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16% Paraformaldehyde (formaldehyde) aqueous solution

多聚甲醛(PFA)

公司名称: Electron Microscopy Sciences
产品编号: 15710
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FM1-43 Photoconversion and Electron Microscopy Analysis at the Drosophila Neuromuscular Junction
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Date:
2017-09-05
[Abstract]  We developed a protocol for photoconversion of endocytic marker FM1-43 followed by electron microscopy analysis of synaptic boutons at the Drosophila neuromuscular junction. This protocol allows detection of stained synaptic vesicle even when release rates are very low, such as during the spontaneous release mode. The preparations are loaded with the FM1-43 dye, pre-fixed, treated and illuminated to photoconvert the dye, and then processed for conventional electron microscopy. This procedure enables clear identification of stained synaptic vesicles at electron micrographs. [摘要]  我们开发了内吞标记FM1-43的光转换方案,然后在果蝇神经肌肉接头处进行突触引物的电子显微镜分析。 即使在释放速率非常低时,例如在自发释放模式期间,该方案允许检测染色的突触小泡。 该制剂装载有FM1-43染料,经预先固定,处理和照射,以使染料转变为染料,然后进行常规电子显微镜处理。 该方法能够在电子显微照片下清楚鉴定染色的突触小泡。
【背景】神经元发射体通过突触小泡与神经元质膜的融合而释放。囊泡可以自发融合或响应动作电位。随后,囊泡通过内吞作用获得回收。通过分子生物学,电生理学和显微镜的工具广泛研究了突触小泡回收的分子机制(Slepnev和De Camilli,2000; Sudhof,2004; Rizzoli和Betz,2005; Kavalali,2006)。加载内参标记FM1-43与染料光转换耦合,然后进行电子显微镜分析是一种强大的技术,允许调查和测量回收囊泡池(Harata et al。,2001; Schikorski and Stevens,2001; Rizzoli和Betz, 2004)。果蝇神经肌肉接头(NMJ)是具有明确定义的突触引物的有利制剂,其能够快速产生具有突变突触蛋白的细胞系和严格评估囊泡回收池(Akbergenova和Bykhovskaia,2009; ...

MHC Class II Tetramer Labeling of Human Primary CD4+ T Cells from HIV Infected Patients
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Date:
2017-03-20
[Abstract]  Major Histocompatibility Complex (MHC) tetramers have been used for two decades to detect, isolate and characterize T cells specific for various pathogens and tumor antigens. In the context of Human Immunodeficiency Virus (HIV) infection, antigen-specific CD8+ T cells have been extensively studied ex vivo, as they can be readily detected by HIV peptide-loaded MHC class I tetramers. In contrast, the detection of HIV-specific CD4+ T cells has proven more challenging, due to the intrinsically lower clonal expansion rates of CD4+ T cells, and to the ... [摘要]  主要组织相容性复合物(MHC)四聚体已经使用二十年来检测,分离和表征各种病原体和肿瘤抗原特异性的T细胞。在人类免疫缺陷病毒(HIV)感染的背景下,抗原特异性CD8 +细胞已经在体外广泛研究,因为它们可以容易地被HIV肽 - 加载MHC I类四聚体。相比之下,HIV特异性CD4 + sup + T细胞的检测已被证明更具挑战性,因为CD4 + sup + T细胞的本质上较低的克隆扩增率以及优先艾滋病毒感染过程中艾滋病毒特异性CD4 + T细胞的消耗。 在以下协议中,我们描述了一种简单的方法,该方法有助于使用肽负载的MHC II类四聚体来鉴定HIV-1衣壳表位特异性的CD4 + / T细胞。可以分析四聚体标记的CD4 T细胞的细胞表面表型和/或FACS分选用于进一步的下游应用。成功检测特异性CD4 + / / T细胞离体的关键是选择导致高亲和力T细胞受体(TCR)的肽/ MHC II组合, (Benati等人,2016)。 MHC II四聚体阳性细胞的可靠检测的第二个关键点是系统地使用负载无关肽的对照四聚体,样品和对照管在相同的条件下进行处理。背景 在用抗PE微珠对四聚体-PE标记的细胞进行磁力富集后,在纯化的CD4 + T细胞中检测到罕见的HIV特异性MHC II四聚体阳性细胞(Seth等人, em>。,2005)。我们发现使用经验证的肽/ MHC ...

Optogenetic Mapping of Synaptic Connections in Mouse Brain Slices to Define the Functional Connectome of Identified Neuronal Populations
Author:
Date:
2017-01-05
[Abstract]  Functional connectivity in a neural circuit is determined by the strength, incidence, and neurotransmitter nature of its connections (Chuhma, 2015). Using optogenetics the functional synaptic connections between an identified population of neurons and defined postsynaptic target neurons may be measured systematically in order to determine the functional connectome of that identified population. Here we describe the experimental protocol used to investigate the excitatory functional connectome of ventral midbrain dopamine neurons, mediated by glutamate cotransmission (Mingote et al., ... [摘要]  神经回路中的功能连通性由其连接的强度,发病率和神经递质特性决定(Chuhma,2015)。使用光遗传学,可以系统地测量识别的神经元群体和定义的突触后靶神经元之间的功能性突触连接,以便确定该识别群体的功能性连接群。这里我们描述了用于研究由谷氨酸共转播介导的腹侧中脑多巴胺神经元的兴奋性功能性连接体的实验方案(Mingote等,2015)。通过将编码通道视紫红质(ChR2)的腺相关病毒(AAV)注射到DATIREScre小鼠的腹侧中脑中,使多巴胺神经元变得光敏。多巴胺合成酶酪氨酸羟化酶的免疫荧光证实ChR2表达在多巴胺神经元中的功效和特异性。然后,切片膜片钳记录由接受多巴胺神经元投影的区域中的神经元产生,并且确定兴奋性连接的发生率和强度。所有接受多巴胺神经元投射的区域的连接发生率和强度的总结构成功能性连接体。
【背景】为了建立特定神经回路的功能,有必要确定解剖连接,解剖连接的映射及其功能连接,连接的强度,发病率和神经递质性质的映射。使用单因素限制的病毒性突触后追踪技术,可以描述包括多巴胺系统在内的神经回路的复杂解剖连接(Callaway and Luo,2015; ...

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