CN103105326A - Method for surface staining and intracellular staining of Th cells and application of method - Google Patents

Method for surface staining and intracellular staining of Th cells and application of method Download PDF

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Publication number
CN103105326A
CN103105326A CN2012105879444A CN201210587944A CN103105326A CN 103105326 A CN103105326 A CN 103105326A CN 2012105879444 A CN2012105879444 A CN 2012105879444A CN 201210587944 A CN201210587944 A CN 201210587944A CN 103105326 A CN103105326 A CN 103105326A
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il
add
pipe
cell
500rpm
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CN2012105879444A
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宁光
谢晓雁
杨颖�
崔斌
郭婷
朱巍
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上海市内分泌代谢病研究所
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Abstract

The invention discloses a method for surface staining and intracellular staining of Th cells and application of the method. The method comprises the steps of; stimulating the cells, blocking off an Fc receptor on the surface of a cell membrane, staining antigen on the surface of the cell membrane, fixing the cells, penetrating the cell membrane, staining intracellular or intranuclear antigens, and carrying out flow cytometry. The method provided by the invention can achieve surface staining and intracellular staining of the Th cells, and has a good experiment effect. The method can be applied in testing the proportion of Th1, Th2, and Th17 cells to CD4<+> cells.

Description

Method and the application thereof of dyeing in a kind of Th cell surface and born of the same parents

Technical field

The present invention relates to method and the application thereof of dyeing in a kind of Th cell surface and born of the same parents, belong to the endocrine technical field.

Background technology

AITD (autoimmune thyroid disease, AITD) is the organ specificity autoimmune disease, take destroyed as feature to the thyroid antigen self tolerance.Hashimoto's thyroiditis (Hashimoto ' s thyroidtis, HT) all belong to typical autoimmune thyroid disease with Graves sick (GD), volume lymphocyte and thick liquid cell, NK cell and macrophages infiltration are all arranged in thyroid gland, the most extensive with the lymphocytic infiltration of HT.Generally believe at present, HT is because the suppressive lymphocyte T function reduces, the helper lymphocyte T effect strengthens, make the bone-marrow-derived lymphocyte differentiation produce a large amount of anti-thyroid antibodies, wherein modal is antiperoxide enzyme antibody (TPOAb), and it is principal feature that the cytotoxicity by the mediation of antibody dependent and natural killer cell causes parathyroid tissue structural failure and hypothyroidism.

CD4 +The T cell is the important cells that body is carried out immune response, initial CD4 +After the T cell is activated by antigenic stimulus, can be divided into effector cell Thl, Th2, Thl7 or regulatory T cells (Regulatory T cells, Treg) under different cell factor microenvironments.The Th1 cell mainly discharges the cell factors such as IFN-γ, IL-12, TNF-β, activating macrophage, neutrophil leucocyte etc., performance cell toxicant and Phagocytosis, mediated cell immunity.The cell factor that the Th2 cell is mainly secreted has IL-4, IL-5, IL-10 and IL-13 etc., promotes the B cell to discharge IgG, IgA, IgE antibody, humoral immunity.Thl7 emiocytosis IL-17A, IL-17F, IL-6 and IL-9 etc., experimental animal model demonstration Th17 cell and organ specificity autoimmune inflammation reaction relation are close.Treg cells specificity suppresses CD4 +, CD8 +The activation of T cell and propagation, thereby the generation of prevention autoimmune response.In the immune response process, can mutually stimulate by secreted cell factor between immunocyte, constraint each other, thus immune response is regulated.

Cell factor participates in, regulates, affects, promotes this process as immunocyte correctives, immune effector molecule, inflammatory reaction promoter.The change of cell factor, prompting Thl/Th2 function has extremely in AITD patient, and this activity that extremely can affect adjusting thyroid cell functional network system promotes thyroid antibody to form and increases, and impels the generation of AITD, development.

Generally believe that the cell immune response that the Th1 cytokines mediates plays a major role in the HT morbidity.Wherein representational cell factor is IFN-γ.It promotes the lymphocytic infiltration in thyroid gland on the one hand, promotes on the other hand lymphocyte and the macrophage activation that infiltrates and discharges the generation of the cell factor such as TNF-α, IL-1, IL-6 and oxygen radical, and these materials can cause parathyroid tissue to destroy.

The generation of the generation of GD and thyrotrophin receptor antibody (TRAb) has direct relation, and it is the main and immediate cause that causes GD.TRAb belongs to the IgG1 subclass, and the Th2 cytokines promotes the generation of IgG1 subclass.The immune response of GD is the Th2 type, and its immune response is take humoral immunity as main.The overaction of TRAb simulation TSH stimulates thyroid follicular cells, causes hyperthyroidism [3]The Th cell has multiple hypotype, comprises the Th17 hypotype that can secrete IL-17A, demonstrates significance in the empirical model of organ specificity autoimmune inflammation.

