CN107064085A - Test Procedur of Splenic Natural Killer Cell Activity - Google Patents

Test Procedur of Splenic Natural Killer Cell Activity Download PDF

Info

Publication number
CN107064085A
CN107064085A CN201710155968.5A CN201710155968A CN107064085A CN 107064085 A CN107064085 A CN 107064085A CN 201710155968 A CN201710155968 A CN 201710155968A CN 107064085 A CN107064085 A CN 107064085A
Authority
CN
China
Prior art keywords
cell
natural killer
target
cell activity
splenic
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
Application number
CN201710155968.5A
Other languages
Chinese (zh)
Other versions
CN107064085B (en
Inventor
张斌
汤永永
陈虎
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Chinese People's Liberation Army Three \/ Seven Hospital
Original Assignee
Chinese People's Liberation Army Three \/ Seven Hospital
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Chinese People's Liberation Army Three \/ Seven Hospital filed Critical Chinese People's Liberation Army Three \/ Seven Hospital
Priority to CN201710155968.5A priority Critical patent/CN107064085B/en
Publication of CN107064085A publication Critical patent/CN107064085A/en
Application granted granted Critical
Publication of CN107064085B publication Critical patent/CN107064085B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Biomedical Technology (AREA)
  • Molecular Biology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Physics & Mathematics (AREA)
  • Chemical & Material Sciences (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • General Physics & Mathematics (AREA)
  • Immunology (AREA)
  • Pathology (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

A kind of Test Procedur of Splenic Natural Killer Cell Activity, " NK cell proportions " and " karyocyte motility rate " parameter is introduced when effector cell and target ration calculate, and carries out quality control in each link, reduces detection otherness.

