CN1493687A - Interleukin-15 gene modified natural killing cell strain and its preparation method - Google Patents
Interleukin-15 gene modified natural killing cell strain and its preparation method Download PDFInfo
- Publication number
- CN1493687A CN1493687A CNA031529682A CN03152968A CN1493687A CN 1493687 A CN1493687 A CN 1493687A CN A031529682 A CNA031529682 A CN A031529682A CN 03152968 A CN03152968 A CN 03152968A CN 1493687 A CN1493687 A CN 1493687A
- Authority
- CN
- China
- Prior art keywords
- cell
- interleukin
- genetic modification
- cell strain
- strain
- 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
Images
Abstract
An interleukin-15 (IL-15) gene modified natural killing cell strain for immunotherapy of tumor is prepared through inserting the cDNA coding region of IL-15 in pcDNA3 eucaryotic expression carrier, configuring secretion-type recombinant eucaryotic expression carrier pc DNA3/IL-15, using liposom to transfecte cell NK-92, adding Geneticin (G418) to screening culture medium, screening, limited dilution to cloned cells, detecting activity, choosing positive cell clones, and amplification.
Description
One. technical field
The present invention relates to cell adoptive immunotherapy tumor area, more specifically, the present invention relates to natural killer cell strain of genetic modification and preparation method thereof.
Two. technical background
(natural killer, NK) cell plays an important role in antitumor, the antiviral process of body natural killer.Be to separate the NK cell to be used for clinical treatment from human peripheral, still, there are technical difficulty in the separation of NK cell, purifying and vitro culture in the past.Therefore, this area need a kind of, growth convenient in vitro culture fast, active high to tumor cytotoxicity, service efficiency height, the NK cell strain that toxic side effect is low.Wherein, the NK-92 cell is a kind of NK clone with high-efficiency broad spectrum antitumor action of setting up from the non_hodgkin lymphoma patient, has good potential applicability in clinical practice (GongJH, et al.Leukemia, 1994,8:652; Tam YK, et al.J Heamatother, 1999,8:281).(interleukin-2 IL-2), when clinical application, needs to inject IL-2 in some cases but the propagation of NK-92 cell and biologic activity depend on interleukin II., because the toxic side effect of IL-2 makes NK-92 be subjected to restriction to a certain degree in clinical application.In order to solve this defective, Nagashima S and Tam YK successively go into the NK-92 cell with the IL-2 gene transfection according to NK-92 to the dependency of IL-2, have obtained NK cell strain (Nagashima S, et al.Blood, 1998,91:3850 that non-IL-2 relies on; Tam YK, et al.Human Gene Therapy, 1999,10:1359).Overcome dependency, when clinical application, do not needed to inject IL-2 though people such as Nagashima S and Tam YK obtain the NK cell strain of IL-2 genetic modification IL-2, but, the NK cell of its biological characteristics and unmodified has similarity, and self can secrete the IL-2 of doses, still have the potential unfavorable factor, do not reach the ideal requirement.
Three. summary of the invention
1. technical problem
The purpose of this invention is to provide NK cell strain of a kind of IL-15 of utilization genetic modification and preparation method thereof,, improve propagation and kill capability, make the NK cell be more suitable for clinical application to change the biological characteristics of NK cell.
2. technical scheme
The NK cell strain of IL-15 genetic modification, its phenotype are the CD56 strong positive, and CD16 and CD3 are all negative, it is characterized in that form is dispersive half an adherent growth state.On this basis, the expression rate of the necessary adhesion molecule CD54 of NK cell killing reaches 90～100%, and average fluorescent strength reaches 220.The expression rate of the surface of cell membrane molecule CD25 relevant with NK cell proliferation reaches 50～80%.The expression rate of the surface of cell membrane molecule CD69 relevant with the NK cell activation reaches 15～65%.The average fluorescent strength of the surface of cell membrane molecule CD48 relevant with the NK cell activation reaches 435.
The method for preparing above-mentioned cell strain is: the pcDNA3 carrier for expression of eukaryon is inserted in the cDNA coding region of IL-15, make up secretor type recombinant eukaryon expression vector pcDNA3/IL-15; Utilize liposome transfection NK-92 cell; Adding Geneticin (G418) in screening culture medium screens; The limited dilution cloning cell, the activity with IL-15 in the dependent cells strain mensuration clone cell culture supernatant of IL-15 selects the positive cell clone of expressing IL-15; This is cloned in α-MEM substratum of the IL-15 that contains 10% calf serum, 8～12.5% horse serums, 20～100 activity units increases, obtain having the engineering cell strain of clinical value.
