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Lipopolysaccharides from Escherichia coli 0111:B4


Company: Sigma-Aldrich
Catalog#: L4391
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Study of Epithelium Barrier Functions by Real-time TER Measurement
[Abstract]  Transepithelial Electrical Resistance (TER) measurement is a reliable and efficient method to quantify the permeability of barrier forming cells such as epithelial cells. Measuring the permeability of the epithelial cells will help the researchers to investigate the barrier function of epithelium in various infectious and inflammatory diseases. Here we provide a real-time and impedance-based approach for measuring the permeability of epithelial cell monolayer using the Electrical Cell Substrate Impedance Sensing (ECIS®) instrumentation. [摘要]  经上皮电阻(TER)测量是量化屏障形成细胞例如上皮细胞的渗透性的可靠和有效的方法。 测量上皮细胞的通透性将有助于研究人员调查上皮在各种感染和炎症疾病中的屏障功能。 在这里,我们提供了使用电细胞底物阻抗感测(ECIS )仪器测量上皮细胞单层的渗透性的基于实时和阻抗的方法。

Cell-based Assays to Monitor AID Activity
[Abstract]  The enzyme Activation induced deaminase (AID) underpins antibody affinity maturation and isotype switching through its mutagenic activity of deaminating deoxycytidine to deoxyuridine in DNA. Subsequent processing of the deoxyuridine initiates the processes of somatic hypermutation (SHM) and class switch recombination (CSR) in B cells. Structure-function analysis of AID requires sensitive and biologically relevant methods to measure its various activities. Here we describe simple but effective methods to measure 1) the ability of AID to mutate the Escherichia coli genome, which provides an indication of its catalytic activity; 2) the capacity of AID to perform SHM by complementing a derivative of the DT40 chicken B cell line; 3) the ability of AID to perform CSR by complementing ... [摘要]  酶活化诱导的脱氨酶(AID)通过其将脱氧胞苷脱氨基到DNA中的脱氧尿苷的诱变活性来支持抗体亲和力成熟和同种型转换。脱氧尿苷的后续加工引发B细胞中体细胞超突变(SHM)和类型转换重组(CSR)的过程。 AID的结构功能分析需要灵敏和生物相关的方法来测量其各种活动。在这里我们描述简单但有效的方法来测量1)AID突变大肠杆菌基因组的能力,其提供其催化活性的指示; 2)AID通过补充DT40鸡B细胞系的衍生物来进行SHM的能力; 3)AID通过补充AID缺乏的原代小鼠B细胞来进行CSR的能力。三种方法的组合,伴随着AID亚细胞定位和蛋白质表达水平和稳定性的必要分析作为对照,允许AID的详细结构功能研究。

Quantification of ex vivo Neutrophil Extracellular Traps
[Abstract]  Neutrophil extracellular traps (NETs) are fibrous mesh-like, web-like, or string-like structures which are composed of DNA, histones, and granule proteins such as neutrophil elastase or myeloperoxidase. When activated by phorbol myristate acetate, interleukin-8, lipopolysaccharide (LPS), and various pathogens, neutrophils release NETs. We reported that NETs were classified as two distinct forms; cell-free NETs that were released away from neutrophils and anchored NETs that were anchored to neutrophils. In general, extracellular DNAs are used as a surrogate marker of NETs. Here, we describe a protocol regarding quantitative procedures of extracellular DNAs released from ex vivo neutrophils activated by LPS using fluorometric double-stranded DNA (dsDNA) quantification assay. [摘要]  中性粒细胞外陷阱(NET)是由DNA,组蛋白和颗粒蛋白如嗜中性粒细胞弹性蛋白酶或髓过氧化物酶组成的纤维状网状,网状或绳状结构。 当被佛波醇肉豆蔻酸乙酯,白细胞介素-8,脂多糖(LPS)和各种病原体激活时,嗜中性粒细胞释放NETs。 我们报告,NETs被分为两种不同的形式; 从嗜中性粒细胞释放的无细胞NETs和锚定于嗜中性粒细胞的锚定NET。 通常,细胞外DNA用作NETs的替代标记。 在这里,我们描述一个议定书关于使用荧光双链DNA(dsDNA)定量测定由LPS激活的离体嗜中性粒细胞释放的细胞外DNA的定量程序。