Description

Test Procedur of Splenic Natural Killer Cell Activity
Technical field
The present invention relates to a kind of Test Procedur of Splenic Natural Killer Cell Activity, belong to field of biological detection.
Background technology
NK cells (Natural killer cells, NK cell, NK) are initially because without presensitization Just autologous tumor cell can be killed, and is referred to as innate immunity lymph toxic cell.By for decades to its phenotype and function Careful research, finding the principal biological function of NK cells mainly has secrete cytokines to participate in immunological regulation and to target cell Direct killing effect [1,2].NK cell distributions Various Functions in different tissues, not only with antitumor, the various diseases of participation Reason process, maintenance immune homeostasis, may additionally facilitate gestational period placenta vascularization process.Correspondingly, malignant tumour, various senses Dye, autoimmune disease etc. all refer to NK cell disorders [3-6].
NK cell killing activities are the important indicators for evaluating immune state, available for immune shape during monitoring immunization therapy In terms of the change of state.Existing document report, its also with Survival after pernicious Chemotherapy of Cervical Cancer curative effect, pancreatic cancer chemotherapy and Ah Alzheimer's disease tempo correlation [7-9].In familial or acquired Hemophagocytic lymphohistocysis disease, inflammation It is one of its feature that NK killing activities are impaired in disease, autoimmune disease and immunologic deficiency disease, but because the index is respectively tested Room result can not compare, and lack normal reference value, and the confirmation frequently as other diagnosis indexs in relevant disease diagnosis refers to Mark [10].The rigorous, determination method of standardization and corresponding normal reference value are badly in need of in the detection of NK killing activities, solve it The bottleneck problem of application.
" the goldstandard method " of NK killing detection methods is 51Cr release experiments, but is limited the problems such as due to radioactive pollution Its application [11,12] in clinical detection.Other methods for being used for the detection of NK cell killing activities also include low toxicity dyestuff Calcein-AM release experiments, LDH release experiments, the detection method [13-17] based on flow cytometry or bioluminescence.Wherein Detection method based on flow cytometry have it is easy to operate, stably, pollute less, baseline it is low and with 51Cr release experiment method phases The advantages of closing property is good, existing clinic is more to carry out NK cell killing activities detection [14,18] using the method.
But the method for prior art is only calculated with number of nucleated cells and target cell number, may result in the inclined of testing result Move, it is impossible to realize the rigorous assessment to NK cellkilling capacities.
The content of the invention
The present invention provides a kind of Test Procedur of Splenic Natural Killer Cell Activity, it is characterised in that comprise the following steps：
1) by target cell Secondary Culture；
2) NK ratio is analyzed in pairing effect cell；
3) by target cell with effector cell according to the effect target of setting than being cultivated；
4) detecting step 3) in target cell death rate to be measured；
5) by step 4) in target cell death rate bring into formula calculate natural killer cell activity：
Natural killer cell activity=(target cell death rate to be measured-control group target cell death rate)/(1- control group targets are thin Born of the same parents' death rate) × 100%.
Further, step 3) in effect target ratio be more than 4.
Further, step 1) in enter the quality control of line density and/or the death rate to passage cell.
Further, step 2) in pairing effect cell carry out cell density, living cell rate and NK ratio extremely One of few quality control.
Further, the method for quality control includes the target cell of Secondary Culture carrying out dye marker.
Further, including with antibody NK is marked.
Further, control target cell natural mortality rate is within 3%.
Further, the ratio between target cell and effector cell's FH1 passages MIF (average fluorescent strength) value are more than 80 times.
Further, step 4) in, give up the following detection number of degrees：Compared with only target cell control tube, effect target cell mixes Close be incubated detection pipe intracellular targets concentration less than the former 85%.
Further, tumour selection effect target ratio whether was suffered from according to detection object.
The present invention establishes a kind of brand-new NK cell killing vigor testing methods, in effector cell and target ration meter " NK cell proportions " and " karyocyte motility rate " parameter is introduced during calculation, the preciseness of the detection is improved, at the same with by NK cells Carry out detection after sorting to compare, in the absence of NK cell magnetic bead sortings are with high costs and mechanical damage of selected by flow cytometry apoptosis etc. is asked Topic.
This method improves NK cell killing vitality assessments between separate sources sample compared with conventional traditional detection method Comparativity, with broader practice prospect.
Present invention firstly provides carry out quality control to each link of the detection, it is therefore an objective to reduces the otherness detected between room, It is expected to solve the present situation that each laboratory detection result can not be referred to mutually.
The present invention assesses the human peripheral N K cell killing vigor normal reference value models for being available for different experiments room to refer to first Enclose.
Brief description of the drawings
The NK cell proportion regularities of distribution in Fig. 1 sample karyocytes to be checked
Fig. 2 tumor patients group organizes peripheral blood NK cell killing activity with healthy volunteer
Embodiment
1 data and method
1.1 research objects are chosen to go to a doctor in affiliated hospital of Military Medical Science Institute and had a medical check-up in June, 2014 in October, 2016 Healthy volunteer and tumor patient totally 108, collects age, sex and NK killing activities detection correlated results.Separately collect other NK killing activities detect the karyocyte NK percent informations of sample 158.Reference value group inclusion criteria：1. healthy population；2. blood As normal；3. informed consent form is signed.Reference value exclusion standard：1. immune correlated disease person；2. there are long-term steroid and immune tune Save agent application history person；3. there is clear and definite the infected in 3 weeks；4. the person that has cardiovascular and cerebrovascular disease；5. it is doubtful or clarify a diagnosis for tumour suffer from Person；6. other influences immune function disease or medical history.
