Western blotting transfer - (Mar/27/2013 )
I am regularly using western blotting technique and often I get distorted (ladder and /or protein) bands after the western blot transfer, though there is no band distortion during the gel run.
I read that it could be due to air bubbles and inadequate uptake of buffer by the filter paper. I noticed no trapped air bubbles each time. I normally do 2 - 4 blots at one time and some of them show the distorted bands but not all of them.
I cant figure out what is the exact problem. Could you please suggest possible reasons and how to overcome to these problems?
What kind of transfer method do you use? Semi-dry vs. wet? If you are doing Semi-dry, I have seen similar patterning develop if you try to transfer to many gels at once. Once you close the lid to the apparatus, gels can become distorted. This is mostly due to uneven size distribution between each "sandwhich". If you do wet transfer, I would possibly use another colleagues buffers if you have the opportunity. Let me know you procedure and we can figure something out.
Edit: Are you using pre-made gels or are you making your own?
Thank you, I make my own gels and I have encountered this problem with both methods.