I am using a 4-12% Bis-Tris NuPage (Invitrogen) ready-made gel , and run in MES buffer to enable me to view the protein size of less than 10k Da.
However, when i transferred the gel to the Immobilon PVDF 0.2um using Trans-Blot SD Semi-Dry (Biorad), at 12V for 45 min for the mini gel , the transferring is still incomplete when i stained my gel with coomasie blue after transferring it.
The transfer buffer which I am using is theglyicine , tris base, SDS and methanol.
I did not use the running buffer as suggested for NuPage transfer buffer.
Is it because of the different transfer buffer which causes the inefficiency of the transferring?