocyte activation and degranulation identification method - Google Patents
Basophilic granulocyte activation and degranulation identification method Download PDFInfo
- Publication number
- CN104777303A CN104777303A CN201510212435.7A CN201510212435A CN104777303A CN 104777303 A CN104777303 A CN 104777303A CN 201510212435 A CN201510212435 A CN 201510212435A CN 104777303 A CN104777303 A CN 104777303A
- Authority
- CN
- China
- Prior art keywords
- human
- basophilic granulocyte
- mark
- streaming
- ccr3
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
- 210000003714 granulocyte Anatomy 0.000 title claims abstract description 75
- 238000000034 method Methods 0.000 title claims abstract description 32
- 230000004913 activation Effects 0.000 title claims abstract description 25
- 101000998120 Homo sapiens Interleukin-3 receptor subunit alpha Proteins 0.000 claims abstract description 59
- 102100033493 Interleukin-3 receptor subunit alpha Human genes 0.000 claims abstract description 59
- 102100024167 C-C chemokine receptor type 3 Human genes 0.000 claims abstract description 53
- 101710149862 C-C chemokine receptor type 3 Proteins 0.000 claims abstract description 53
- 102000006354 HLA-DR Antigens Human genes 0.000 claims abstract description 43
- 108010058597 HLA-DR Antigens Proteins 0.000 claims abstract description 43
- 210000004369 blood Anatomy 0.000 claims abstract description 39
- 238000012360 testing method Methods 0.000 claims abstract description 36
- 102100025222 CD63 antigen Human genes 0.000 claims abstract description 34
- 101000934368 Homo sapiens CD63 antigen Proteins 0.000 claims abstract description 34
- 239000008280 blood Substances 0.000 claims abstract description 29
- 210000004027 cell Anatomy 0.000 claims abstract description 21
- 210000003743 erythrocyte Anatomy 0.000 claims abstract description 21
- 239000007850 fluorescent dye Substances 0.000 claims abstract description 20
- 238000001215 fluorescent labelling Methods 0.000 claims abstract description 20
- 239000006228 supernatant Substances 0.000 claims abstract description 15
- 238000000684 flow cytometry Methods 0.000 claims abstract 4
- 239000007788 liquid Substances 0.000 claims description 34
- 238000011068 loading method Methods 0.000 claims description 21
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 claims description 16
- 239000012530 fluid Substances 0.000 claims description 15
- 238000013016 damping Methods 0.000 claims description 14
- 230000012447 hatching Effects 0.000 claims description 14
- 238000002156 mixing Methods 0.000 claims description 14
- 238000002372 labelling Methods 0.000 claims description 9
- 238000005119 centrifugation Methods 0.000 claims description 7
- 239000008363 phosphate buffer Substances 0.000 claims description 7
- 229910021642 ultra pure water Inorganic materials 0.000 claims description 7
- 239000012498 ultrapure water Substances 0.000 claims description 7
- 239000003643 water by type Substances 0.000 claims description 7
- 238000001556 precipitation Methods 0.000 claims description 5
- 238000005070 sampling Methods 0.000 claims description 5
- 101000897042 Homo sapiens Nucleotide pyrophosphatase Proteins 0.000 abstract description 7
- 102100021969 Nucleotide pyrophosphatase Human genes 0.000 abstract description 7
- 238000001514 detection method Methods 0.000 abstract description 6
- 210000005259 peripheral blood Anatomy 0.000 abstract description 2
- 239000011886 peripheral blood Substances 0.000 abstract description 2
- 238000011534 incubation Methods 0.000 abstract 3
- 239000013592 cell lysate Substances 0.000 abstract 1
- 239000013566 allergen Substances 0.000 description 7
- 206010020751 Hypersensitivity Diseases 0.000 description 6
- 238000003745 diagnosis Methods 0.000 description 6
- 201000010099 disease Diseases 0.000 description 6
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 6
- 210000003630 histaminocyte Anatomy 0.000 description 5
- NTYJJOPFIAHURM-UHFFFAOYSA-N Histamine Chemical compound NCCC1=CN=CN1 NTYJJOPFIAHURM-UHFFFAOYSA-N 0.000 description 4
- 208000030961 allergic reaction Diseases 0.000 description 4
- 208000035474 group of disease Diseases 0.000 description 4
- 230000001404 mediated effect Effects 0.000 description 4
- 208000003455 anaphylaxis Diseases 0.000 description 3
- 208000006673 asthma Diseases 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 238000010998 test method Methods 0.000 description 3
- 206010002216 Anaphylactoid reaction Diseases 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- 208000026935 allergic disease Diseases 0.000 description 2
- 230000007815 allergy Effects 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 230000018109 developmental process Effects 0.000 description 2
- 230000036541 health Effects 0.000 description 2
- 229960001340 histamine Drugs 0.000 description 2
- 210000003712 lysosome Anatomy 0.000 description 2
- 230000001868 lysosomic effect Effects 0.000 description 2
- 238000012797 qualification Methods 0.000 description 2
- 206010061623 Adverse drug reaction Diseases 0.000 description 1
- 206010002198 Anaphylactic reaction Diseases 0.000 description 1
- 206010012735 Diarrhoea Diseases 0.000 description 1
- 206010013700 Drug hypersensitivity Diseases 0.000 description 1
- 208000030453 Drug-Related Side Effects and Adverse reaction Diseases 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 208000001718 Immediate Hypersensitivity Diseases 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 208000031662 Noncommunicable disease Diseases 0.000 description 1
- 206010039085 Rhinitis allergic Diseases 0.000 description 1
- 206010045240 Type I hypersensitivity Diseases 0.000 description 1
- 230000002009 allergenic effect Effects 0.000 description 1
- 201000009961 allergic asthma Diseases 0.000 description 1
- 230000000172 allergic effect Effects 0.000 description 1
- 201000010105 allergic rhinitis Diseases 0.000 description 1
- 230000036783 anaphylactic response Effects 0.000 description 1
- 208000010216 atopic IgE responsiveness Diseases 0.000 description 1
- 208000010668 atopic eczema Diseases 0.000 description 1
- 210000003651 basophil Anatomy 0.000 description 1
- 210000001772 blood platelet Anatomy 0.000 description 1
- 210000000170 cell membrane Anatomy 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000003759 clinical diagnosis Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 201000005311 drug allergy Diseases 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000002757 inflammatory effect Effects 0.000 description 1
- 230000004054 inflammatory process Effects 0.000 description 1
- 201000010659 intrinsic asthma Diseases 0.000 description 1