That is to say that the cytokine of the present invention's regulating effect from NK cytodifferentiation growth course is started with, on the one hand consider in the multiple in vivo tissue of Interleukin-15 (IL-15) that expression is all arranged, the differentiation and development process of NK cell is played an important role; On the other hand, because the NK-92 cell is the NK cell of not differentiation and maturation, cell surface can be expressed the IL-15 acceptor.And the toxic side effect of IL-15 is more much smaller than IL-2.So, the present invention adopts the secretor type recombinant eukaryon expression vector transfection NK-92 cell of IL-15, in the hope of obtaining to express the NK cell of IL-15, by IL-15 autocrine loop, improve the multiplication capacity of NK cell, to the service efficiency of the killing activity and the NK cell of tumour cell, obviously reduce toxic side effect, more be applicable to clinical application.Its phenotype of NK cell strain of the IL-15 genetic modification that is obtained is still all negative for CD56 strong positive, CD16 and CD3, and form is dispersive, half adherent growth state by poly-growth alteration before modifying, and helps the growth of NK cell and killing and wounding target cell more.And, the necessary adhesion molecule CD54 of NK cell killing, the surface of cell membrane molecule CD25 relevant, surface of cell membrane molecule CD69 and the expression rate of CD48 or the raising that average fluorescent strength all have significance degree relevant with the NK cell activation with NK cell proliferation.Described average fluorescent strength is meant the density of the surface of cell membrane molecule that utilizes Flow cytometry.The kind of employed liposome is unrestricted in preparation process.The dependent cells strain of described IL-15 is meant a kind of cell strain CTLL-2 of the IL-15 of dependence growth.The engineering cell strain that is obtained just can carry out clinical application through conventional amplification in vitro.
Preservation information about the NK cell strain of IL-15 genetic modification: at present, this cell strain has been submitted China Committee for Culture Collection of Microorganisms's common micro-organisms center preservation to, its preservation accession designation number is CGMCC No.0996, the classification name is people's natural killer cell system, and storage life is 30 years that 2003.8.27 rises.
3. beneficial effect
One aspect of the present invention provides a kind of NK cell strain (NK-92/IL-15) of new IL-15 genetic modification, compares with unmodified NK-92 cell, and this cell strain can stably express IL-15; Can under the IL-15 of low dosage culture condition, have high-caliber multiplication capacity, help large scale culturing; Under low dosage IL-15 culture condition, the kill capability of the tumour cell in multiple source is significantly improved, improve service efficiency, reaching under the prerequisite of equal antitumous effect, reduce the dosage of NK cell, thereby reduce cost, be more suitable for the clinical adoptive immunotherapy tumour that is used for.
Four. description of drawings
Fig. 1 is the electrophoretogram of the PCR product of human IL-15 cDNA, and wherein swimming lane 1 is the PCR product of IL-15, and swimming lane 2 is DNA Marker;
Fig. 2 is a pcDNA3/IL-15 construction of recombinant plasmid schema;
Fig. 3 is a pcDNA3/IL-15 stable transfection NK-92 cell synoptic diagram;
Fig. 4 is that RT-PCR detects NK cell IL-15mRNA transcriptional level, and wherein swimming lane 1 is DNA Marker, and swimming lane 2 is an IL-15mRNA level in the NK-92/IL-15 cell, and swimming lane 3 is an IL-15mRNA level in the NK-92 cell;
Fig. 5 is the comparison of cellular form before and after the genetic modification, and wherein A is NK-92 cellular form before modifying, and B is for modifying back NK-92/IL-15 cellular form;
Fig. 6 is the comparison of NK-92/IL-15 multiplication effect and NK-92 cell, and wherein A is a cell increment trend under the 100U/ml IL-15 culture condition, and B is a 25U/ml IL-15 culture condition cell increment trend;
Fig. 7 is that NK-92/IL-15 is to the comparison of tumor cytotoxicity activity in multiple source with the NK-92 cell, wherein A is the killing activity to colorectal carcinoma HT-29 cell, B is the killing activity that height is shifted lung cancer PG5 cell, C is the killing activity to laryngeal cancer cell Hep2, D is the killing activity to cervical cancer cell Hela, and E is the killing activity to ovarian cancer cell 3AO;
Fig. 8 is the comparison of relevant murderer transcriptional level of NK-92/IL-15 cell and NK-92 cell;
Fig. 9 is the comparison of NK-92/IL-15 cell CD25 developed by molecule level and NK-92 cell;
Figure 10 is the comparison of NK-92/IL-15 cell CD48 developed by molecule level and NK-92 cell;
Figure 11 is the comparison of NK-92/IL-15 cell CD69 developed by molecule level and NK-92 cell;
Figure 12 is the comparison of NK-92/IL-15 cell CD54 developed by molecule level and NK-92 cell.