And how from the different intracellular cell factors of unicellular horizontal detection, be that association area has technical matters to be solved.

Summary of the invention

The objective of the invention is method and the application thereof of dyeing in a kind of Th cell surface and born of the same parents in order to provide, so that a kind of method from the different intracellular cell factors of unicellular horizontal detection to be provided.

Purpose of the present invention can be achieved through the following technical solutions:

In a kind of Th cell surface and born of the same parents, the method for dyeing, the steps include:

1) add the aseptic RPMI1640 training liquid mixing that contains 10% calf serum of 1ml anticoagulant heparin whole blood and 1ml in a streaming mensuration pipe.

2) add Buddhist ripple ester 2ul, ionomycin 2ul, coban 2ul at above-mentioned Guan Zhongzai; Cover lid, at 37 ℃, 5% CO 2Cultivate in incubator and be no more than 3.5 hours and (must not surpass 3.5 hours, otherwise the cd4 cell yield reduce.Lid does not cover too tight).

3) in 3 streaming pipes, add respectively IFN γ/CD4/IL4 system, be labeled as IL-4, add IFN-γ/CD4/IL-17 system, be labeled as IL-17, and negative control pipe (not adding any antibody).Get step 2) the light and slow mixing of putting upside down of whole blood nutrient solution, take out 600 μ l and be added to respectively in three streaming pipes.

4) every streaming pipe adds the aseptic erythrocyte cracked liquid of 4ml, puts upside down mixing for several times, places 10min, splitting erythrocyte.

5) sample after splitting erythrocyte is centrifugal 1,500rpm, 5min abandons supernatant.

6) every pipe adds 1ml PBS damping fluid and plays gently for several times washing and precipitating, and is centrifugal 1,500rpm, and 5min abandons the supernatant antibody PE-Cy5-CD4 antibody 10 μ l that label, and negative tube does not add, and 4 ℃ of lucifuges are hatched 30min.

7) every pipe adds 1ml PBS damping fluid and plays gently for several times washing, and is centrifugal 1,500rpm, and 5min abandons supernatant, and control is done.

8) every Guan Jialeng fixing agent: 500 μ l, 4% paraformaldehyde; Fixing 20min, lucifuge is placed.

9) every pipe adds 1ml PBS damping fluid and plays gently for several times and clean, and is centrifugal 1,500rpm, and 5min abandons supernatant.Add 500 μ l, 1% saponin is worn film, and lucifuge is placed 10min.

10) 1,500rpm, 5min, the centrifugal supernatant of abandoning.

11) add FITC-IFN-gamma antibodies 0.5ug in being labeled as the pipe of IL-4, PE-IL-4 antibody 0.125ug, FITC-IFN-gamma antibodies 0.5ug in the pipe of IL-17, PE-IL-17 antibody 0.25ug.

12) above-mentioned 3 streaming pipes are placed in the darkroom, after incubated at room was spent the night, the up flow type instrument detected.

Described flow cytometer detects and adopts BECTON DICKINSON FACSCalibur flow cytometer to detect, and CellsQuest software is used in interpretation of result.

Activator: under static state, substantially do not express in non-activated lymphocyte or only express a small amount of cell factor, thereby being difficult to detect.Only have after it is activated, the cell intrinsic factor could be synthesized increase, therefore detects the cell within a cell factor and must utilize activator that it is activated.

Blocking agent is selected: cell is after activation, cell factor is synthetic to be increased, but constantly secrete to the extracellular and have an effect, this has just caused the difficulty that detects its cell within a cell factor level with fluidic cell, therefore also wants blocking agent to stop its cytokine secretion just can detect to born of the same parents.The blocking agent that uses in this test is coban.Coban is by suppressing the cell within a cell factor to cell exocrine, be accumulated in cell after making cell factor synthetic, but, coban can produce dosage, time dependent cytotoxicity, therefore should grope its experiment condition in use, usually finish to use in front 6~12 hours in irritant reaction.

Activationary time: according to the difference that detects index and sample, need to select different stimulant and stimulation time, to obtain best experimental result.

Sample disposal: avoid using the anti-coagulants of complexing calcium, can limit Ca-dependent activation as EDTA(because of it), so the recommendation liquaemin.In addition, blood sample is tested 8h planted agent, surpasses 8h and can cause the cytoactive loss, and general cell factor positive cell can reduce 5%.As not testing in 8h, the horizontal room temperature of vacuum test tube should be placed.