- 210000002540 macrophage Anatomy 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 239000012528 membrane Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 210000001616 monocyte Anatomy 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- 230000035778 pathophysiological process Effects 0.000 description 1
- 230000002265 prevention Effects 0.000 description 1
- 230000009290 primary effect Effects 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N15/00—Investigating characteristics of particles; Investigating permeability, pore-volume, or surface-area of porous materials
- G01N15/10—Investigating individual particles
- G01N15/14—Electro-optical investigation, e.g. flow cytometers
-
- 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
-
- 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
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56966—Animal cells
-
- G01N15/149—
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2800/00—Detection or diagnosis of diseases
- G01N2800/24—Immunology or allergic disorders
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Immunology (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Cell Biology (AREA)
- Physics & Mathematics (AREA)
- Pathology (AREA)
- General Physics & Mathematics (AREA)
- General Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Analytical Chemistry (AREA)
- Medicinal Chemistry (AREA)
- Biotechnology (AREA)
- Food Science & Technology (AREA)
- Microbiology (AREA)
- Virology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Zoology (AREA)
- Dispersion Chemistry (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention provides a basophilic granulocyte activation and degranulation identification method. The basophilic granulocyte activation and degranulation identification method comprises the following steps that test samples are numbered in sequence, and flow type sample feeding pipes required by testing are marked; a required anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD203c and/or anti-human CD63 fluorescence labeling flow type antibody combination is added into each flow type sample feeding pipe; mixed whole blood is added into the marked flow type pipes, and shading incubation is conducted at the indoor temperature; red blood cell lysate is added into each sample pipe, and shading incubation is conducted at the indoor temperature again; supernatant is removed in a centrifugal mode after incubation; detection is conducted by means of a flow cytometry, and the number of basophilic granulocytes and the average fluorescence intensity are analyzed. According to the basophilic granulocyte activation and degranulation identification method, 80%-100% of the basophilic granulocytes in peripheral blood can be distinguished from other types of cells under the condition that the basophilic granulocytes do not need to be separated or purified, the basophilic granulocyte activation and/or degranulation state can be identified, whole blood of no more than 100 microlitres is needed by each sample to be tested, and the repeatability is good.
Description
Technical field
The present invention relates to the technical field of anaphylactia diagnosis, be specially the activation of a kind of basophilic granulocyte, degranulated authentication method.
Background technology
The allergic disease incidence of disease accounts for total world population's more than 30%, by one of the World Health Organization's four large noninfectious diseases being classified as 21 century.Along with the development of industrial economy, the change of ecologic environment, this type of disease is increasing in recent years, becomes common disease, frequently-occurring disease, is the significant problem that China's health and economic development field need to solve.
Since the dawn of human civilization, the definition of disease and the diagnosis scheme overwhelming majority are proposed by developed country.In the first that what splendid weighing apparatus in 2011, Zhang Huiyun hold in Tokyo on the irritated annual exchanging meeting of--Korea Spro three state, at home and abroad take the lead in having carried out revising discussion to the definition of the anaphylactia continuing to use nearly 40 years, original " anaphylactia is one group of disease mediated by IgE " is revised as " being one group of disease mediated by mast cell/basophilic granulocyte (MC/Bas) ".And the anaphylactia of IgE mediation is one of them hypotype, thus have modified the deviation that people are familiar with anaphylactia.This is because the root problem of allergic reaction (immediate hypersensitivity) be activate after primary effect cell-MC/Bas discharge inflammatory mediators, thus startup inflammation pathophysiological process, cause the appearance of clinical symptoms.Based on the definition of original " anaphylactia is one group of disease mediated by IgE ", what more external experts were artificial has descended the definition of " anaphylactoid reaction " the anaphylactia of those non-IgE mediations, namely " anaphylactoid reaction " clinical manifestation is identical with allergic reaction, methods for the treatment of is identical, but the IgE level in blood does not increase.People are strengthened further to the deviation that anaphylactia is familiar with.We will make people re-recognize this type of disease to the new definition of anaphylactia, greatly will improve prevention and the treatment level of anaphylactia.
Also be the definition based on " anaphylactia is one group of disease mediated by IgE ", in current serum clinically, the detection of allergenic specific IgE level has almost become the exclusive diagnosis method of anaphylactia, what even have human factor error thinks that this detection is anaphylactia diagnosis " goldstandard ", people are strengthened further to the deviation that anaphylactia is familiar with, and " goldstandard " that have forgotten anaphylactia diagnosis is completely this objective fact of allergen specificity provocative test.The clinical meaning detected due to specific IgE is to tell that IgE dependence autopath is to the former allergy of any Typical allergic, and to the allergic reaction of non-IgE dependence as most drug anaphylaxis, the food hypersenstivity of more than 80%, the patient of most of asthma, Small molecular in physical environment is as vehicle exhaust, the allergies such as ethanol are without any diagnostic significance, therefore the method worked out anaphylactia difinite quality diagnostic value is badly in need of clinically, thus help doctor to drug allergy or adverse drug reaction, food hypersenstivity or the diarrhoea of other reason, allergic asthma or intrinsic asthma etc. carry out antidiastole.