Five. embodiment
Embodiment
One, pcDNA3/IL-15 construction of recombinant plasmid
Stimulating the back to extract total RNA peripheral blood mononuclear cell is that template is carried out RT-polymerase chain reaction (RT-PCR), the cDNA fragment of amplification human IL-15, expectation length is 486bp (Fig. 1), wherein the RT parameter is: 37 ℃ 1 hour, 95 ℃ 10 minutes; The PCR parameter is: 94 ℃ 1 minute, 58 ℃ 1 minute, 72 ℃ 1 minute, circulate 30 times, last prolongs 7 minutes.PcDNA3/IL-15 construction of recombinant plasmid flow process (Fig. 2): at first, the RT-PCR product is connected with the T-carrier, product changes the competence bacillus coli DH 5 alpha over to, screening positive clone, entrust Shanghai Bo Ya bio-engineering corporation to carry out determined dna sequence, the result shows that the IL-15cDNA sequence is in full accord with report.Then, with EcoR I and BamH I double digestion T-IL-15, electrophoresis purifying IL-15 fragment is connected with the pcDNA3 that same enzyme is cut, and product changes the competence bacillus coli DH 5 alpha over to, the screening positive recombinant.
Two, pcDNA3/IL-15 stable transfection NK-92 cell
PcDNA3/IL-15 stable transfection NK-92 cell and screening (Fig. 3): the NK-92 cell cultures is in the α that contains 10% calf serum, 12.5% horse serum, 100U IL-2-MEM substratum, go down to posterity and grow to exponential phase of growth, get 2-3ugpcDNA3/IL-15 liposome LIPOFECTAMINE
TMTransfection NK-92 cell, 37 ℃ of 5%CO2 were cultivated after 48 hours, (contained the α that final concentration is 200mg/L G418-MEM) by importing screening culture medium at 1: 4 into.Cultivated 7 days, and be changed to the complete α-MEM substratum that contains final concentration 600mg/L G418, continue to cultivate about 10 days.Get a part of cell extraction RNA and carry out RT-PCR detection IL-15mRNA transcriptional level, another part cell dilution is that 10/ml carries out the limited dilution cloning cell, cultivated about 10 days, thereby get supernatant and pick out positive cell clone, obtain to have the cell strain of clinical value with CTLL-2 cell detection IL-15 activity.
Three, the qualification process of the positive cell clone of IL-15 genetic modification:
After pcDNA3/IL-15 was with liposome method stable transfection NK-92 cell, the at first complete α through containing G418-MEM substratum screening was got a part of cell extraction RNA then and is carried out RT-PCR detection IL-15 transcriptional level; Another part cell limiting dilution assay clone cell, cultivate about 10 days after, take out the 100ul supernatant and be used for the IL-15 determination of activity.
1.CTLL-2 the activity of IL-15 in the raji cell assay Raji cell clone culture supernatant
Get these 6 extent of dilution of sesquialter dilution in 96 porocyte culture plates of rhIL-15 standard substance, rhIL-2 and institute's test sample, 2 multiple holes of each extent of dilution, and to establish simple RPMI1640 liquid be blank.Abandon supernatant after getting the CTLL-2 cell centrifugation, physiological saline washing 3 times transfers to 1 * 10 with the RPMI1640 nutrient solution
5/ ml adds the 100ul/ hole respectively.37 ℃, 5%CO
2Cultivate 24h under the condition.After mirror was observed the whole death of blank porocyte down, every hole added MTT, continued to cultivate 4h.The 100ul culture supernatant is abandoned in every hole, adds 10%SDS solution 100ul/ hole, and 37 ℃ are spent the night, and measure 570nm OD value.The employing probit method calculates the biological respinse rate of each sample, finally calculates the biological value (IU/ml) of sample IL-15.
2.mRNA transcription analysis
NK cell after the G418 screening, guanidinium isothiocyanate method extract RNA and carry out the RT-PCR analysis, have the mRNA of IL-15 to transcribe (Fig. 4).