1.2 reagents and instrument PI fluorescent dyes, the analysis of NK cells of human beings subgroup monoclonal antibody (CD45-APC, CD56- PE, CD3-PerCP) and its Isotype control be purchased from U.S. company BD, CSFE fluorescent dyes are purchased from Japanese colleague company, and people's lymph is thin Born of the same parents' separating liquid is purchased from the foreign China Tech bio tech ltd of Tianjin Hao, and cell culture is purchased from 1640 culture mediums and hyclone Thremo Fisher companies.Detection BD Accuri C6 flow cytometers.
1.3 experimental method
1.3.1 the Secondary Culture of target cell：K562 cells press 1-2 × 105Cells/ml density, is inoculated in containing 20ml's In 1640 complete medium T75 blake bottles, passage in every three days once, maintains its good vegetative state.
1.3.2 the dye marker of target cell：K562 suspension cultures are collected in 15ml centrifuge tubes, horizontal centrifugal is placed in 1000rpm is centrifuged 10 minutes in machine.With brine twice after, unicellular is resuspended in 500ul serum-frees 1640 and trained Support in base, sampled after fully mixing, count stand-by using C6.It is another to take the serum free medium of certain volume 1640 to dilute CSFE dyestuffs Working solution is simultaneously fully mixed, and is rapidly added in stand-by K562 cell suspensions and mixed, and it is 5 μM to make CSFE dye concentrations, Cell suspension density is 1 × 107cells/ml.It is placed in 37 DEG C of 5%CO2In incubator, shake once within every 5 minutes.It is incubated 10 points The hyclone of isometric 4 DEG C of precoolings is added after clock, is placed in 4 DEG C of refrigerators and terminates staining reaction 10 minutes.Supply physiological saline 1000rpm is centrifuged 10 minutes after mixing, is abandoned supernatant and is resuspended in 1640 complete mediums, is placed in incubator and is incubated 1 hour.Receive Collect cell, once, cell is resuspended in 1640 complete mediums to brine, and sampling counts stand-by with C6.
1.3.3 the separation of effector cell：If sample is whole blood, then PMNC (Peripheral is carried out Blood mononuclear cell, PBMC) separation, i.e., by whole blood sample 1800rpm centrifuge 10 minutes, go after blood plasma wait body Product physiological saline gravity treatment cell, is gently added in after being centrifuged on lymphocyte separation medium, carefully siphons away tunica albuginea layer.Washed with physiology salt Wash twice, cell is resuspended in 1640 complete mediums, sampling is diluted in white blood cell count(WBC) liquid, count stand-by with C6.If sample is to exempt from Epidemic disease cell culture product, then only with brine twice.Sampling is diluted in 1640 complete mediums, is counted and treated with C6 With.
1.3.4 the mark of effector cell NK cell subsets：Take containing about 5 × 105The 100ul effects of individual mononuclearcell (PBMC) Cell suspension is answered, CD3-CD56+NK cell subsets antibody labelings is carried out by flow cytometer detection antibody specification, uses BD FACS Calibur flow cytometers are detected, analyze NK cell proportions in karyocyte.PBMC then carries out erythrocyte splitting step in this way Suddenly, in this way immunocyte cultured products then without erythrocyte splitting step.
1.3.5 the detection of effector cell's living cell rate：Such as sample is immunocyte cultured products, then needing detection, it has core thin Born of the same parents' living cell rate.100ul cell suspensions are taken, PI dyestuffs is added to final concentration 3.75ug/ml, balances 5-10 minutes, fluidic cell Instrument detects effector cell's death rate.
1.3.6 effector cell is mixed with target cell：Setting effect target ratio (NK cells and mark CSFE target cell it Than) it is 0: 1,1: 1,2: 1,4: 1 and 6: 1, according in effector cell's karyocyte density, effector cell's motility rate and its karyocyte Effector cell's volume, melange effect cell and target cell, are complemented to 1640 complete mediums needed for NK cell proportions are calculated 200ul co-culture systems.The multiple holes of each group at least three, mix and are incubated 2.5 hours altogether.
1.3.7 target cell death rate is detected：The mixed culture cell of 1.3.6 steps is resuspended, PI dyestuffs are added to final concentration 3.75ug/ml, is balanced 5-10 minutes, target cell death rate positive each pipe CSFE of flow cytomery.
1.3.8NK the calculating of cell killing vigor：Bring each pipe target cell death rate into formula (experimental group target cell death Rate-control group target cell death rate)/(1- control group target cell deaths rate) × 100%, calculate each effect target and lived than NK cell killing Mild-natured average.
1.4 quality control
1.4.1 the quality control of target cell observes growth conditions and speed by constant density inoculation passage K562 cells, The indexs such as cell cycle are detected if necessary.Control balance durations of the K562 in 1640 complete mediums after CSFE marks, it is desirable to K562 cell lines natural mortality rate after mark is less than 3%, target cell and effector cell's FH1 passage average fluorescent strengths The ratio between (Median Fluorescence Intensity, MIF) value is more than 80 times, ensures that the target used of detection every time is thin with this The uniformity of born of the same parents.
1.4.2 the quality control EDTA anticoagulant tubes collection periphery blood specimen of effector cell, in vitro to be sent to reality in 3 hours afterwards Room is tested, room temperature placement is placed in, is resuspended in when effector cell is standby in 1640 complete mediums.As non-fresh blood sample or it is long when Between be positioned over other conditions, karyocyte motility rate need to be detected, with the cell density of karyocyte, living cell rate and NK ratios calculate Each effect target is than required PBMC suspension volumes.
1.4.3 the quality control of flow cytometer is calibrated by instrument fixed maintenance, with FH3 Air conduct measurement PI fluorescent dyes Signal, the manual measuring effect cell of cell absolute number function replacement or target cell suspension density are measured with Accuri C6.
1.4.4 the same technical staff of the quality control of data analysis carries out reading under same standard, if any data Accept or reject and analysis is then completed jointly by fixing three technical staff.Compared with only target cell control tube, effect target cell mixing is incubated Educate detection pipe intracellular targets concentration less than the former 85% when, it is believed that be completely severed after the target cell death of part, cause testing result Less than actual target cell death rate, then the testing result is insincere.
1.5 statistical methods carry out statistical analysis using SPSS21.0 softwares, and each index is represented with X ± s, and factor is related Property analysis use between multiple linear regression analysis, two groups mean to compare to use independent samples t test.P ＜ 0.05 are that difference has system Meter learns meaning.