Allergen specificity provocative test is divided into provocative test and external provocative test two kinds in body, and the former is anaphylactia diagnosis " goldstandard ", but has certain risk, even can bring out severe allergic reaction, seldom carry out clinically; The latter's (comprising the provocative test of allergen specificity mast cell and allergen specificity basophilic granulocyte provocative test two kinds) is although be not as direct as the former on clinical meaning, and devoid of risk, therefore obtains and approve widely.But people's making great efforts many times through many decades, cannot obtain reliable external provocative test method all the time.For many years, as the specific marker thing of mast cell degranulation, histamine has been a great concern, but histamine detection system not only costly regrettably, function singleness, and data are stablized in more difficult acquisition, are difficult to clinically at present carry out.In addition, because mast cell is arranged in tissue, and except the nasal lavage fluid of Allergic Rhinitis, the mast cell being difficult to obtain autopath carries out provocative test, so cannot be used for clinical diagnosis at all.
The provocative test of allergen specificity basophilic granulocyte is the current uniquely potential provocative test method being applied to anaphylactia clinical detection.Be understood that, this provocative test method has three basic demands, and one is the specific recognition of basophilic granulocyte, and it two is that basophilic granulocyte activates, degranulated qualification, and it three is specific anaphylactogen.But up to the present not yet there is the method for the activation of a kind of reliable, reproducible specificity identification basophilic granulocyte in whole blood, retting conditions state regrettably, cause the provocative test of allergen specificity basophilic granulocyte to carry out.If our nearest research finds to detect CD123, HLA-DR, CCR3 simultaneously, CCR3+CD123+HLA-DR-cell mass 98-100% is basophilic granulocyte, thus take the lead in substantially solving the specific immunity identification problem of basophilic granulocyte, but do not solve that basophilic granulocyte activates, degranulated qualification problem.
Over nearly 10 years, it is found that CD203c can be used as the label of basophilic granulocyte activation, and be applied to clinical and experimental study gradually.Owing to only there being basophil cellular expression CD203c in blood, so the expression change of basophilic granulocyte CD203c, should accurately reflect its state of activation.CD63 is a kind of lysosome glycoprotein.Be expressed on basophilic granulocyte, activated blood platelet, monocyte/macrophage.CD63 molecule is mainly present on lysosome membrane, migrates to surface of cell membrane when basophilic granulocyte retting conditions, so detect the change of CD63 developed by molecule, can represent basophilic granulocyte retting conditions state.If thus detect CD123, HLA-DR, CCR3 simultaneously, CD203c and/or CD63, wherein CCR3+CD123+HLA-DR-CD203c+ or CCR3+CD123+HLA-DR-CD63+ cell mass represents the quantity of activation or degranulated basophilic granulocyte respectively, for solid foundation has been laid in the provocative test of allergen specificity basophilic granulocyte further.
Summary of the invention
Technical matters solved by the invention is to provide the activation of a kind of basophilic granulocyte, degranulated authentication method, to solve the problem in above-mentioned background technology.
Technical matters solved by the invention realizes by the following technical solutions: a kind of basophilic granulocyte activates, degranulated authentication method, comprises the steps:
(1) first according to the ratio of 9:1, with the ultrapure waters of 9 times, the concentrated erythrocyte cracked liquid storage liquid in kit is diluted to duty; Similarly with the ultrapure water of 9 times, the storage liquid of the concentrated phosphate buffer in kit is diluted to duty.Attention: erythrocyte cracked liquid suggestion matching while using.
(2) in sequence test sample is numbered, and the good streaming loading pipe needed for test of mark;
(3) in each streaming loading pipe, required Antibody Combination is added:
Combination 1: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-humen CD 20 3c of PE mark, consumption is each 5 μ L of every person-portion four kinds of antibody, to identify basophilic granulocyte state of activation;
Combination 2: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD63 fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-human CD63 of PE mark, consumption is each 5 μ L of every person-portion four kinds of antibody, to identify basophilic granulocyte retting conditions state;
Combination 3: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c, anti-human CD63 fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-human CD63 of anti-humen CD 20 3c, PE/Cy7 mark of PE mark, consumption is each 5 μ L of every person-portion five kinds of antibody, to identify that basophilic granulocyte activates and retting conditions state;
(4), after the mixing of patient whole blood's low speed whirlpool, draws with sample loading gun 25 μ L to the 125 μ L whole bloods mixed and be added in the streaming pipe that marked, in vortex oscillator, low speed whirlpool mixes 3 ~ 5s, and room temperature lucifuge hatches 15min;
(5) after hatching end, add the erythrocyte cracked liquid of 1mL in each sample tube, low speed whirlpool mixing 3 ~ 5s in vortex oscillator, room temperature lucifuge hatches 10min;
(6), after hatching end, the centrifugal 6min of 1200rpm/min rotating speed, after centrifugal end, abandoning supernatant, adds the PBS damping fluid of 1mL, with same centrifugation, (during centrifugal end sampling operation, should avoid acutely rocking cause cell precipitation to suspend);
(7) after centrifugal, remove supernatant in the same way, add the PBS damping fluid of 300 μ L, use flow cytomery.