Its ultimate principle is to use fluorescein-labeled specificity antibacterial agent monoclonal antibody (as IFN, IL-4, IL-17 etc.) and anti-cell surface molecular monoclonal antibody (as CD4, CD25 etc.), can be from the different intracellular cell factors of unicellular horizontal detection by FCM immunofluorescence technique (as FACS), and can judge thus the cell category, celluar localization, distribution density and the cell factor that produce the specific cells factor and lesion tissue relation etc.

The present invention by irritation cell → blocking-up surface of cell membrane Fc acceptor → surface of cell membrane antigen dyeing → cell fix → cell permeable membrane → born of the same parents in or the step such as core endoantigen dyeing → flow cytometry, can be to dyeing in Th cell surface and born of the same parents, and obtain experiment effect preferably.Can be applicable to test Th1, Th2, the Th17 cell accounts for CD4 +The ratio of cell.

Embodiment

Further set forth technical scheme of the present invention below in conjunction with specific embodiment.

Embodiment

1, selected crowd

GD group 25 examples and Hashimoto first subtract group 25 routine patients and all arrive 2011-01 Shanghai City Ruijin Hospital endocrine outpatient service from 2010-06.The physical examination of healthy population that Normal group comes 25 examples to be complementary from age and sex.Above-mentioned onset GD patient 25 examples, not yet treatment.Control GD patient's 25 examples stationary phase.Above-mentioned physical examination of healthy population 25 examples.Onset GD group patient's diagnosis is with reference to the diagnostic criteria of GD in " clinical practice " sixth version and " the consonance endocrine metabolism is learned " sixth version.The GD patient who makes a definite diagnosis meets one of following condition and can be selected in.There is thyroid gland

Symptom hyperactivity, companion or do not accompany expophthalmos, thyroid gland anthorisma (palpation or B ultrasonic confirm), matter are soft; Thyroid function checks (CMIA method; Architect system, Abbott Laboratories, USA): free T3 (FT3), free T4 (FT4) raise, super quick thyrotropic hormone (sensitive thyroidStimulatinghormone, STSH) level reduces.Positive (the ECLIA method of TA (TRAb); RocheDiagnostics).

The Hashimoto first subtracts diagnosis reference " clinical practice " sixth version of group patient and the diagnostic criteria in " the consonance endocrine metabolism is learned " sixth version: the thyroid gland moderate swelling, quality is hard.Total T4(TT4), FT4 lowers, STSH significantly raises thyroid function checks:.Thyroglobulin antibody (TgAb) and antiperoxide enzyme antibody (TPOAb) titre obviously raise.

The inspection of the stable group of GD treatment thyroid function: free T3 (FT3), free T4 (FT4), super quick thyrotropic hormone level are in normal range, and TRAb the moon turns.All selected research objects are all got rid of history, women's gestation, the smoking of infection in the recent period, merge other immunity diseases and take immunopotentiator or the inhibitor history.All select newly to examine still untreated patient for two groups.Biochemical analysis and hormone determination blood thyroid hormone are detected by Shanghai City Ruijin Hospital endocrine research institute.THE (chemoluminescence method; Architectsystem, AbbottLaboratories, USA); TRAb (ECLIA method; Roche Diagnostics).TgAb (CMIA method; Architect system, Abbott Laboratories.USA); TPOAb (CMIA method; Architect system, Abbott Laboratories.USA).

2, sample is prepared: all research objects all 9 o'clock to the 11 o'clock morning empty stomach gather venous blood 6ml, 3ml be from blood coagulation, the packing of extraction serum is frozen in-80 ℃, centralized detecting.3ml collects with aseptic heparin anticoagulant tube, is placed in room temperature, and namely processed noon on the same day, and standby flow cytometer detects.

3, dyeing in Th cell surface and born of the same parents:

1) add the aseptic RPMI1640 training liquid mixing that contains 10% calf serum of 1ml anticoagulant heparin whole blood and 1ml in a streaming mensuration pipe.

2) add Buddhist ripple ester 2ul, ionomycin 2ul, coban 2ul at above-mentioned Guan Zhongzai; Cover lid, at 37 ℃, 5% CO 2Cultivate in incubator and be no more than 3.5 hours and (must not surpass 3.5 hours, otherwise the cd4 cell yield reduce.Lid does not cover too tight).

3) in 3 streaming pipes, add respectively IFN-γ/CD4/IL-4 system, be labeled as IL-4, add IFN-γ/CD4/IL-17 system, be labeled as IL-17, and negative control pipe (not adding any antibody).Get step 2) the light and slow mixing of putting upside down of whole blood nutrient solution, take out 600 μ l and be added to respectively in three streaming pipes.