2 results
2.1 include research object this basic condition
Normal reference value group includes healthy volunteer 47, women 22 [- 68 years old 28 years old, average (47.5 ± 9.1) year], Male 25 [- 69 years old 27 years old, average (47.3 ± 11.2) year].Tumor group includes all kinds of tumor patient 61 [liver cancer 23, pancreas Gland cancer 8, lung cancer and each 5 of colon cancer, the carcinoma of the rectum, stomach cancer, breast cancer and each 3 of oophoroma, thyroid cancer 2, kidney, the moon Road cancer, hypopharyngeal cancer, cancer of the esophagus, hepatobiliary cancer and each 1 of cholangiocarcinoma], women 18 [- 84 years old 36 years old, average (60.2 ± 11.9) Year], male 43 [- 85 years old 17 years old, average (53.9 ± 13.5) year] refers to table 1.
The ordinary circumstance table (example) of table 1
2.2NK proportional difference
NK cells occupy nucleus ratio distribution situation (see Fig. 1) in each source sample in analysis NK killing activity detections, As a result the parameter distribution situation is 10.8% ± 7.87% (3%-42%) in prompting tumor patient group, and the degree of bias is 1.714, kurtosis For 3.609；The parameter distribution situation is 12.31% ± 5.25% (4%-23%) in healthy volunteer person's group, and the degree of bias is 0.576, kurtosis is -0.860；The parameter distribution situation is 13.48% ± 12.30% (0.11%- in other source groups 75.19%), the degree of bias is 3.095, and kurtosis is 11.283, it can be seen that in each source NK killing activities sample karyocyte to be checked The NK cell proportion regularities of distribution are different, do not consider the greatest differences of the parameter when calculating effector cell with target ration such as, Only calculated with number of nucleated cells and target cell number, may result in the skew of testing result, it is impossible to realized to NK cell killing energy The rigorous assessment of power.
2.3NK cell killing activity Study on Relative Factors
It will be grouped by less than 44 years old, -59 years old 45 years old, more than 60 years old at age, analyze age, sex and whether suffered from It is related between tumour and NK killing activities.As a result point out age, sex and whether suffered from tumour three when imitating target ratio and being 1 and 2 Non-correlation (P ＞ 0.05) between person and NK killing activities.When effect target ratio is 4 and 6, sex, age property associated therewith are without statistics Meaning (P ＞ 0.05) is learned, suffered from tumour property whether associated therewith statistically significant (P ＜ 0.05).Based on this data Analysis, age and Effect of gender peripheral blood NK cell killing ability are not found, and whether suffering from tumour will produce to the index Influence, and effect target ratio should be set in 4 or more than 6.
2.4NK cell killing activity normal reference values
It is that age and sex are unrelated with NK cell killing activities to consider this research healthy control group Data Representation, therefore true When determining reference range, no longer it is grouped according to age or sex.The determination of term of reference uses method of percentiles, sets NK The reference range of cell killing activity bilateral 90%, the results are shown in Table 2.
The peripheral blood NK cell killing activity normal reference value (%) of table 2
The NK killing activity features of 2.5 tumor patients
Comparison of tumor patient organizes organizes peripheral blood NK cell killing activity difference with healthy volunteer, as a result the target score of prompting effect Not Wei 4 and 6 when, two group differences are statistically significant (P ＞ 0.05).When it is 1 and 2 to imitate target ratio, two group differences are without statistics Learn meaning (P ＜ 0.05).Point out the tumor patient index to be less than health donors, and the detection project be considered as set imitate target ratio as 4 and 6.
Bibliography
1.Cooper, M.A., et al., Human natural killer cells：a unique innate Immunoregulatory role for the CD56 (bright) subset.Blood, 2001.97 (10)：p.3146-51.
2.Cerwenka, A.and L.L.Lanier, Natural killer cells, viruses and Cancer.Nat Rev Immunol, 2001.1 (1)：p.41-9.
3.Yu, J., A.G.Freud, and M.A.Caligiuri, Location and cellular stages of natural killer cell development.Trends Immunol.34(12)：p.573-82.
4.Vivier, E., et al., Functions of natural killer cells.Nat Immunol, 2008.9(5)：p.503-10.
5.Parham, P.and A.Moffett, Variable NK cell receptors and their MHC Class I ligands in immunity, reproduction and human evolution.Nat Rev Immunol.13(2)：p.133-44.
6.Orange, J.S., Natural killer cell deficiency.J Allergy Clin Immunol.132(3)：p.515-25；quiz 526.
7.Cosiski Marana, H.R. .J.Santana da Silva, and J.Moreira de Andrade, NK cell activity in the presence of IL-12is a prognostic assay to neoadjuvant Chemotherapy in cervical cancer.Gynecol Oncol, 2000.78 (3Pt 1)：p.318-23.
8.Prolo, P., et al., Physiologic modulation of natural killer cell Activity as an index of Alzheimer ' s disease progression.Bioinformation, 2007.1 (9)：p.363-6.
9.Karakhanova, S., et al., Prognostic and predictive value of immunological parameters for chemoradioimmunotherapy in patients with pancreatic adenocarcinoma.Br J Cancer.112(6)：p.1027-36.
10.Ishii, E., Hemophagocytic Lymphohistiocytosis in Children： Pathogenesis and Treatment.Front Pediatr.4：p.47.
11.Brunner, K.T., et al., Quantitative assay of the lytic action of immune lymphoid cells on 51-Ct-labelled allogeneic target cells in vitro； Inhibition by isoantibody and by drugs.Immunology, 1968.14 (2)：p.181-96.
12.Fehniger, T.A., et al., Potential mechanisms of human natural killer Cell expansion in vivo during low-dose IL-2therapy.J Clin Invest, 2000.106 (1)： p.117-24.
13.Lichtenfels, R., et al., CARE-LASS (calcein-release-assay), an improved fluorescence-based test system to measure cytotoxic T lymphocyte activity.J Immunol Methods, 1994.172 (2)：p.227-39.
14.Valiathan, R., et al., Evaluation of a flow cytometry-based assay for natural killer cell activity in clinical settings.Scand J Immunol.75(4)： p.455-62.
15.Cho, D., et al., Cytotoxicity of activated natural killer cells against pediatric solid tumors.Clin Cancer Res.16(15)：p.3901-9.
16.Konjevic, G., V.Jurisic, and I.Spuzic, Association of NK cell dysfunction with changes in LDH characteristics of peripheral blood Lymphocytes (PBL) in breast cancer patients.Breast Cancer Res Treat, 2001.66 (3)： p.255-63.
17.Karimi, M.A., et al., Measuring cytotoxicity by bioluminescence imaging outperforms the standard chromium-51 release assay.PLoS One.9(2)： p.e89357.
18.Kim, G.G., et al., A novel multiparametric flow cytometry-based cytotoxicity assay simultaneously immunophenotypes effector cells：comparisons To a 4 h 51Cr-release assay.J Immunol Methods, 2007.325 (1-2)：p.51-66.
19.Defining, establishing, and verifying RIs in the clinical aboratory.Approved guideline 3rd Ed.Clinical Laboratory and Standards Institute, 2008.
20.Mace, E.M., et al., Cell biological steps and checkpoints in accessing NK cell cytotoxicity.Immunol Cell Biol.92(3)：p.245-55.