(8) with counting and the average fluorescent strength of the corresponding analysis software basophilic granulocyte of different flow cytometer.
Described erythrocyte cracked liquid is provided by U.S. company BD, as please diluted according to reagent instructions with the erythrocyte cracked liquid of other companies.
Described fluorescence labeling streaming antibody comprises the fluorescently-labeled anti-human CD123 of FITC, APC, APC/Cy7, PE, PE/Cy7 or PerCP, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c and anti-human CD63 antibody.
Described Antibody Combination is any fluorescently-labeled anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c, anti-human CD63 five kinds of Antibody Combination.
Described Antibody Combination is any fluorescently-labeled anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c tetra-kinds of Antibody Combination.
Described Antibody Combination is any fluorescently-labeled anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD63 tetra-kinds of Antibody Combination.
Described Antibody Combination is the anti-human CD123 of any molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c, anti-human CD63 five kinds of Antibody Combination.
Described Antibody Combination is the anti-human CD123 of any molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c tetra-kinds of Antibody Combination.
Described Antibody Combination is the anti-human CD123 of any molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-human CD63 tetra-kinds of Antibody Combination.
Compared with public technology, there is following advantage in the present invention:
(1) basophilic granulocyte of peripheral blood 98-100% and other cell types can be distinguished;
(2) without the need to separation and purification basophilic granulocyte;
(3) every routine pattern detection only needs the whole blood being no more than 100 μ L;
(4) the inventive method is reproducible, and error is less than 10%
(5) can activate and/or retting conditions state by secured identification basophilic granulocyte.
Accompanying drawing explanation
Expressed by Fig. 1 .1 is CD123 positive cell group in blood;
Expressed by Fig. 1 .2 is CD123 positive HLA negative cells group in blood;
Expressed by Fig. 1 .3 is the negative CCR3 positive cell group of the positive HLA of CD123 and basophilic granulocyte in blood;
Expressed by Fig. 1 .4 is the negative CCR3 positive cell CD63 expression of the positive HLA of CD123 and retting conditions testing result in blood;
Expressed by Fig. 1 .5 is that in blood, namely the positive HLA of CD123 negative CCR3 positive cell CD203c expression activates testing result;
Testing result of the present invention, basophilic granulocyte activation, retting conditions testing result in blood.
Expressed by Fig. 2 .1 is CD123 positive cell group in blood;
Expressed by Fig. 2 .2 is CD123 positive HLA negative cells group in blood;
Expressed by Fig. 2 .3 is the negative CCR3 positive cell group of the positive HLA of CD123 and basophilic granulocyte in blood;
Expressed by Fig. 2 .4 is that in blood, namely the positive HLA of CD123 negative CCR3 positive cell CD203c expression activates testing result;
Testing result of the present invention, in blood, basophilic granulocyte activates testing result.
Expressed by Fig. 3 .1 is CD123 positive cell group in blood;
Expressed by Fig. 3 .2 is CD123 positive HLA negative cells group in blood;
Expressed by Fig. 3 .3 is the negative CCR3 positive cell group of the positive HLA of CD123 and basophilic granulocyte in blood;
Expressed by Fig. 3 .4 is the negative CCR3 positive cell CD63 expression of the positive HLA of CD123 and retting conditions testing result in blood;
Testing result of the present invention, basophilic granulocyte retting conditions testing result in blood.
Embodiment
Object is reached and effect is easy to understand in order to make technological means of the present invention, creation characteristic, workflow, using method, below in conjunction with the embodiment of the present invention, technical scheme in the embodiment of the present invention is clearly and completely described, obviously, described embodiment is only the present invention's part embodiment, instead of whole embodiments.Based on the embodiment in the present invention, those of ordinary skill in the art, not making the every other embodiment obtained under creative work prerequisite, belong to the scope of protection of the invention.
Embodiment 1
A kind of basophilic granulocyte activates, degranulated authentication method, as shown in Fig. 1 .1,1.2,1.3,1.4,1.5, comprises the steps:
(1) first according to the ratio of 9:1, with the ultrapure waters of 9 times, the concentrated erythrocyte cracked liquid storage liquid in kit is diluted to duty; Similarly with the ultrapure water of 9 times, the storage liquid of the concentrated phosphate buffer in kit is diluted to duty.Attention: erythrocyte cracked liquid suggestion matching while using;
(2) in order test sample is numbered, and the good streaming loading pipe needed for test of mark;
(3) in each streaming loading pipe, required Antibody Combination is added: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c and anti-human CD63 fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-human CD63 of anti-humen CD 20 3c, PE/Cy7 mark of PE mark, consumption is each 5 μ L of every person-portion five kinds of antibody, to identify that basophilic granulocyte activates and retting conditions state;
(4), after the mixing of patient whole blood's low speed whirlpool, draws with sample loading gun 25 μ L to the 125 μ L whole bloods mixed and be added in the streaming pipe that marked, in vortex oscillator, low speed whirlpool mixes 3 ~ 5s, and room temperature lucifuge hatches 15min;
(5) after hatching end, add the erythrocyte cracked liquid of 1mL in each sample tube, low speed whirlpool mixing 3 ~ 5s in vortex oscillator, room temperature lucifuge hatches 10min;
(6), after hatching end, the centrifugal 6min of 1200rpm/min rotating speed, after centrifugal end, abandoning supernatant, adds the PBS damping fluid of 1mL, with same centrifugation, (during centrifugal end sampling operation, should avoid acutely rocking cause cell precipitation to suspend);
(7) after centrifugal, remove supernatant in the same way, add the PBS damping fluid of 300 μ L, with flow cytomery;
(8) with counting and the average fluorescent strength of the corresponding analysis software basophilic granulocyte of different flow cytometer.
