| Thioglycollate-elicited Peritoneal Macrophages Preparation and Arginase Activity Measurement in IL-4 Stimulated Macrophages
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Author:
Date:
2015-09-05
[Abstract] Macrophages are an essential cell population of innate immunity that plays important roles in inflammatory processes. Two main different phenotypes have been described with opposing activities: The classically activated macrophages (M1) and the alternatively activated macrophages (M2). Alternative activation of mouse macrophages can be induced by type 2 cytokines such as IL-4 and it is characterized by the regulation of the L-arginine metabolism. M2 macrophages convert arginine to ornithine and urea through the action of Arginase-1. Here we described a method for the isolation of peritoneal macrophages from thioglycollate-elicited mice and alternative activation by stimulation with IL-4. Intraperitoneal injection of thioglycollate elicits large numbers of macrophages into peritoneal ...
[摘要] 巨噬细胞是在炎症过程中起重要作用的先天免疫的必需细胞群体。 已经描述了具有相反活性的两种主要不同表型：经典激活的巨噬细胞（M1）和交替激活的巨噬细胞（M2）。 小鼠巨噬细胞的备选激活可以由2型细胞因子例如IL-4诱导，并且其特征在于L-精氨酸代谢的调节。 M2巨噬细胞通过精氨酸酶-1的作用将精氨酸转化为鸟氨酸和尿素。 在这里我们描述了从巯基乙酸诱发小鼠分离腹膜巨噬细胞和IL-4刺激的替代激活的方法。 腹膜内注射巯基乙酸盐引发大量巨噬细胞进入腹膜腔。
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| In vitro Colorimetric Method to Measure Plant Glutamate Dehydrogenase Enzyme Activity
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Author:
Date:
2015-08-20
[Abstract] Glutamate dehydrogenase (GDH) is an NAD(H) dependent enzyme that catalyzes, in vitro, the reversible amination of glutamate. Here we describe how to determine spectrophotometrically GDH activity monitoring NADH evolution. This protocol is described here for Arabidopsis thaliana (A. thaliana) although it is also valid for other plant species. GDH protein is a hexamer composed, in the case of Arabidopsis, of a combination of GDHα, GDHβ and GDHγ subunits. Every combination of subunits is possible; however, it is still barely known whether different combinations affect the enzymatic properties of the hexamers. In other species, hexamers are a combination of GDHα and GDHβ but it cannot be discarded the existence of other genes since for instance GDHγ subunit ...
[摘要] 谷氨酸脱氢酶（GDH）是NAD（H）依赖性酶，其在体外催化谷氨酸盐的可逆胺化。在这里我们描述如何确定分光光度法GDH活动监测NADH进化。此协议在这里描述为拟南芥（ A thaliana ），虽然它也适用于其他植物物种。 GDH蛋白是在拟南芥属的情况下由GDHα，GDHβ和GDHγ亚基的组合组成的六聚体。每个亚基的组合是可能的;然而，仍然几乎不知道不同的组合是否影响六聚体的酶性质。在其他物种中，六聚体是GDHα和GDHβ的组合，但不能丢弃其它基因的存在，因为例如在Fontaine等人（2012）中描述了拟南芥中的GDHγ亚基。
谷氨酸盐+ NAD + + H + →2-氧代戊二酸+ NADH + NH4 +
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| Protein Flotation Assay to Isolate Lipids Rafts from Soft Tissue or Cells
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Author:
Date:
2013-08-20
[Abstract] The arrangement in eukaryotic cell membranes of liquid-ordered states surrounded by liquid-disordered phases allows for the existence of organized membrane microdomains called rafts. Rafts play a crucial role in signal-transduction events, in lipid transport and in various internalization processes, and for all these reasons need to be purified for further characterization.
[摘要] 在由液体无序相包围的液体有序状态的真核细胞膜中的排列允许存在称为筏的有组织的膜微结构域。 木筏在信号转导事件，脂质运输和各种内化过程中发挥关键作用，并且由于所有这些原因，需要进行纯化以进一步表征。
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