Four, the biological characteristics of IL-15 genetic modification NK cell
1.NK-92/IL-15 multiplication capacity analysis
Cell counting is observed the time-effect relationship (Fig. 6) of NK cell proliferation: add 10 in 24 orifice plates
4Cell/ml and the substratum that contains 100U/ml or 25U/ml rhIL-15, per two days countings once and are partly measured and are changed liquid, cultured continuously 10 days.NK cell behind the IL-15 genetic modification has rate of propagation faster.Perhaps the horse serum concentration in the substratum is reduced to 8%, also can obtains same result.
2.NK-92/IL-15 to the active analysis of tumor cytotoxicity
Measure the killing activity (Fig. 7) of NK cell with mtt assay: be the effector cell with NK-92 and NK-92/IL-15 cell respectively, with 3AO, PG5, Hen2, Hela, when HT-29 is target cell, at first target cell is used trysinization, transfer to 5 * 10 after the washing
4/ ml adds the 100ul/ hole in 96 orifice plates, cultivate 24hr, adds the effector cell according to 10: 1,5: 1,2.5: 1,1.25: 1,0.625: 1 then, and each effect target ratio is established 4 multiple holes, and every hole cumulative volume is 200ul, and establishes perfect medium and do the zeroing hole.37 ℃, 5%CO2 overnight incubation continue to cultivate 4h behind the adding MTT, discard supernatant, add 100ul 5%SDS, and 37 ℃, 5%CO2 overnight incubation are measured OD570 then, calculate the NK cell killing activity according to following formula:
3.NK cell killing associated molecule transcriptional level
Detect the transcriptional level (Fig. 8) of NK cell killing associated molecule before and after the genetic modification with the RT-PCR method: collecting cell, guanidine isothiocyanate method extract RNA and carry out RT-PCR, with β-actin as system's internal reference.
4. the mensuration of cell phenotype
Flow cytometer detects NK cell surface molecule (Fig. 9-13): collecting cell, the PBA washed twice, add 0.25%BSA sealing 30min, the PBA washed twice, add monoclonal antibody (being contrast with the homotype monoclonal antibody simultaneously), ice bath 30min, PBA washed twice, last machine analysis or fix with the formaldehyde solution of 1-4%.Used antibody is the product of U.S. BD-Pharmingen company, and detected result shows that the phenotype of NK cell is CD56 strong positive (average fluorescent strength is 684.3), and CD16 and CD3 are all negative.In addition, the expression rate of the necessary adhesion molecule CD54 of NK cell killing reaches 99.6%, and average fluorescent strength reaches 220; The expression rate of the surface of cell membrane molecule CD25 relevant with NK cell proliferation reaches 77%; The expression rate of the surface of cell membrane molecule CD69 relevant with the NK cell activation reaches 64.25%; The average fluorescent strength of the surface of cell membrane molecule CD48 relevant with the NK cell activation reaches 435.9.
Claims (7)
1, the natural killer cell strain of Interleukin-15 genetic modification, its phenotype is the CD56 strong positive, CD16 and CD3 are all negative, it is characterized in that form is dispersive, half adherent growth state.
2, the natural killer cell strain of Interleukin-15 genetic modification according to claim 1 is characterized in that the expression rate of the necessary adhesion molecule CD54 of NK cell killing reaches 90～100%, and average fluorescent strength reaches 220.
3, the natural killer cell strain of Interleukin-15 genetic modification according to claim 1 is characterized in that the expression rate of the surface of cell membrane molecule CD25 relevant with NK cell proliferation reaches 50～80%.
4, the natural killer cell strain of Interleukin-15 genetic modification according to claim 1 is characterized in that the expression rate of the surface of cell membrane molecule CD69 relevant with the NK cell activation reaches 15～65%.
5, the natural killer cell strain of Interleukin-15 genetic modification according to claim 1 is characterized in that the average fluorescent strength of the surface of cell membrane molecule CD48 relevant with the NK cell activation reaches 435.
6, the preparation method of the natural killer cell strain of Interleukin-15 genetic modification is characterized in that the pcDNA3 carrier for expression of eukaryon is inserted in the cDNA coding region of IL-15, makes up secretor type recombinant eukaryon expression vector pcDNA3/IL-15; Utilize liposome transfection NK-92 cell; Adding Geneticin (G418) in screening culture medium screens; The limited dilution cloning cell, the activity with IL-15 in the dependent cells strain mensuration clone cell culture supernatant of IL-15 selects the positive cell clone of expressing IL-15; This is cloned in the complete α-MEM substratum of IL-15 of 20～100 activity units increases, obtain having the engineering cell strain of clinical value.
