ImmunoCult™树突状细胞成熟添加物

用于人单核细胞来源树突状细胞成熟的添加物

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产品号 #10989_C

用于人单核细胞来源树突状细胞成熟的添加物

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使用ImmunoCult™树突状细胞成熟添加物，将未成熟树突状细胞（DC）转化为成熟DC。

该产品作为ImmunoCult™树突状细胞培养试剂盒的组分，也可单独购买，以方便您的使用。ImmunoCult™树突状细胞成熟添加物专为搭配ImmunoCult™-ACF树突状细胞培养基和ImmunoCult™-ACF树突状细胞分化添加物使用而设计，用以支持未成熟DC的成熟过程。

关于使用ImmunoCult™树突状细胞成熟添加物进行DC成熟的详细方法，请参阅产品说明书（PIS）。

实验数据

Figure 1. Protocol Diagram.

Mature DCs were generated by culturing EasySep™ isolated monocytes at 1 x 10⁶ cells/mL in ImmunoCult™-ACF Dendritic Cell Medium (Catalog #10987) with added ImmunoCult™-ACF Dendritic Cell Differentiation Supplement (Catalog #10988). At day 3, the medium with differentiation supplement was replaced and cells were incubated for 2 more days. At day 5, without changing the medium, ImmunoCult™ Dendritic Cell Maturation Supplement (Catalog #10989) was added to the culture. At day 7, fully mature DCs were harvested for downstream applications.

Figure 2. Mature DCs generated with ImmunoCult™-ACF Dendritic Cell Medium with Supplements show desired phenotype.

EasySep™ isolated monocytes were cultured and differentiated into mature DCs as described in Figure 1. (A) The percentage of CD14 and CD83 expression in cells at day 7 (mature DCs) was determined by flow cytometry. At day 7, a total of 93 ± 5% of the cells expressed the mature DC marker CD83 and only 1 ± 1% of cells still expressed the monocyte marker CD14 (mean ± SD, n=39). Yield of mature DCs was determined by count of total viable cells at day 7 relative to the count of viable monocytes used for initial culture at day 0. At day 7, the yield of viable mature DCs corresponded to 45 ± 25% (mean ± SD, n=39). (B) Immature DCs were cultured as described in Figure 1. At day 5, cells were cultured with maturation supplement for 2 days (mature DCs) or without maturation supplement (immature DCs). Supernatant was collected at day 7 and IL-12p70 levels were determined by ELISA. Concentrations of IL-12p70 in supernatant of mature and immature DCs were 361 ± 81 and 5 ± 2 pg/mL, respectively (mean ± SEM, n=27).

Figure 3. Mature DCs generated with ImmunoCult™-ACF Dendritic Cell Medium and Supplements induce T cell proliferation.

Mature DCs generated with ImmunoCult™-ACF Dendritic Cell Medium and Supplements (ImmunoCult) or other serum-free competitor media (competitor 1 and 2) and corresponding supplements when applicable (competitor 2), were cultured in ImmunoCult™-XF T Cell Expansion Medium with 1 x 10⁵ CFSE labeled (A) allogeneic CD3+ T cells (MLR assay) or (B) autologous CD8+ T cells (antigen-specific T cell response). (A) Cells were cultured at a DC:T cell ratio of 1:25. (B) Prior to culture with T cells, immature DCs were loaded with HLA Class I peptides derived from the human Cytomegalovirus, Epstein-Barr Virus and Influenza Virus (CEF peptide pool) and stimulated with maturation supplement for 2 days. Cells were cultured at a DC:T cell ratio of 1:4 or 1:10. (A,B) CFSE labeled T cells were incubated in media alone (negative control) or with ImmunoCult™ Human CD3/CD28 T Cell Activator (positive control). After 5-7 days in culture the number of dividing T cells ( CD3⁺CFSE^lo) was assessed by flow cytometry (mean ± SEM) (A) n=5 (B) n=4 (competitor 1 and 2, n=3). Mature DCs generated in ImmunoCult™-ACF Dendritic Cell Medium induced proliferation of allogeneic and antigen-specific T cells similar to DCs generated in either competitor media. Competitors 1 and 2, include in no particular order, CellGro DC Medium (CellGenix) and PromoCell DC Generation Medium DXF.

产品说明书及文档

请在《产品说明书》中查找相关支持信息和使用说明，或浏览下方更多实验方案。

文献 (1)

DNA of neutrophil extracellular traps promote NF-κB-dependent autoimmunity via cGAS/TLR9 in chronic obstructive pulmonary disease J. Chen et al. Signal Transduction and Targeted Therapy 2024 Jun

Abstract

Chronic obstructive pulmonary disease (COPD) is characterised by persistent airway inflammation even after cigarette smoking cessation. Neutrophil extracellular traps (NETs) have been implicated in COPD severity and acute airway inflammation induced by short-term cigarette smoke (CS). However,whether and how NETs contribute to sustained airway inflammation in COPD remain unclear. This study aimed to elucidate the immunoregulatory mechanism of NETs in COPD,employing human neutrophils,airway epithelial cells (AECs),dendritic cells (DCs),and a long-term CS-induced COPD mouse model,alongside cyclic guanosine monophosphate-adenosine monophosphate synthase and toll-like receptor 9 knockout mice ( cGAS -−/−,TLR9 −/− ); Additionally,bronchoalveolar lavage fluid (BALF) of COPD patients was examined. Neutrophils from COPD patients released greater cigarette smoke extract (CSE)-induced NETs (CSE-NETs) due to mitochondrial respiratory chain dysfunction. These CSE-NETs,containing oxidatively-damaged DNA (NETs-DNA),promoted AECs proliferation,nuclear factor kappa B (NF-κB) activation,NF-κB-dependent cytokines and type-I interferons production,and DC maturation,which were ameliorated/reversed by silencing/inhibition of cGAS/TLR9. In the COPD mouse model,blocking NETs-DNA-sensing via cGAS − /− and TLR9 − /− mice,inhibiting NETosis using mitoTEMPO,and degrading NETs-DNA with DNase-I,respectively,reduced NETs infiltrations,airway inflammation,NF-κB activation and NF-κB-dependent cytokines,but not type-I interferons due to IFN-α/β receptor degradation. Elevated NETs components (myeloperoxidase and neutrophil elastase activity) in BALF of COPD smokers correlated with disease severity and NF-κB-dependent cytokine levels,but not type-I interferon levels. In conclusion,NETs-DNA promotes NF-κB-dependent autoimmunity via cGAS/TLR9 in long-term CS exposure-induced COPD. Therefore,targeting NETs-DNA and cGAS/TLR9 emerges as a potential strategy to alleviate persistent airway inflammation in COPD. Subject terms: Inflammation,Respiratory tract diseases

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物种	人
配方	不含动物成分, 无血清

ImmunoCult™树突状细胞成熟添加物

产品号 #10989

产品号 #（选择产品）

产品号 #10989_C

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ImmunoCult™树突状细胞成熟添加物

产品号 #10989

产品号 #（选择产品）

产品号 #10989_C

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实验数据

产品说明书及文档

应用领域

相关材料与文献

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