apoB做Western Blot转膜的时间

题目

apoB做Western Blot转膜的时间
我在做apoB100的Western,它的分子量是512kD,我老是做不出来,我想问问做出来的朋友们,你们用百分之几的SDS-PAGE胶跑的电泳,转膜的条件是什么?

答案

512KD?这个蛋白太大了一点.我最多转过200多KD的,你试试看转过夜呢?
网上找了一篇文献,Plos One 2008
ApoB100-LDL Acts as a Metabolic Signal from Liver to Peripheral Fat Causing Inhibition of Lipolysis in Adipocytes
里面没有特别讲条件,但确实做了western,ApoB100和48都有,估计转膜时间要很长很长,他们的条件如下：
Mouse plasma samples (2 μl) were fractionated by SDS-PAGE using 4% gels.The apoB proteins were detected with rabbit antiserum against mouse apoB (Biosite,Taby,Sweden).The binding of primary antibodies was assessed with horseradish peroxidase–labeled donkey anti-rabbit antibodies and ECL western blotting reagents (Amersham Biosciences,GE Healthcare Biosciences,Little Chalfont,Buckinghamshire,England).