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Antibiotic/antimitotic solution

N-乙基马来酰亚胺

Company: Thermo Fisher Scientific
Catalog#: 15240062
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Single-cell qPCR Assay with Massively Parallel Microfluidic System
Author:
Date:
2020-03-20
[Abstract]  The single-cell transcriptome is the set of messenger RNA molecules expressed in one cell. It is extremely variable and changes according to external, physical and biochemical conditions. Due to sensitivity shortages, most of genetic studies use bulk samples, providing only the average gene expression. Single-cell technologies have provided a powerful approach to a more detailed understanding of the heterogenic populations and minority cells. However, since it is still a quite novel technique, standardized protocol has to be established. Single-cell qPCR, although partly limited by the number of genes, is relatively simple to analyze. Therefore, its use is accessible without the necessity to recourse to complex bioinformatics analyses. The main steps for single-cell qPCR, as illustrated ... [摘要]  [摘要 ] 单细胞转录组是在一个细胞中表达的信使RNA分子的集合。它变化很大,并会根据外部,物理和生化条件而变化。由于敏感性不足,大多数基因研究使用大量样品,仅提供平均基因表达。单细胞技术提供了一种强大的方法,可以更详细地了解异质群体和少数细胞。然而,因为它仍然是一个相当新的技术,标准化协议具有至b e建立。尽管单细胞qPCR受基因数量的限制,但分析起来相对简单。因此,无需使用复杂的生物信息学分析就可以使用它。如本协议所述,单细胞qPCR的主要步骤包括单细胞分离,细胞裂解液,cDNA逆转录合成,cDNA库生成的扩增以及最终的定量聚合酶链反应。

[背景 ] 的单细胞转录是一套完整的在一个细胞中表达的信使RNA(mRNA)分子。它变化很大,并会根据外部,物理和生化条件而变化。因此,这是生物异质性的来源,其中来自相同环境的细胞与其他细胞相似但不相同。

由于灵敏度不足,大多数遗传研究使用大量样本,其中有数百至数千个细胞,仅提供平均基因表达。这极大地限制了少数细胞群体的研究,这可能会带来特定的特性,例如在癌症情况下的耐药性或转移能力。单细胞技术能够在单细胞水平上分析转录组,从而揭示了异源群体的复杂性。这些技术不仅应用于癌症,而且还应用于许多细胞生物学研究中,包括成年组织,干细胞,免疫细胞等。以及微生物学和病毒学等其他领域(Wen and ...

Long-term in vitro Culture of Cryptosporidium parvum
Author:
Date:
2018-08-05
[Abstract]  Continuous in vitro growth of Cryptosporidium parvum has proved difficult and conventional in vitro culture techniques result in short-term (2-5 days) growth of the parasite resulting in thin-walled oocysts that fail to propagate using in vitro cultures, and do not produce an active infection using immunosuppressed or immunodeficient mouse models (Arrowood, 2002). Here we describe the use of hollow fiber bioreactors (HFB) that simulate in vivo conditions by providing oxygen and nutrients to host intestinal cells from the basal surface and permit the establishment of a low redox, high nutrient environment on the apical surface. When inoculated with 105 C. parvum (Iowa isolate) oocysts the bioreactor produced 108 ... [摘要]  Cryptosporidium parvum 的连续体外生长已证明是困难的,并且常规体外培养技术导致短期(2-5天)生长寄生虫导致薄壁卵囊不能使用体外培养物繁殖,并且不使用免疫抑制或免疫缺陷小鼠模型产生活跃感染(Arrowood,2002)。在这里,我们描述了中空纤维生物反应器(HFB)的使用,通过提供氧气和营养物质从基础表面宿主肠细胞模拟体内条件,并允许建立低氧化还原,高营养环境顶面。当接种10 5 C时。 parvum (爱荷华州分离物)卵囊生物反应器在14天后每ml产生10个 8 卵囊(20ml额外毛细血管体积),并保持2年以上。使用TCR-α免疫缺陷小鼠模型的体内感染性研究显示,在6,12和18个月时从生物反应器产生的卵囊与用于启动培养的亲本Iowa分离物无法区分。 HFB产生的卵囊具有与亲本爱荷华分离物类似的百分比分析。

【背景】 Cryptosporidium parvum 是人和其他哺乳动物肠道的细胞内专性寄生虫,导致急性腹泻。该疾病在免疫功能正常的个体中是自限性的,然而,在免疫功能低下的成人和幼儿中,该疾病可能危及生命(Kotloff,2017)。它是经济资源低的国家中三种被诊断出的儿童肠道疾病之一(Kotloff et al。,2013; Sow et ...

Generation of Chemically Induced Liver Progenitors (CLiPs) from Rat Adult Hepatocytes
Author:
Date:
2018-01-20
[Abstract]  Primary mature hepatocytes (MHs) or their progenitor cells are candidate cell sources for cell transplantation therapy in severe liver diseases. However, stable culture of these cells or generation of equivalent cells from pluripotent stem cells has been limited. Using a cocktail of small molecules that we previously found useful in stable culture of multiple types of stem/progenitor cells, we recently established a novel method to generate bipotent liver progenitor cells, named chemically induced liver progenitors (CLiPs), from adult rat MHs. Here, we describe a detailed protocol for the induction of rat CLiPs. We first describe the method to isolate primary rat MHs and then describe how to induce CLiPs from these MHs. In addition, we describe a method to evaluate the bipotentiality of ... [摘要]  原代成熟肝细胞(MH)或其祖细胞是重症肝病中细胞移植治疗的候选细胞来源。然而,这些细胞的稳定培养或多能干细胞的等效细胞的产生受到限制。我们使用先前在多种类型的干/祖细胞稳定培养中发现有用的小分子混合物,最近建立了一种从成年大鼠MHs产生双能肝脏祖细胞(命名为化学诱导肝祖细胞(CLiPs))的新方法。在这里,我们描述了诱导大鼠CLiPs的详细方案。我们首先描述分离原代鼠MH的方法,然后描述如何从这些MH中诱导CLiPs。另外,我们描述了一种评估产生的CLiPs分化成肝细胞和胆管上皮细胞的双能性的方法。我们还介绍了如何通过长期的文化和详细的示例数据建立稳定的CLiP。可以在2周内产生初级CLiPs,并且可以在2.5-4个月内建立经历10次传代的稳定的CLiPs,批次间变异性。
【背景】对于实现肝病再生医学的新型细胞来源有着强烈的需求。目前唯一的治疗终末期肝病的方法是肝移植,但是由于供者短缺,其应用受到限制。最近,我们小组提出了一种产生能够在体外稳定地扩增的新型LPC的方法,并且可以以广泛的效率重新繁殖慢性肝炎动物模型的损伤肝脏(Katsuda等人, / ...

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