Claims (10)

1. a kind of Test Procedur of Splenic Natural Killer Cell Activity, it is characterised in that comprise the following steps：
1) by target cell Secondary Culture；
2) NK ratio is analyzed in pairing effect cell；
3) by target cell with effector cell according to the effect target of setting than being cultivated；
4) detecting step 3) in target cell death rate to be measured；
5) by step 4) in target cell death rate bring into formula calculate natural killer cell activity：
Natural killer cell activity=(target cell death rate to be measured-control group target cell death rate)/(1- control group target cells are dead Die rate) × 100%.
2. Test Procedur of Splenic Natural Killer Cell Activity as claimed in claim 1, it is characterised in that step 3) in effect target ratio be More than 4.
3. Test Procedur of Splenic Natural Killer Cell Activity as claimed in claim 1, it is characterised in that step 1) in passage cell Enter the quality control of line density and/or the death rate.
4. Test Procedur of Splenic Natural Killer Cell Activity as claimed in claim 1, it is characterised in that step 2) in pairing effect cell Carry out the quality control of at least one cell density, living cell rate and NK ratio.
5. Test Procedur of Splenic Natural Killer Cell Activity as claimed in claim 3, it is characterised in that the method for quality control bag Include and the target cell of Secondary Culture is subjected to dye marker.
6. Test Procedur of Splenic Natural Killer Cell Activity as claimed in claim 4, it is characterised in that including with antibody to killing naturally Hinder cell to be marked.
7. Test Procedur of Splenic Natural Killer Cell Activity as claimed in claim 3, it is characterised in that control target cell natural death Rate is within 3%.
8. Test Procedur of Splenic Natural Killer Cell Activity as claimed in claim 3, it is characterised in that target cell and effector cell The ratio between FH1 passages MIF (average fluorescent strength) value is more than 80 times.
9. the Test Procedur of Splenic Natural Killer Cell Activity as described in one of claim 1-8, it is characterised in that step 4) in, house Abandon the following detection number of degrees：Compared with only target cell control tube, effect target cell mixing is incubated detection pipe intracellular targets concentration and is less than The former 85%.
10. Test Procedur of Splenic Natural Killer Cell Activity as claimed in claim 1, it is characterised in that whether foundation detection object Suffered from tumour selection effect target ratio.
CN201710155968.5A 2017-03-16 2017-03-16 Method for measuring natural killer cell activity Active CN107064085B (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201710155968.5A CN107064085B (en) 2017-03-16 2017-03-16 Method for measuring natural killer cell activity

