样品分析
概述
准备样品(微孔板孔)从板上取下液体
加入100μL/孔的iFluor 546-鬼笔环肽溶液
在室温下染色细胞15到60分钟洗涤细胞
使用TRITC通道在显微镜下观察样品(Ex / Em = 550/575 nm)
注意:打开前将所有组件加热至室温
操作步骤
1.准备1X iFluor 546-鬼笔环肽工作溶液:
将10μL的iFluor 546-鬼笔环肽(组分A)加入10mL标记缓冲液(组分B)中。
注意1:未使用的1X iFluor 546-鬼笔环肽原液应等分并保存在-20 ℃,避光。
注意2:不同的细胞类型可能染色不同。 应相应地制备iFluor 546-鬼笔环肽工作溶液的浓度。
2.染色细胞:
2.1执行甲醛固定。 在室温下用3.0-4.0%甲醛的PBS孵育细胞10-30分钟。
注意:避免使用任何含甲醇的固定剂,因为甲醇会在固定过程中破坏肌动蛋白。 优选的固定剂是不含甲醇的甲醛。
2.2用PBS冲洗固定的细胞2-3次。
2.3可选:在PBS中加入0.1%Triton X-100固定细胞(步骤2.2)3至5分钟,以增加渗透性。 用PBS冲洗细胞2-3次。
2.4将100μL/孔(96孔板)的iFluor 546-鬼笔环肽工作溶液(来自步骤1)加入固定的细胞(来自步骤2.2或2.3),并在室温下将细胞染色15至60分钟。
2.5用PBS轻轻冲洗细胞2至3次以去除多余的染料,然后使用TRITC通道进行板密封和成像。
图1.用甲醛固定并用Cell Navigator F-肌动蛋白标记试剂盒染色的CPA细胞的图像*在Costar黑色96孔板中的橙色荧光* A:用1X iFluor 546-鬼笔环肽标记细胞仅30分钟。 B:用鬼笔环肽处理细胞10分钟,然后用1X iFluor 546-鬼笔环肽染色30分钟。
试剂应用文献
Mussel-inspired conductive nanofibrous membranes repair myocardial infarction by enhancing cardiac function and revascularization
Authors: Yutong He, Genlan Ye, Chen Song, Chuangkun Li, Weirong Xiong, Lei Yu, Xiaozhong Qiu, Leyu Wang
Journal: Theranostics. 2018; 8(18): 5159–5177.
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