4) every streaming pipe adds the aseptic erythrocyte cracked liquid of 4ml, puts upside down mixing for several times, places 10min, splitting erythrocyte.

5) sample after splitting erythrocyte is centrifugal 1,500rpm, 5min abandons supernatant.

6) every pipe adds 1ml PBS damping fluid and plays gently for several times washing and precipitating, and is centrifugal 1,500rpm, and 5min abandons the supernatant antibody PE-Cy5-CD4 antibody 10 μ l that label, and negative tube does not add, and 4 ℃ of lucifuges are hatched 30min.

7) every pipe adds 1ml PBS damping fluid and plays gently for several times washing, and is centrifugal 1,500rpm, and 5min abandons supernatant, and control is done.

8) every Guan Jialeng fixing agent: 500 μ l, 4% paraformaldehyde; Fixing 20min, lucifuge is placed.

9) every pipe adds 1ml PBS damping fluid and plays gently for several times and clean, and is centrifugal 1,500rpm, and 5min abandons supernatant.Add 500 μ l, 1% saponin is worn film, and lucifuge is placed 10min.

10) 1,500rpm, 5min, the centrifugal supernatant of abandoning.

11) add FITC-IFN-gamma antibodies 0.5ug in being labeled as the pipe of IL-4, PE-IL-4 antibody 0.125ug, FITC-IFN-gamma antibodies 0.5ug in the pipe of IL-17, PE-IL-17 antibody 0.25ug.

12) above-mentioned 3 streaming pipes are placed in the darkroom, after incubated at room was spent the night, the up flow type instrument detected.

Testing result: Th1 cell proportion: the Hashimoto first is kept to (17.82% ± 1.70%), and GD is that (15.72% ± 0.86%) is all higher than Normal group (12.47% ± 1.09%) (P<0.05); GD group Th2 ratio (1.78% ± 0.19%) is significantly higher than Normal group (0.99% ± 0.13%) (P<0.01), and the Hashimoto first subtracts group Th2 ratio (1.59% ± 0.20%), higher than Normal group (P<0.05).Th17 cell proportion: GD, Hashimoto first subtract, control group is respectively (2.30% ± 0.12%), (2.87% ± 0.31%), (1.90% ± 0.11%).Two groups of patients' Th17 accounts for CD4 +Cell proportion all higher than control group just, (P<005).

Claims (1)

1. the method for dyeing in a Th cell surface and born of the same parents, is characterized in that: the steps include:
1) add the aseptic RPMI1640 training liquid mixing that contains 10% calf serum of 1ml anticoagulant heparin whole blood and 1ml in a streaming mensuration pipe;
2) add Buddhist ripple ester 2ul, ionomycin 2ul, coban 2ul at above-mentioned Guan Zhongzai; Cover lid, at 37 ℃, 5% CO 2Cultivate in incubator and be no more than 3.5 hours
3) in 3 streaming pipes, add respectively IFN-γ/CD4/IL-4 system, be labeled as IL-4, add IFN-γ/CD4/IL-17 system, be labeled as IL-17, and the negative control pipe; Get step 2) the light and slow mixing of putting upside down of whole blood nutrient solution, take out 600 μ l and be added to respectively in three streaming pipes;
4) every streaming pipe adds the aseptic erythrocyte cracked liquid of 4ml, puts upside down mixing for several times, places 10min, splitting erythrocyte;
5) sample after splitting erythrocyte is centrifugal 1,500rpm, 5min abandons supernatant;
6) every pipe adds 1ml PBS damping fluid and plays gently for several times washing and precipitating, and is centrifugal 1,500rpm, and 5min abandons the supernatant antibody PE-Cy5-CD4 antibody 10 μ l that label, and negative tube does not add, and 4 ℃ of lucifuges are hatched 30min;
7) every pipe adds 1ml PBS damping fluid and plays gently for several times washing, and is centrifugal 1,500rpm, and 5min abandons supernatant, and control is done;
8) every Guan Jialeng fixing agent: 500 μ l, 4% paraformaldehyde; Fixing 20min, lucifuge is placed;
9) every pipe adds 1ml PBS damping fluid and plays gently for several times and clean, and is centrifugal 1,500rpm, and 5min abandons supernatant; Add 500 μ l, 1% saponin is worn film, and lucifuge is placed 10min;
10) 1,500rpm, 5min, the centrifugal supernatant of abandoning;
11) add FITC-IFN-gamma antibodies 0.5ug in being labeled as the pipe of IL-4, PE-IL-4 antibody 0.125ug, FITC-IFN gamma antibodies 0.5ug in the pipe of IL-17, PE-IL-17 antibody 0.25ug;
12) above-mentioned 3 streaming pipes are placed in the darkroom, after incubated at room was spent the night, the up flow type instrument detected.
CN2012105879444A 2012-12-30 2012-12-30 Method for surface staining and intracellular staining of Th cells and application of method CN103105326A (en)

