样品实验方案
以下是我们推荐的方案,仅提供指导。具体实验应根据您的特定需求进行修改。
简要概述
- 根据需要处理样品
- 准备Xite Red β-D-吡喃半乳糖苷工作溶液并将其添加到样品中
- 在37°C下孵育样品15至45分钟
- 使用带有575/26 nm滤光片的流式细胞仪(PE通道)或带有Cy3/TRITC滤光片组的荧光显微镜检测荧光强度
溶液配制
储备溶液配制
Xite Red β-D-吡喃半乳糖苷储备溶液:在Xite Red β-D-吡喃半乳糖苷中加入适量的DMSO,制成2-5 mM Xite Red β-D-吡喃半乳糖苷原液。注意:将未使用的Xite Red β D-吡喃半乳糖苷原液以等份储存在-20°C下。
工作溶液配制
Xite Red β-D-吡喃半乳糖苷工作溶液:在自备的缓冲液中制备1-20 µM Xite Red β-D-吡喃半乳糖苷工作溶液。注意1:Xite Red β-D-吡喃半乳糖苷工作溶液应立即使用。注意2:Xite Red β-D-吡喃半乳糖苷的浓度应针对不同的细胞类型和条件进行优化。
操作步骤
- 根据需要处理样品。
- 处理并用自备的缓冲液(例如DPBS)洗涤细胞。
- 加入Xite Red β-D-吡喃半乳糖苷工作溶液15-45分钟,然后在37°C的培养箱中培养样品。
注意:孵育的最佳时间需要通过实验确定。
- 取出工作溶液并用自备的缓冲液洗涤细胞。
- 将细胞重悬在自备的缓冲液中,并使用流式细胞仪使用575/26 nm滤光片(PE通道)或带有Cy3/TRITC滤光片组的荧光显微镜检测荧光强度。
图示
图1.用Xite Red β-D-吡喃半乳糖苷测量β-gal的表达。将9L-LacZ细胞(过度表达β-gal的细胞)与Xite Red β-D-吡喃半乳糖苷在37°C下孵育30分钟。使用NovoCyte流式细胞仪(ACEA Biosciences)通过PE通道获取信号。
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