Protocols, optimization tips,
and more for Western Blot.
Western Blot Transfer Optimization
Selecting the Right Transfer Type
Transferring electrophoretically separated proteins to a membrane allows easier handling and better accessibility to antibodies, stains, and signal development reagents. All transfer types rely on the electrophoretic mobility of proteins to push them out of the gel and onto the membrane; the biggest differences between transfer types arise in the type of equipment needed and the speed and efficiency of the transfer. Pick your transfer method carefully to get the best results in your own time and for the right price.
Wet transfer is the most common transfer method. After assembling the transfer sandwich with saturated filter paper, the assembly is locked into a transfer cassette and immersed in a transfer tank filled with buffer before the electric field is activated. This type of transfer is usually run for several hours or even overnight.
Wet transfers are more consistent than faster methods, allowing more quantitative interpretation, but are slow and prone to errors caused by bubbles trapped between the layers of the sandwich.
Semi-dry transfer sandwiches are of the same basic setup as for wet transfe, but rather than immersion in buffer, the componens of the sandwich are soaked in buffer and held between transfer pads and electrode layers. Semi-dry transfer is much faster than wet transfer, typically taking only about 7 minutes. The rapidity of the transfer can cause uneven, blotchy, and inconsistent results. This method is more expensive due to the specialized equipment and consumable materials required.
Dry transfers use a unique gel matrix in place of buffer tanks or soaked filter paper. The high ionic density of the gel matrix enables more rapid protein transfer. Since this method does not involve immersing the electrodes in liquid, no oxygen is produced during electrotransfer, which reduces blot distortion. Dry transfer is fast and produces more consistent results, but is also the most expensive option.
Keywords: Western blotting optimization, protein transfer, wet transfer, semi-dry transfer, dry transfer, best transfer methodClick for more optimization tips