树突细胞根据其独特形态而得名,是一组分布广泛、由骨髓来源的、具有迁移能力的免疫细胞,是目前功能最强大的抗原递呈细胞(APC),也是唯一能活化静息型T细胞(Naïve T cells)的APC,引起T细胞初级免疫应答。
1、DC树突状细胞的分化发育Overview
2、Pathways of human dendritic cell (DC) development
3、Regulation of DC development and homeostasis in mice
4、DC controllers of adaptive immunity DC与获得性免疫之间的关系
1、Phenotype of human DC subsets
2、Phenotype of mouse lymphoid and nonlymphoid tissue DC
1、如何获取DC细胞?
(1)使用CD34、CD133磁珠分选得到高纯度造血干细胞,体外培养,诱导成DC细胞;
(2)磁珠分选单核细胞(CD14+)诱导成DC细胞;
(3)使用BDCA1、4、3直接分选DC细胞 ( BDCA-1(CD1c) DC Iso kit; CD304( BDCA-2) MicroBeads, kit; CD141(BDCA-3) MicroBeads, kit);
所需产品:
分选PDC用磁珠
130-090-509
CD303 (BDCA-2) MicroBead Kit, human
Kit
130-097-240
Diamond Plasmacytoid Dendritic Cell Isolation Kit II, human
Kit
分选MDC1用磁珠
130-119-475
CD1c (BDCA-1)+ Dendritic Cell Isolation Kit, human
Kit
分选MDC2用磁珠
130-090-512
CD141 (BDCA-3) MicroBead Kit, human
Kit
分选单核细胞磁珠
130-050-201
CD14 MicroBeads, human
2mL
分选造血干细胞磁珠
130-100-453
CD34 MicroBead Kit UltraPure, human
2mL
130-094-913
Diamond CD133 Isolation Kit, human
2mL
Mo-DC诱导用细胞因子
130-093-919
Human IL-4, premium grade
5μg
130-093-864
Human GM-CSF, premium grade
10μg
130-093-897
Human IL-1β, premium grade
10μg
130-094-015
Human TNF-α, research grade
10μg
Human IL-6, research grade
10μg
130-096-481
Human IFN-γ1b, premium grade
10μg
2、Mo-DC Workflow
Day 0
分离出的单核细胞(3×108 cells)用3 mL的完全培养基(RPMI 1640, 2 mM L-glutamine, 1% autologous plasma),添加250 IU/mL IL-4 、800 IU/mL GM-CSF进行培养。
Day 2
从培养体系吸出1.5 mL,离心。用1.5 mL完全培养基加2倍浓度的IL-4和 GM-CSF,再添加到原培养体系中
Day 6
从培养体系吸出1.5 mL,离心。用1.5 mL培养基(添加2000 IU/mL IL-6, 400 IU/mL IL-1β, 2000 IU/mL TNF-α, 2 µg/mL PGE2),再添加到原培养体系中,培养24小时。
Day 7
通过流式检测成熟Mo-DCs (mMo-DCs)的活性为95.4±2.4%,得率为23%。
3、MLR混合淋巴细胞反应
4、DC细胞亚群分析
Panel一:人DC细胞亚群鉴定:
4-Color Assay;
R3: lin-HLA-DR-CD123+嗜碱性粒细胞;
R4:lin-HLA-DR+CD123+淋巴系细胞DC(DC2);
R5:lin-HLA-DR+CD11C+髓系DC(DC1)
Panel二:小鼠DC细胞亚群鉴定:
reference :
1:Alexander M. Steffen J(2014)Development and Function of Dendritic Cell Subsets.Immunity Review
2: Barbara U. Caetano R.(2015)Defining dendritic cells Current Opinion in Immunology 2015, 32:13–20
3:Miriam M. Priyanka S. Julie H. Jennifer M. and Arthur M.(2013)The Dendritic Cell Lineage: Ontogeny and Function of Dendritic Cells and Their Subsets in the Steady State and the Inflamed Setting Annu Rev Immunol. 2013 ; 31