7, the preparation method of the natural killer cell strain of Interleukin-15 genetic modification according to claim 6, it is characterized in that it being this to be cloned in α-MEM substratum of the IL-15 that contains 10% calf serum, 8～12.5% horse serums, 20～100 activity units increase, obtain having the engineering cell strain of clinical value.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB031529682A CN1233822C (en) | 2003-09-05 | 2003-09-05 | Interleukin-15 gene modified natural killing cell strain and its preparation method |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB031529682A CN1233822C (en) | 2003-09-05 | 2003-09-05 | Interleukin-15 gene modified natural killing cell strain and its preparation method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1493687A true CN1493687A (en) | 2004-05-05 |
| CN1233822C CN1233822C (en) | 2005-12-28 |
Family
ID=34240741
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB031529682A Expired - Lifetime CN1233822C (en) | 2003-09-05 | 2003-09-05 | Interleukin-15 gene modified natural killing cell strain and its preparation method |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN1233822C (en) |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101878034B (en) * | 2007-09-28 | 2013-11-20 | 细胞基因细胞疗法公司 | Tumor suppression using human placental perfusate and human placenta-derived intermediate natural killer cells |
| CN105017429A (en) * | 2010-09-21 | 2015-11-04 | 阿尔托生物科学有限公司 | Multimeric IL-15 soluble fusion molecules and methods of making and using same |
| CN108531458A (en) * | 2018-04-27 | 2018-09-14 | 赛诺（深圳）生物医药研究有限公司 | Treat the genetic engineering natural killer cells product of tumour |
| CN109207430A (en) * | 2018-09-25 | 2019-01-15 | 华东师范大学 | A kind of Chimeric antigen receptor NK cell and its preparation method and application |
| CN110863000A (en) * | 2019-11-22 | 2020-03-06 | 北京鼎成肽源生物技术有限公司 | Gene and transfection vector for transfecting NK cell |
| US11053299B2 (en) | 2010-09-21 | 2021-07-06 | Immunity Bio, Inc. | Superkine |
| US11173191B2 (en) | 2014-06-30 | 2021-11-16 | Altor BioScience, LLC. | IL-15-based molecules and methods of use thereof |
| US11318201B2 (en) | 2016-10-21 | 2022-05-03 | Altor BioScience, LLC. | Multimeric IL-15-based molecules |
| US11365231B2 (en) | 2007-05-11 | 2022-06-21 | Altor Bioscience, Llc | Interleukin 15 (IL-15) variants |
-
2003
- 2003-09-05 CN CNB031529682A patent/CN1233822C/en not_active Expired - Lifetime
Cited By (20)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US11673932B2 (en) | 2007-05-11 | 2023-06-13 | Altor BioScience, LLC. | Fusion molecules and IL-15 variants |
| US11498950B1 (en) | 2007-05-11 | 2022-11-15 | Altor Bioscience, Llc | Fusion molecules and IL-15 variants |
| US11365231B2 (en) | 2007-05-11 | 2022-06-21 | Altor Bioscience, Llc | Interleukin 15 (IL-15) variants |
| CN101878034B (en) * | 2007-09-28 | 2013-11-20 | 细胞基因细胞疗法公司 | Tumor suppression using human placental perfusate and human placenta-derived intermediate natural killer cells |
| CN105017429A (en) * | 2010-09-21 | 2015-11-04 | 阿尔托生物科学有限公司 | Multimeric IL-15 soluble fusion molecules and methods of making and using same |
| US11845783B2 (en) | 2010-09-21 | 2023-12-19 | Altor BioScience, LLC. | Multimeric IL-15 soluble fusion molecules and methods of making and using same |
| US11046747B2 (en) | 2010-09-21 | 2021-06-29 | Altor Bioscience Llc | Multimeric IL-15 soluble fusion molecules and methods of making and using same |
| US11053299B2 (en) | 2010-09-21 | 2021-07-06 | Immunity Bio, Inc. | Superkine |
| US11104716B2 (en) | 2010-09-21 | 2021-08-31 | Altor BioScience, LLC. | Multimeric IL-15 soluble fusion molecules and methods of making and using same |
| US11890323B2 (en) | 2014-06-30 | 2024-02-06 | Altor Bioscience, Llc | Method of treating cancer with composition comprising IL-15-based molecules and BCG |
| US11173191B2 (en) | 2014-06-30 | 2021-11-16 | Altor BioScience, LLC. | IL-15-based molecules and methods of use thereof |
| US11679144B2 (en) | 2014-06-30 | 2023-06-20 | Altor BioScience, LLC. | IL-15-based molecules and methods of use thereof |