The streaming of fluorescence labeling described in the present embodiment antibody comprises the fluorescently-labeled anti-human CD123 of FITC, APC, APC/Cy7, PE, PE/Cy7 or PerCP, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c and anti-human CD63 antibody.
Antibody Combination described in the present embodiment is any fluorescently-labeled anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c, anti-human CD63 five kinds of Antibody Combination.
Antibody Combination described in the present embodiment is the anti-human CD123 of any molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c, anti-human CD63 five kinds of Antibody Combination.
Embodiment 2
The authentication method that basophilic granulocyte activates, as shown in Fig. 2 .1,2.2,2.3,2.4, comprises the steps:
(1) first according to the ratio of 9:1, with the ultrapure waters of 9 times, the concentrated erythrocyte cracked liquid storage liquid in kit is diluted to duty; Similarly with the ultrapure water of 9 times, the storage liquid of the concentrated phosphate buffer in kit is diluted to duty.Attention: erythrocyte cracked liquid suggestion matching while using;
(2) in order test sample is numbered, and the good streaming loading pipe needed for test of mark;
(3) in each streaming loading pipe, required Antibody Combination is added: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-humen CD 20 3c of PE mark, consumption is each 5 μ L of every person-portion four kinds of antibody, to identify basophilic granulocyte state of activation;
(4), after the mixing of patient whole blood's low speed whirlpool, draws with sample loading gun 25 μ L to the 125 μ L whole bloods mixed and be added in the streaming pipe that marked, in vortex oscillator, low speed whirlpool mixes 3 ~ 5s, and room temperature lucifuge hatches 15min;
(5) after hatching end, add the erythrocyte cracked liquid of 1mL in each sample tube, low speed whirlpool mixing 3 ~ 5s in vortex oscillator, room temperature lucifuge hatches 10min;
(6), after hatching end, the centrifugal 6min of 1200rpm/min rotating speed, after centrifugal end, abandoning supernatant, adds the PBS damping fluid of 1mL, with same centrifugation, (during centrifugal end sampling operation, should avoid acutely rocking cause cell precipitation to suspend);
(7) after centrifugal, remove supernatant in the same way, add the PBS damping fluid of 300 μ L, with flow cytomery;
(8) with counting and the average fluorescent strength of the corresponding analysis software basophilic granulocyte of different flow cytometer.
The streaming of fluorescence labeling described in the present embodiment antibody comprises the fluorescently-labeled anti-human CD123 of FITC, APC, APC/Cy7, PE, PE/Cy7 or PerCP, anti-human CCR3, anti-human HLA-DR and anti-humen CD 20 3c antibody.
Antibody Combination described in the present embodiment is any fluorescently-labeled anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c tetra-kinds of Antibody Combination.
Antibody Combination described in the present embodiment is the anti-human CD123 of any molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c tetra-kinds of Antibody Combination.
Embodiment 3
The degranulated authentication method of a kind of basophilic granulocyte, as shown in Fig. 3 .1,3.2,3.3,3.4, comprises the steps:
(1) first according to the ratio of 9:1, with the ultrapure waters of 9 times, the concentrated erythrocyte cracked liquid storage liquid in kit is diluted to duty; Similarly with the ultrapure water of 9 times, the storage liquid of the concentrated phosphate buffer in kit is diluted to duty.Attention: erythrocyte cracked liquid suggestion matching while using.
(2) in sequence test sample is numbered, and the good streaming loading pipe needed for test of mark;
(3) in each streaming loading pipe, required Antibody Combination is added: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD63 fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-human CD63 of PE mark, consumption is each 5 μ L of every person-portion four kinds of antibody, to identify basophilic granulocyte retting conditions state;
(4), after the mixing of patient whole blood's low speed whirlpool, draws with sample loading gun 25 μ L to the 125 μ L whole bloods mixed and be added in the streaming pipe that marked, in vortex oscillator, low speed whirlpool mixes 3 ~ 5s, and room temperature lucifuge hatches 15min;
(5) after hatching end, add the erythrocyte cracked liquid of 1mL in each sample tube, low speed whirlpool mixing 3 ~ 5s in vortex oscillator, room temperature lucifuge hatches 10min;
(6), after hatching end, the centrifugal 6min of 1200rpm/min rotating speed, after centrifugal end, abandoning supernatant, adds the PBS damping fluid of 1mL, with same centrifugation, (during centrifugal end sampling operation, should avoid acutely rocking cause cell precipitation to suspend);
(7) after centrifugal, remove supernatant in the same way, add the PBS damping fluid of 300 μ L, use flow cytomery.
(8) with counting and the average fluorescent strength of the corresponding analysis software basophilic granulocyte of different flow cytometer.
The streaming of fluorescence labeling described in the present embodiment antibody comprises the fluorescently-labeled anti-human CD123 of FITC, APC, APC/Cy7, PE, PE/Cy7 or PerCP, anti-human CCR3, anti-human HLA-DR and anti-human CD63 antibody.
Antibody Combination described in the present embodiment is any fluorescently-labeled anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD63 tetra-kinds of Antibody Combination.
Antibody Combination described in the present embodiment is the anti-human CD123 of any molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-human CD63 tetra-kinds of Antibody Combination.