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Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103884562A (en) * 2014-04-10 2014-06-25 上海太阳生物技术有限公司 Chloroacetate AS-D naphythol AS-D chloroacetate esterase (AS-DNCE) staining solution (chemical coloring process)
CN105593685A (en) * 2013-08-01 2016-05-18 苏伯利莫尔公司 In vitro method for determining the stability of compositions comprising soluble Fc gamma receptor(s)
CN107314965A (en) * 2017-04-26 2017-11-03 马鞍山易廷生物科技有限公司 The Sample pretreatment method detected based on streaming combination ICP MS single cell proteins

Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20040229368A1 (en) * 2003-05-14 2004-11-18 Beckman Coulter, Inc. Method and apparatus for preparing cell samples for intracellular antigen detection using flow cytometry
US20060013900A1 (en) * 2000-12-26 2006-01-19 Council Of Scientific & Industrial Research Organisation Use of betel leaf extract to induce IFN-gamma production from human peripheral blood T cells and as a Th1 type immunomodulator
CN1824303A (en) * 2005-12-23 2006-08-30 中国农业大学 Medicine for treating and/or preventing hepatitis B
CN101130074A (en) * 2007-09-12 2008-02-27 中国农业大学 Medicament for treating and/or preventing hepatitis B
CN101298615A (en) * 2004-10-26 2008-11-05 中国医学科学院基础医学研究所 Novel anti-plasmodium falciparum epitope and vaccine containing the same
WO2010007213A1 (en) * 2008-07-18 2010-01-21 Suomen Punainen Risti, Veripalvelu A method of evaluating the integrity of the plasma membrane of cells by detecting glycans found only intracellularly
CN102198192A (en) * 2011-05-26 2011-09-28 浙江省中医院 Traditional Chinese medicine composition for treating HINI and application thereof

Patent Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20060013900A1 (en) * 2000-12-26 2006-01-19 Council Of Scientific & Industrial Research Organisation Use of betel leaf extract to induce IFN-gamma production from human peripheral blood T cells and as a Th1 type immunomodulator
US20040229368A1 (en) * 2003-05-14 2004-11-18 Beckman Coulter, Inc. Method and apparatus for preparing cell samples for intracellular antigen detection using flow cytometry
CN101298615A (en) * 2004-10-26 2008-11-05 中国医学科学院基础医学研究所 Novel anti-plasmodium falciparum epitope and vaccine containing the same
CN1824303A (en) * 2005-12-23 2006-08-30 中国农业大学 Medicine for treating and/or preventing hepatitis B
CN101130074A (en) * 2007-09-12 2008-02-27 中国农业大学 Medicament for treating and/or preventing hepatitis B
WO2010007213A1 (en) * 2008-07-18 2010-01-21 Suomen Punainen Risti, Veripalvelu A method of evaluating the integrity of the plasma membrane of cells by detecting glycans found only intracellularly
CN102198192A (en) * 2011-05-26 2011-09-28 浙江省中医院 Traditional Chinese medicine composition for treating HINI and application thereof

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
王勇等: "《系统性红斑狼疮患者Th0、Thl和Th2细胞水平检测》", 《安徽医科大学学报》, vol. 41, no. 3, 30 June 2006 (2006-06-30), pages 321 *
高伟等: "《佛波酯对人外周血CD4+T细胞产生IL-17的影响》", 《西南大学学报》, vol. 32, no. 4, 30 April 2010 (2010-04-30) *

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN105593685A (en) * 2013-08-01 2016-05-18 苏伯利莫尔公司 In vitro method for determining the stability of compositions comprising soluble Fc gamma receptor(s)
CN105593685B (en) * 2013-08-01 2017-11-24 苏伯利莫尔公司 For the in-vitro method for the stability for determining the composition comprising soluble Fc γ acceptors
CN103884562A (en) * 2014-04-10 2014-06-25 上海太阳生物技术有限公司 Chloroacetate AS-D naphythol AS-D chloroacetate esterase (AS-DNCE) staining solution (chemical coloring process)
CN107314965A (en) * 2017-04-26 2017-11-03 马鞍山易廷生物科技有限公司 The Sample pretreatment method detected based on streaming combination ICP MS single cell proteins

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