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201710155968.5A CN107064085B (en) 2017-03-16 2017-03-16 Method for measuring natural killer cell activity

Publications (2)

Publication Number Publication Date
CN107064085A true CN107064085A (en) 2017-08-18
CN107064085B CN107064085B (en) 2021-05-11

Family

ID=59618292

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201710155968.5A Active CN107064085B (en) 2017-03-16 2017-03-16 Method for measuring natural killer cell activity

Country Status (1)

Country Link
CN (1) CN107064085B (en)

Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110231275A (en) * 2019-05-20 2019-09-13 西安交通大学 A method of based on Flow cytometry NK cell killing activity
CN112285081A (en) * 2020-10-28 2021-01-29 上海睿钰生物科技有限公司 Method for detecting cell killing efficacy and application thereof
CN112285083A (en) * 2020-10-28 2021-01-29 上海睿钰生物科技有限公司 Method for evaluating cell killing efficacy
CN112304851A (en) * 2020-10-28 2021-02-02 上海睿钰生物科技有限公司 Evaluation method of in vitro natural killer cell immunocompetence and application thereof
CN113899725A (en) * 2021-10-12 2022-01-07 江苏省人民医院（南京医科大学第一附属医院） Method for real-time quantitative detection of degranulation and killing capacity of NK effector cells

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1399675A (en) * 1999-11-26 2003-02-26 理化学研究所 Method of proliferating natural killer cells
US20110104128A1 (en) * 2009-10-30 2011-05-05 The Board Of Regents, The University Of Texas System Device and Method for Transfecting Cells for Therapeutic Use

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1399675A (en) * 1999-11-26 2003-02-26 理化学研究所 Method of proliferating natural killer cells
US20110104128A1 (en) * 2009-10-30 2011-05-05 The Board Of Regents, The University Of Texas System Device and Method for Transfecting Cells for Therapeutic Use

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
李媛等: "HBV感染者自然杀伤细胞活性分析", 《肝脏》 *