More than show and describe ultimate principle of the present invention, principal character and advantage of the present invention.The technician of the industry should understand; the present invention is not restricted to the described embodiments; what describe in above-described embodiment and instructions just illustrates principle of the present invention; without departing from the spirit and scope of the present invention; the present invention also has various changes and modifications, and these changes and improvements all fall in the claimed scope of the invention.Claimed scope of the present invention is defined by appending claims and equivalent thereof.
Claims (15)
1. basophilic granulocyte activation, a degranulated authentication method, is characterized in that: comprise the steps:
1. first according to the ratio of 9:1, with the ultrapure waters of 9 times, the concentrated erythrocyte cracked liquid storage liquid in kit is diluted to duty; Similarly with the ultrapure water of 9 times, the storage liquid of the concentrated phosphate buffer in kit is diluted to duty;
2. in order test sample is numbered, and the good streaming loading pipe needed for test of mark;
3. in each streaming loading pipe, required Antibody Combination is added: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c and anti-human CD63 fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-human CD63 of anti-humen CD 20 3c, PE/Cy7 mark of PE mark, consumption is each 5 μ L of every person-portion five kinds of antibody, to identify that basophilic granulocyte activates and retting conditions state;
4., after the mixing of patient whole blood's low speed whirlpool, draws with sample loading gun 25 μ L to the 125 μ L whole bloods mixed and be added in the streaming pipe that marked, in vortex oscillator, low speed whirlpool mixes 3 ~ 5s, and room temperature lucifuge hatches 15min;
5. after hatching end, add the erythrocyte cracked liquid of 1mL in each sample tube, low speed whirlpool mixing 3 ~ 5s in vortex oscillator, room temperature lucifuge hatches 10min;
6. after hatching end, the centrifugal 6min of 1200rpm/min rotating speed, after centrifugal end, abandoning supernatant, adds the PBS damping fluid of 1mL, with same centrifugation;
7., after centrifugal, remove supernatant in the same way, add the PBS damping fluid of 300 μ L, with flow cytomery;
8. counting and the average fluorescent strength of the corresponding analysis software basophilic granulocyte of different flow cytometer is used.
2. basophilic granulocyte activation according to claim 1, degranulated authentication method, is characterized in that: described fluorescence labeling streaming antibody comprises the fluorescently-labeled anti-human CD123 of FITC, APC, APC/Cy7, PE, PE/Cy7 or PerCP, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c and anti-human CD63 antibody.
3. basophilic granulocyte activation according to claim 1, degranulated authentication method, is characterized in that: described Antibody Combination is the anti-human CD123 of any fluorescence labeling or molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c, anti-human CD63 five kinds of Antibody Combination.
4. basophilic granulocyte activation according to claim 1, degranulated authentication method, is characterized in that: described 25 μ L to the 125 μ L whole bloods mixed are any amount between 25 μ L to 125 μ L.
5. basophilic granulocyte activation according to claim 1, degranulated authentication method, is characterized in that: described flow cytometry analysis comprises counting and the average fluorescent strength of basophilic granulocyte.
6. an authentication method for basophilic granulocyte activation, is characterized in that: comprise the steps:
1. first according to the ratio of 9:1, with the ultrapure waters of 9 times, the concentrated erythrocyte cracked liquid storage liquid in kit is diluted to duty; Similarly with the ultrapure water of 9 times, the storage liquid of the concentrated phosphate buffer in kit is diluted to duty;
2. in order test sample is numbered, and the good streaming loading pipe needed for test of mark;
3. in each streaming loading pipe, required Antibody Combination is added: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-humen CD 20 3c of PE mark, consumption is each 5 μ L of every person-portion four kinds of antibody, to identify basophilic granulocyte state of activation;
4., after the mixing of patient whole blood's low speed whirlpool, draws with sample loading gun 25 μ L to the 125 μ L whole bloods mixed and be added in the streaming pipe that marked, in vortex oscillator, low speed whirlpool mixes 3 ~ 5s, and room temperature lucifuge hatches 15min;
5. after hatching end, add the erythrocyte cracked liquid of 1mL in each sample tube, low speed whirlpool mixing 3 ~ 5s in vortex oscillator, room temperature lucifuge hatches 10min;
6. after hatching end, the centrifugal 6min of 1200rpm/min rotating speed, after centrifugal end, abandoning supernatant, adds the PBS damping fluid of 1mL, with same centrifugation;
7., after centrifugal, remove supernatant in the same way, add the PBS damping fluid of 300 μ L, with flow cytomery;
8. counting and the average fluorescent strength of the corresponding analysis software basophilic granulocyte of different flow cytometer is used.
7. the authentication method of a kind of basophilic granulocyte activation according to claim 6, is characterized in that: described fluorescence labeling streaming antibody comprises the fluorescently-labeled anti-human CD123 of FITC, APC, APC/Cy7, PE, PE/Cy7 or PerCP, anti-human CCR3, anti-human HLA-DR and anti-humen CD 20 3c antibody.
8. the authentication method of a kind of basophilic granulocyte activation according to claim 6, is characterized in that: described Antibody Combination is the anti-human CD123 of any fluorescence labeling or molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-humen CD 20 3c tetra-kinds of Antibody Combination.
9. the authentication method of a kind of basophilic granulocyte activation according to claim 6, is characterized in that: described 25 μ L to the 125 μ L whole bloods mixed are any amount between 25 μ L to 125 μ L.
10. the authentication method of a kind of basophilic granulocyte activation according to claim 6, is characterized in that: described flow cytometry analysis comprises counting and the average fluorescent strength of basophilic granulocyte.