| US11925676B2 (en) | 2014-06-30 | 2024-03-12 | Altor BioScience, LLC. | Method for treating neoplasia with an anti-CD38 antibody and an IL-15:IL-15R complex |
| US11471511B2 (en) | 2014-06-30 | 2022-10-18 | Altor Bioscience, Llc | IL-15-based molecules and methods of use thereof |
| US11318201B2 (en) | 2016-10-21 | 2022-05-03 | Altor BioScience, LLC. | Multimeric IL-15-based molecules |
| US11369679B2 (en) | 2016-10-21 | 2022-06-28 | Altor Bioscience, Llc | Multimeric IL-15-based molecules |
| WO2019205403A1 (en) * | 2018-04-27 | 2019-10-31 | 赛诺（深圳）生物医药研究有限公司 | Genetically engineered natural killer cell product for treating tumor |
| CN108531458A (en) * | 2018-04-27 | 2018-09-14 | 赛诺（深圳）生物医药研究有限公司 | Treat the genetic engineering natural killer cells product of tumour |
| CN109207430A (en) * | 2018-09-25 | 2019-01-15 | 华东师范大学 | A kind of Chimeric antigen receptor NK cell and its preparation method and application |
| CN110863000A (en) * | 2019-11-22 | 2020-03-06 | 北京鼎成肽源生物技术有限公司 | Gene and transfection vector for transfecting NK cell |
Also Published As
| Publication number | Publication date |
|---|---|
| CN1233822C (en) | 2005-12-28 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Seiffert et al. | Soluble CD14 is a novel monocyte-derived survival factor for chronic lymphocytic leukemia cells, which is induced by CLL cells in vitro and present at abnormally high levels in vivo | |
| CN101519646B (en) | CIK cell, as well as preparation method and cell preparation thereof | |
| CN102816735B (en) | Method for culturing autologous peripheral blood lymphocytes | |
| CN1233822C (en) | Interleukin-15 gene modified natural killing cell strain and its preparation method | |
| KR20100012586A (en) | Method for proliferating natural killer cell | |
| CN115651903B (en) | High-lethality immune cell population, and culture method, reagent composition and application thereof | |
| CN115558641B (en) | High-purity effector immune cell population, culture method, reagent composition and application thereof | |
| CN102027104A (en) | Method for production of cell mass containing cytokine-induced killer cell | |
| CN115466726B (en) | NK cell efficient gene transduction scheme | |
| CN102766654A (en) | Slow virus expression vector specially promoting high expression of liver cell CYP1A2 gene, and construction method and application thereof | |
| CN114250198A (en) | Method for enhancing anti-tumor effect of immune cells | |
| CN116179596A (en) | Long-acting recombinant human interleukin 2 fusion protein and preparation method and application thereof | |
| CN101988049A (en) | Genetically modified cell of coexpression mouse membranous type interleukin 15 and retinoic acid early transcript 1 epsilon (Rae-1 epsilon) and preparation method thereof | |
| CN111826354B (en) | NK cell and application thereof in tumor treatment | |
| CN1244696C (en) | Highly effective expressing method of interleukin-12 | |
| CN1656214A (en) | Cytokine-free growth and maintenance of progenitor cells | |
| CN103194428A (en) | Culture method for in vitro differentiating and amplifying human natural killer cells through insulin-like growth factors and various cell factors | |
| CN109294988B (en) | NK cell induction kit | |
| CN104962518A (en) | New application of CD44 plasmid | |
| Kern et al. | Eosinophil differentiating activity in sera of patients with AIDS under recombinant IL-2 substitution | |
| CN110904048A (en) | CAR-CD123T2 chimeric antigen receptor T cell and application thereof | |
| Luo et al. | In Vitro Generation of Human Cross-Presenting Type 1 Conventional Dendritic Cells (cDC1s) and Plasmacytoid Dendritic Cells (pDCs) | |
| CN105861484B (en) | Cell culture medium composition containing resveratrol and silk sericin | |
| CN112300992A (en) | NK cell culture solution and multistage activation NK cell culture method | |
| CN105624109A (en) | Method for culturing NKT cells by using bioreactor |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CX01 | Expiry of patent term | ||
| CX01 | Expiry of patent term |
Granted publication date: 20051228 |