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PE-CyTM 7 Mouse Anti-Human CD4

PE-Cy TM 7小鼠抗人CD4

Company: BD
Catalog#: 557852
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HIV-1 Fusion Assay
[Abstract]  The HIV-1 fusion assay measures all steps in the HIV-1 life cycle up to and including viral fusion. It relies on the incorporation of a β-lactamase–Vpr (BlaM-Vpr) protein chimera into the virion and the subsequent transfer of this chimera into the target cell by fusion (Figure 1). The transfer is monitored by the enzymatic cleavage of CCF2, a fluorescent dye substrate of β-lactamase, loaded into the target cells. Cleavage of the β-lactam ring in CCF2 by β-lactamase changes the fluorescence emission spectrum of the dye from green (520 nm) to blue (447 nm). This change reflects virion fusion and can be detected by flow cytometry (Figure 2). [摘要]  HIV-1融合测定测量HIV-1生命周期中直至并包括病毒融合的所有步骤。 它依赖于将β-内酰胺酶-Vpr(BlaM-Vpr)蛋白嵌合体并入病毒体中,并且随后通过融合将该嵌合体转移到靶细胞中(图1)。 通过CCF2(一种β-内酰胺酶的荧光染料底物)的酶裂解来监测转移,将其装载到靶细胞中。 β-内酰胺酶在CCF2中的β-内酰胺环的切割将染料的荧光发射光谱从绿色(520nm)改变为蓝色(447nm)。 这种变化反映了病毒体融合,并且可以通过流式细胞术检测(图2)。

Transfection of Human Naive CD4+ T Cells with PHA Activation and Neon Electroporation
[Abstract]  Transfection of primary T cells can be challenging. This protocol describes a method to transfect primary human naive CD4+ T cells with an AP-1 luciferase reporter using low-level activation by phytohemagglutinin (PHA) and electroporation, as published (Palin et al., 2013). This technique is a modification of one previously described by our group (Cron et al., 2013). Anyone wishing to transfect murine T cells should consult the publication by Cron et al., 2013. This technique may be adapted for other primary T cell types by optimizing the Neon electroporation conditions, as described in the text. Other luciferase or GFP reporters may be used, and will require optimization of the stimulation conditions for that particular reporter. [摘要]  原代T细胞的转染可能是具有挑战性的。 该方案描述了使用植物凝集素(PHA)和电穿孔的低水平激活,用AP-1荧光素酶报道子转染初级人初始CD4 + T细胞的方法,如公开的(Palin等人 。,2013)。 这种技术是先前由我们组描述的技术的修改(Cron等人,2013)。 任何希望转染鼠T细胞的人都应参考Cron等人2013年的出版物。该技术可以通过优化氖电穿孔条件来适应其它原代T细胞类型,如文中所述。 可以使用其他荧光素酶或GFP报道分子,并且将需要优化该特定报道分子的刺激条件。