11. 1 kinds of degranulated authentication methods of basophilic granulocyte, is characterized in that: comprise the steps:
1. first according to the ratio of 9:1, with the ultrapure waters of 9 times, the concentrated erythrocyte cracked liquid storage liquid in kit is diluted to duty; Similarly with the ultrapure water of 9 times, the storage liquid of the concentrated phosphate buffer in kit is diluted to duty;
2. in order test sample is numbered, and the good streaming loading pipe needed for test of mark;
3. in each streaming loading pipe, required Antibody Combination is added: anti-human CD123, anti-human CCR3, anti-human HLA-DR, anti-human CD63 fluorescence labeling streaming Antibody Combination, as the anti-human HLA-DR of anti-human CCR3, APC/Cy7 mark of anti-human CD123, APC mark of FITC mark, the anti-human CD63 of PE mark, consumption is each 5 μ L of every person-portion four kinds of antibody, to identify basophilic granulocyte retting conditions state;
4., after the mixing of patient whole blood's low speed whirlpool, draws with sample loading gun 25 μ L to the 125 μ L whole bloods mixed and be added in the streaming pipe that marked, in vortex oscillator, low speed whirlpool mixes 3 ~ 5s, and room temperature lucifuge hatches 15min;
5. after hatching end, add the erythrocyte cracked liquid of 1mL in each sample tube, low speed whirlpool mixing 3 ~ 5s in vortex oscillator, room temperature lucifuge hatches 10min;
6. after hatching end, the centrifugal 6min of 1200rpm/min rotating speed, after centrifugal end, abandoning supernatant, adds the PBS damping fluid of 1mL, with same centrifugation, (during centrifugal end sampling operation, should avoid acutely rocking cause cell precipitation to suspend);
7., after centrifugal, remove supernatant in the same way, add the PBS damping fluid of 300 μ L, with flow cytomery;
8. counting and the average fluorescent strength of the corresponding analysis software basophilic granulocyte of different flow cytometer is used.
The degranulated authentication method of 12. basophilic granulocyte according to claim 11, is characterized in that: described fluorescence labeling streaming antibody comprises the fluorescently-labeled anti-human CD123 of FITC, APC, APC/Cy7, PE, PE/Cy7 or PerCP, anti-human CCR3, anti-human HLA-DR and anti-human CD63 antibody.
The degranulated authentication method of 13. basophilic granulocyte according to claim 11, is characterized in that: described Antibody Combination is the anti-human CD123 of any fluorescence labeling or molecular labeling, anti-human CCR3, anti-human HLA-DR, anti-human CD63 tetra-kinds of Antibody Combination.
The degranulated authentication method of 14. basophilic granulocyte according to claim 11, is characterized in that: described 25 μ L to the 125 μ L whole bloods mixed are any amount between 25 μ L to 125 μ L.
The degranulated authentication method of 15. basophilic granulocyte according to claim 1, is characterized in that: described flow cytometry analysis comprises counting and the average fluorescent strength of basophilic granulocyte.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510212435.7A CN104777303B (en) | 2015-04-29 | 2015-04-29 | Basophilic granulocyte activation and degranulation identification method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510212435.7A CN104777303B (en) | 2015-04-29 | 2015-04-29 | Basophilic granulocyte activation and degranulation identification method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104777303A true CN104777303A (en) | 2015-07-15 |
| CN104777303B CN104777303B (en) | 2017-02-01 |
Family
ID=53618900
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510212435.7A Active CN104777303B (en) | 2015-04-29 | 2015-04-29 | Basophilic granulocyte activation and degranulation identification method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104777303B (en) |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105004705A (en) * | 2015-08-07 | 2015-10-28 | 天津中医药大学 | Detection method of medicine injection anaphylactic and anaphylactoid reactions and new application of adopted dye |
| CN109387636A (en) * | 2017-08-08 | 2019-02-26 | 欧蒙医学诊断技术有限公司 | Method for examining basicyte to activate |
| CN111549091A (en) * | 2020-05-21 | 2020-08-18 | 核工业总医院 | Test method for basophil activation and degranulation |
| WO2023046111A1 (en) * | 2021-09-23 | 2023-03-30 | 上海市第一人民医院 | Basophil activation detection method and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP2037269A1 (en) * | 2007-09-11 | 2009-03-18 | Bühlmann Laboratories AG | Allergy test based on flow cytometric analysis |
| WO2010043724A1 (en) * | 2008-10-17 | 2010-04-22 | Mead Johnson & Company | Method for identifying allergenic proteins and peptides |
| CN102590492A (en) * | 2012-02-22 | 2012-07-18 | 江苏苏中药业集团股份有限公司 | Method for detecting anaphylactoid reaction of Shengmai liquid preparation |
-
2015
- 2015-04-29 CN CN201510212435.7A patent/CN104777303B/en active Active
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| EP2037269A1 (en) * | 2007-09-11 | 2009-03-18 | Bühlmann Laboratories AG | Allergy test based on flow cytometric analysis |
| WO2010043724A1 (en) * | 2008-10-17 | 2010-04-22 | Mead Johnson & Company | Method for identifying allergenic proteins and peptides |
| CN102590492A (en) * | 2012-02-22 | 2012-07-18 | 江苏苏中药业集团股份有限公司 | Method for detecting anaphylactoid reaction of Shengmai liquid preparation |
Non-Patent Citations (3)
| Title |
|---|
| GUILLAUME MONNERET ET AL.: "Monitoring of Basophil Activation Using CD63 and CCR3 in Allergy to Muscle Relaxant Drugs", 《CLINICAL IMMUNOLOGY》 * |
| 孙林等: "外周血嗜碱粒细胞的定量研究", 《中国医药指南》 * |
