Histone Purification - Troubleshooting (Jul/01/2008 )
hey guys,
I need some help again. I have been trying to purify histone from the ACTIVE MOTIF kit. But I had tried 3 times but still doesnt work. There was NO pellet at all after precipitation. Other people in the lab used different cell lines but they can purify it with the same kit.
Could it be the cell lines? I can't think of anything else. Thanks!
-timjim-
QUOTE (timjim @ Jul 1 2008, 05:48 AM)
hey guys,
I need some help again. I have been trying to purify histone from the ACTIVE MOTIF kit. But I had tried 3 times but still doesnt work. There was NO pellet at all after precipitation. Other people in the lab used different cell lines but they can purify it with the same kit.
Could it be the cell lines? I can't think of anything else. Thanks!
I need some help again. I have been trying to purify histone from the ACTIVE MOTIF kit. But I had tried 3 times but still doesnt work. There was NO pellet at all after precipitation. Other people in the lab used different cell lines but they can purify it with the same kit.
Could it be the cell lines? I can't think of anything else. Thanks!
Check if you are using roughly the same amount of cells?
-genehunter-1-
I did. 80-90% of confluency on 150mm dish. It is the same with the manual. Unless we have different way of judging the %?
-timjim-
QUOTE (timjim @ Jul 1 2008, 08:34 AM)
I did. 80-90% of confluency on 150mm dish. It is the same with the manual. Unless we have different way of judging the %?
cells come with different sizes, therefore the number of cells per sq of area would be different, therefore, keeping cell mumber roghtly the same would be important.
-genehunter-1-
so do you recommend me to count the cells? How can I do that? I have cells on 150mm dish.
Have you use the same kit too?
-timjim-
If I have some other protein from the elution, does that mean I have sufficient cells?
-timjim-