小中大
这是CST技术支持的反馈意见,除了hepg2细胞是我的实验用细胞外，其他细胞全是说明书上指定的阳性细胞株。显示出条带的p15是兔多抗，cdk4是鼠单抗，二抗也是cst的，按说明书1：2000稀释。
下面是CSt的回复，他们提出了一些建议，同时他们想送一个阳性对照给您。
具体内容见下：
************Original email************
Dear Fugang:
The customer was very thorough in the information provided, but I cannot find any problems with the procedure. I am concerned however,because the western blot images are not as clean and clear as I would expect. It is possible that a transfer using a 0.45 micron membrane will work with p15, but it is unlikely. I would like to know if they had indeed performed sonication of their samples, using a tip sonicator and not a water bath sonication? The p15 band is wavy, like it was not sonicated. So, I cannot explain any technical trouble shooting that can help. I can only offer control lysates
that the customer can use with their western blots. Normally I would have thought that the secondary antibodies failed, but the two antibodies to cdk4 and p15 worked, so maybe there is something wrong with the detection. If there is too much protein
loaded, the HRP detection substrate may be depleted before exposure to film.
Let me know if the sample lysate will be helpful.
Sincerely,
Rami Najjar
Product Manager
Cell Signaling Technology, Inc.