Cited By (8)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN110231275A (en) * 2019-05-20 2019-09-13 西安交通大学 A method of based on Flow cytometry NK cell killing activity
CN110231275B (en) * 2019-05-20 2021-05-28 西安交通大学 Method for detecting killing activity of NK (natural killer) cells based on flow cytometry
CN112285081A (en) * 2020-10-28 2021-01-29 上海睿钰生物科技有限公司 Method for detecting cell killing efficacy and application thereof
CN112285083A (en) * 2020-10-28 2021-01-29 上海睿钰生物科技有限公司 Method for evaluating cell killing efficacy
CN112304851A (en) * 2020-10-28 2021-02-02 上海睿钰生物科技有限公司 Evaluation method of in vitro natural killer cell immunocompetence and application thereof
CN112285081B (en) * 2020-10-28 2021-11-19 上海睿钰生物科技有限公司 Method for detecting cell killing efficacy and application thereof
CN113899725A (en) * 2021-10-12 2022-01-07 江苏省人民医院（南京医科大学第一附属医院） Method for real-time quantitative detection of degranulation and killing capacity of NK effector cells
CN113899725B (en) * 2021-10-12 2022-04-29 江苏省人民医院（南京医科大学第一附属医院） Method for real-time quantitative detection of degranulation and killing capacity of NK effector cells

Also Published As

Publication number Publication date
CN107064085B (en) 2021-05-11

Similar Documents

Publication Publication Date Title
CN107064085A (en) Test Procedur of Splenic Natural Killer Cell Activity
Kandarian et al. A flow cytometry-based cytotoxicity assay for the assessment of human NK cell activity
Doulabi et al. Analysis of Th22, Th17 and CD4+ cells co-producing IL-17/IL-22 at different stages of human colon cancer
Dean et al. The relative proliferation index as a more sensitive parameter for evaluating lymphoproliferative responses of cancer patients to mitogens and alloantigens
CN109100501A (en) Measure the test of cell mediated immune response
CN103725648A (en) Novel CTC (circulating tumor cell) enrichment technology and preparation method of kit
JP2024529420A (en) A platform for co-culture imaging to characterize the in vitro efficacy of heterotypic effector cell therapy in cancer
Holmberg-Thyden et al. A user's guide to multicolor flow cytometry panels for comprehensive immune profiling
CN103792364A (en) Reagent for detection of ROR1 protein of circulating tumor cells in peripheral blood and application thereof
Niu et al. Characteristics and reference ranges of CD4+ T cell subpopulations among healthy adult Han Chinese in Shanxi Province, North China
CN103243074A (en) Human colorectal adenocarcinoma tumor cell line as well as preparation method and application thereof
Huang et al. Preliminary report on interleukin-22, GM-CSF, and IL-17F in the pathogenesis of acute anterior uveitis
Dai et al. Balance of CD8+ CD28+/CD8+ CD28− T lymphocytes is vital for patients with ulcerative colitis
Misra et al. Flow-based sorting of neonatal lymphocyte populations for transcriptomics analysis
JP2000510687A (en) How to measure lymphocyte function
CN114791411A (en) Index combination, kit and method for evaluating human immune function
Duriancik et al. The identification and enumeration of dendritic cell populations from individual mouse spleen and Peyer's patches using flow cytometric analysis
Silva et al. Stabilization of blood for long-term storage can affect antibody-based recognition of cell surface markers
CN104458539A (en) Method and kit for detecting lymphocyte proliferation conditions through non-diagnostic purpose
Patuzzo et al. Immunophenotypic analysis of B lymphocytes in patients with common variable immunodeficiency: identification of CD23 as a useful marker in the definition of the disease
CN105823882B (en) ATP is used to evaluate the purposes and evaluation method of immunocompetence as biomarker
Wu et al. A comprehensive multiparameter flow cytometry panel for immune profiling and functional studies of frozen tissue, bone marrow, and spleen
Zeng et al. FoxP3 promotes lymph node metastasis in patients with papillary thyroid carcinoma complicated with Hashimoto’s thyroiditis
CN110373445A (en) A kind of preparation method and application method of the quick drug effect test kit of anti-tumor drug
RU2581925C2 (en) Method for assessing allogenic immune response in short-term mixed mononuclear culture of unrelated donors

Legal Events

Date Code Title Description
PB01 Publication
PB01 Publication
SE01 Entry into force of request for substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant