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1.5 ml microtubes

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Company: SARSTEDT
Catalog#: 72.690.001
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Direct Reprogramming of Mouse Embryonic Fibroblasts to Conventional Type 1 Dendritic Cells by Enforced Expression of Transcription Factors
Author:
Date:
2020-05-20
[Abstract]  Ectopic expression of transcription factor combinations has been recently demonstrated to reprogram differentiated somatic cells towards the dendritic cell (DC) lineage without reversion to a multipotent state. DCs have the ability to induce potent and long-lasting adaptive immune responses. In particular, conventional type 1 DCs (cDC1s) excel on antigen cross-presentation, a critical step for inducing CD8+ T cell cytotoxic responses. The rarity of naturally occurring cDC1s and lack of in vitro methodologies for the generation of pure cDC1 populations strongly hinders the study of cDC1 lineage specification and function. Here, we describe a protocol for the generation of induced DCs (iDCs) by lentiviral-mediated expression of the transcription factors PU.1, IRF8 and ... [摘要]  [摘要] 转录因子组合的异位表达最近被证明可以将分化的体细胞重编程为树突状细胞(DC)谱系,而不会回复到多能状态。DC具有诱导有效和持久的适应性免疫应答的能力。在特定的常规类型1的DC(cDC1s)练成抗原交叉呈递,用于诱导CD8的关键步骤+ T细胞的细胞毒性应答。天然存在的cDC1的稀有性和缺乏用于生成纯cDC1群体的体外方法论,严重阻碍了对cDC1谱系规格和功能的研究。在这里,我们描述了用于生成感应DC(iDC)的协议 慢病毒介导的转录因子PU.1,IRF8和BATF3在小鼠胚胎成纤维细胞中的表达。iDC 在9天内获得DC形态,cDC1表型和转录特征。使用此协议生成的iDC 具有对炎症刺激,吞噬死细胞,将抗原加工并交叉呈递给CD8 + T细胞的功能。DC重新编程提供了一个简单易处理的系统,可以生成大量的cDC1类细胞用于高内涵筛选,从而开辟了新途径,可以更好地了解cDC1的规格和功能。将来,在成纤维细胞中忠实诱导cDC1命运可能会导致产生患者特定的疫苗接种DC。

[背景技术树突状细胞(DC)是专业的抗原呈递细胞,专门用于识别,加工和呈递T细胞抗原,在诱导适应性免疫应答和免疫记忆中起关键作用(Me rad 等,2013)。DC可以分为4个主要子集:浆细胞样DC(pDC ),大量1型干扰素的产生者,循环单核细胞衍生的单核细胞衍生DC 和常规DC(cDC ...

Confocal and Super-resolution Imaging of RNA in Live Bacteria Using a Fluorogenic Silicon Rhodamine-binding Aptamer
Author:
Date:
2020-05-05
[Abstract]  Genetically encoded light-up RNA aptamers have been shown to be promising tools for the visualization of RNAs in living cells, helping us to advance our understanding of the broad and complex life of RNA. Although a handful of light-up aptamers spanning the visible wavelength region have been developed, none of them have yet been reported to be compatible with advanced super-resolution techniques, mainly due to poor photophysical properties of their small-molecule fluorogens. Here, we describe a detailed protocol for fluorescence microscopy of mRNA in live bacteria using the recently reported fluorogenic silicon rhodamine binding aptamer (SiRA) featuring excellent photophysical properties. Notably, with SiRA, we demonstrated the first aptamer-based RNA visualization using super-resolution ... [摘要]  [摘要 ] 遗传编码的点亮适体是显示活细胞中RNA的有前途的工具,可帮助我们加深对RNA广泛而复杂的生命的理解。可见光波长区已经被开发,他们都没有然而,据报道,在兼容先进的超分辨率技术,主要是由于不良的光物理性质其小分子荧光团。在这里,我们描述了一个详细的协议对于荧光显微镜mRNA的使用最近报道的具有优异光物理性质的荧光罗丹明结合适体(SiRA )在活细菌中进行检测。值得注意的是,我们利用SiRA 展示了首个使用超分辨率(STED)显微镜进行的基于适体的RNA可视化。这种成像方法可能特别有价值用于可视化原核生物中的RNA,因为细菌的大小仅比光学分辨率大几倍 传统显微镜的分辨率。

[背景 ] 可视化的具体RNA分子通过荧光显微镜具有不可估量的价值在过去二十年中扩大我们的知识RNA功能内的细胞在时空精气神(特亚吉,2009年;夏等人,2017年),由于缺乏。固有的荧光RNA,用于活细胞成像的荧光RNA标记工具的开发以及它们对最新显微镜的适应性 –特别是对于超分辨率显微镜– 势在必行。超分辨率显微镜(SRM)对于原核系统中的RNA成像特别有吸引力,因为细菌很小(〜2.5MYU中号长,0.5-1〜MYU 中号宽)和分辨率的标准荧光显微镜被限制在200〜300〜牛米,由于衍射极限光(Reshes ...

Optogenetic Tuning of Ligand Binding to The Human T cell Receptor Using The opto-ligand-TCR System
Author:
Date:
2020-03-05
[Abstract]  T cells are one major cell type of the immune system that use their T cell antigen receptor (TCR) to bind and respond to foreign molecules derived from pathogens. The ligand-TCR interaction half-lives determine stimulation outcome. Until recently, scientists relied on mutating either the TCR or its ligands to investigate how varying TCR-ligand interaction durations impacted on T cell activation. Our newly created opto-ligand-TCR system allowed us to precisely and reversibly control ligand binding to the TCR by light illumination. This system uses phytochrome B (PhyB) tetramers as a light-regulated TCR ligand. PhyB can be photoconverted between a binding (ON) and non-binding (OFF) conformation by 660 nm and 740 nm light illumination, respectively. PhyB ON is able to bind to a synthetic ... [摘要]  [摘要] T细胞是免疫系统的一种主要细胞类型,利用其T细胞抗原受体(TCR)结合并响应源自病原体的外来分子。配体-TCR相互作用的半衰期决定了刺激的结果。直到最近,科学家还是依靠突变TCR或其配体来研究变化的TCR-配体相互作用持续时间如何影响T细胞活化。我们新创建的光配体-TCR系统使我们能够通过光照精确且可逆地控制配体与TCR的结合。该系统使用植物色素B(PhyB )四聚体作为光调节的TCR配体。PHYB 可光转化的结合(ON)之间,并且通过分别为660nm和740nm的光照射,非结合性(OFF)构象。PhyB ON能够结合通过将PhyB 相互作用因子(PIF)融合到TCRβ链而产生的合成TCR 。将PhyB 切换为OFF构象会破坏这种相互作用。PhyB 四聚体与PIF-TCR 足够长的结合导致T细胞活化(通过钙内流测量)。在这里,我们描述了有关如何为我们的光配体-TCR系统生成四聚体配体,如何通过流式细胞仪测量配体-TCR结合以及如何通过钙内流量化T细胞活化的协议。

[背景 ] ...

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