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看完这篇文章,不要再傻傻分不清nitrocellulose和PVDF

在做Western Blot的时候,膜的选择是整个实验的一个关键点,所以大家要谨慎选择。毕竟它是一个物理支撑物,不但要hold住宝贵的样品,还要有“坚韧不拔”的精神来抵抗住各种实验条件。每个研究人员的目标蛋白特点和下游检测方法不尽相同,要想找到适合自己的转渍膜,需要通过一系列的实验来找到。为了方便大家比较和参考,小编在这里对比和总结了Nitrocellulose 和PVDF膜的特点(如下表所示)。如果大家有更多建议,也欢迎和我们取得联系。

 

表格1:Nitrocellulose 和 PVDF 性质比较。

 

Nitrocellulose

PVDF

中文名称

硝酸纤维素

聚偏二氟乙烯

与蛋白质的结合能力(μg/cm2

80-100

170-200

详细用途

·Western blotting (尤其针对分子量小于20kDa的蛋白质)

·核酸分析

·氨基酸分析

·Dot/slot blotting

·Western blotting (尤其针对分子量较大的蛋白质)

·蛋白质测序

·氨基酸分析

与蛋白质结合的作用力

疏水性相互作用

疏水性相互作用和偶极偶极相互作

物理特性

相对脆弱

较高的机械强度

膜孔径的基本信息

是的。产品通常都有0.1,
0.2
或是 0.45μm孔径的。0.45μm 适用于大多数蛋白质,而0.1, 0.2μm适合分子量小的蛋白质。

化学耐性

没有

敏感度

敏感程度相对较低,但是可以大大减少背景信号。

可以检测蛋白质含量非常低的样品,但同时也会有很多背景信号作为干扰。

蛋白质剥离并重新探测的过程

不建议。信号会大大减弱。

可以。蛋白质在膜上的吸附程度更紧密。

甲醇的使用

需要。转渍过程中必须使用。但是可以导致大分子量的蛋白质沉淀。

不需要。膜必须在使用之前用甲醇浸泡,但在转渍过程中可以使用不含有甲醇的缓冲液。这个方法可以预防分子量较大的蛋白质沉淀。

SDS的添加

不需要。这样可以增大小分子量的蛋白质与膜结合。

需要。这样可以大大提高转渍效率。

 



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