this will be difficult since the sizes are so different. I would use a gradient gel to separate them (4-20%) then cut the membrane and transfer separately as you will likely lose the 14kD band to try to get the 280kD band.

I would do a traditional overnight wet transfer for the larger protein and omit methanol from the transfer buffer.

Wet tank only

For proteins 280-14 kDa, the best western blotting conditions for the Invitrogen Power Blotter are as follows: Protein Size: 280-14 kDa Transfer Buffer: Tris-Glycine, pH 8.3 Transfer Voltage: 50-100 V Transfer Time: 1-2 hours Blocking Buffer: 5% nonfat milk in PBS-Tween Primary Antibody Concentration: 1:1000-1:5000 Secondary Antibody Concentration: 1:5000-1:10000 Detection Method: Chemiluminescent or fluorescent

Bandla Ramesh Thank you, I will go through it.

Pakorn Pengin Thank you.

Julie Ann Dougherty thank you, I will look into it.

The conditions for western blotting, including the transfer and detection methods, are generally optimized for the specific proteins being analyzed and the antibodies used for detection. However, some general guidelines can be followed for western blotting of proteins of different sizes using the Invitrogen Power Blotter system.

For proteins in the range of 280-14 kDa, the following conditions can be used as a starting point:

It is important to note that these conditions are a general starting point and may need to be optimized for individual proteins and antibodies. Additionally, the Invitrogen Power Blotter system may have specific recommendations for use that should be followed.