| 张皓月等: "流式细胞术分析嗜碱性粒细胞活化试验在过敏性疾病诊断中的应用", 《中国医药导报》 * |
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105004705A (en) * | 2015-08-07 | 2015-10-28 | 天津中医药大学 | Detection method of medicine injection anaphylactic and anaphylactoid reactions and new application of adopted dye |
| CN105004705B (en) * | 2015-08-07 | 2018-05-01 | 天津中医药大学 | The detection method of drug injection allergy anaphylactoid reaction and dyestuff new application used |
| CN109387636A (en) * | 2017-08-08 | 2019-02-26 | 欧蒙医学诊断技术有限公司 | Method for examining basicyte to activate |
| CN111549091A (en) * | 2020-05-21 | 2020-08-18 | 核工业总医院 | Test method for basophil activation and degranulation |
| CN111549091B (en) * | 2020-05-21 | 2023-11-17 | 核工业总医院 | Method for testing activation and degranulation of basophils |
| WO2023046111A1 (en) * | 2021-09-23 | 2023-03-30 | 上海市第一人民医院 | Basophil activation detection method and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104777303B (en) | 2017-02-01 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104777303A (en) | Basophilic granulocyte activation and degranulation identification method | |
| Jung et al. | Immature platelet fraction: establishment of a reference interval and diagnostic measure for thrombocytopenia | |
| NZ630277A (en) | Methods and compositions for diagnosis and prognosis of renal injury and renal failure | |
| CN103805696B (en) | A kind of microRNA molecule mark of diagnostics classes rheumatic arthritis and detection kit thereof | |
| Saxena et al. | Sonoclot signature analysis in patients with liver disease and its correlation with conventional coagulation studies | |
| Mansoor et al. | Reverse pseudohyperkalemia: an important clinical entity in chronic lymphocytic leukemia | |
| CN106399510A (en) | Method for analyzing urine exosome miRNA | |
| JP2019032317A5 (en) | ||
| Tailliar et al. | Urinary peptides as potential non-invasive biomarkers for Lupus Nephritis: results of the Peptidu-LUP Study | |
| CN104807988A (en) | Specific immunity recognition method for basophilic granulocyte | |
| Prihirunkit et al. | Hemostatic markers in congestive heart failure dogs with mitral valve disease | |
| CN108795692A (en) | Rare cell capture systems and its application | |
| CN203625357U (en) | Circulating tumor cell separation system | |
| CN102818890A (en) | Method for detecting autoantibodies of systemic lupus erythematosus patient | |
| CN104792687B (en) | A kind of authentication method of basophilic granulocyte sensitization | |
| CN107121551B (en) | Biomarker combinations, detection kit and the application of nasopharyngeal carcinoma | |
| CN113125757B (en) | Protein biomarker for early pregnancy diagnosis of sows and method for detecting early pregnancy of sows by using protein biomarker | |
| Wee et al. | A composite of urinary biomarkers for differentiating between tubulointerstitial inflammation and interstitial fibrosis/tubular atrophy in kidney allografts | |
| CN102654502A (en) | Protein marker for chronic hepatitis liver fibrosis and detection method thereof | |
| CN107102127B (en) | Monokaryon sample marrow source inhibits the detection method of cell in a kind of human peripheral blood | |
| Zahedipour et al. | Development of flow cytometry-fluorescent in situ hybridization (Flow-FISH) method for detection of PML/RARa Chromosomal Translocation in acute promyelocytic leukemia cell line | |
| Haque et al. | Evaluation of Biochemical marker for the diagnosis of Rheumatoid arthritis | |
| CN209450535U (en) | Haemolysis indicates constant volume heparin tube | |
| CN103540658A (en) | Method, primer and kit for detecting hot mutation site of human XPD (Xeroderma Pigmentosum group D) gene | |
| CN204188440U (en) | Cell mass gatherer |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| EXSB | Decision made by sipo to initiate substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C41 | Transfer of patent application or patent right or utility model | ||
| TA01 | Transfer of patent application right |
Effective date of registration: 20160516 Address after: 121004, Fairview District, Taihe District, Jinzhou City, Liaoning Province, D, 35-97 Applicant after: He Shaoheng Address before: 121001 No. five, Section 2, Renmin Street, Guta District, Liaoning, Jinzhou Applicant before: No.1 Hospital Attached To Liaoning Medical College |
|
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20170630 Address after: 121010, 29, Yu Jie street, Taihe District, Liaoning, Jinzhou Patentee after: Liaoning universal source of biomedical science and Technology Development Co., Ltd. Address before: 121004, Fairview District, Taihe District, Jinzhou City, Liaoning Province, D, 35-97 Patentee before: He Shaoheng |
|
| PE01 | Entry into force of the registration of the contract for pledge of patent right | ||
| PE01 | Entry into force of the registration of the contract for pledge of patent right |
Denomination of invention: Basophilic granulocyte activation and degranulation identification method Effective date of registration: 20190808 Granted publication date: 20170201 Pledgee: Jinzhou Bank Co., Ltd. Shifulu Branch of Science and Technology Pledgor: Liaoning universal source of biomedical science and Technology Development Co., Ltd. Registration number: Y2019990000043 |
|
| PC01 | Cancellation of the registration of the contract for pledge of patent right | ||
| PC01 | Cancellation of the registration of the contract for pledge of patent right |
Date of cancellation: 20200408 Granted publication date: 20170201 Pledgee: Jinzhou Bank Co., Ltd. Shifulu Branch of Science and Technology Pledgor: LIAONING HUIPUYUAN BIOMEDICAL TECHNOLOGY DEVELOPMENT Co.,Ltd. Registration number: Y2019990000043 |