LMW proteins not transfer of gel to membrane - (Jul/19/2013 )
I have a Question! please help me!
I always done the western blot without any problem but recently, I have a prblem, the LMW don't transfer of gel into membrane correctly while all situations are equal in compare to past ( 0.2 micron membrane, low voltage and time related to transfer etc.) I used nitrocellulose membrane, Other proteins (for example upper 40 kd are transferred heterogenous while homogenous in past ) I mean of hetero-homo related to use ponceau S staining. Can you help me? only element changed is PAG! Do you think change polyacrylamide gel can help me?
what is different this time from when the transfer worked perfectly (eg: fresh made buffer, new lot or brand of nitrocellulose, etc)?
i know you said that your gel was changed. what is different about it from what worked in the past (eg: %acrylamide, gradient, chemistry, etc)?
can you show a picture of one that worked and one that didn't?
when something that used to work stops working, you have to determine what has changed since it worked. that is where you will usually find the problem.