Hi,
We use the bio-rad precast gels for SDS-PAGE and the mini-protean tetra setup for running and transferring them. We don't do too many westerns but every now and then.
My issue is with the transfer. We use both an older 3 cell tank mini trans-blot module as well as a newer tetra tank mini trans-blot module. The older 3 cell tank transfers very well and after 90 min at 100V, RT, there are no markers left in the gel and mostly everything has transferred to the membrane. However, with the newer tetra tank, even after 90, or even 120min at the same voltage, there are always high MW markers still left in the gel. We've remade the buffers, added filter paper to increase contact, even bought a new core electrode assembly and tried a new power source from a neighbor lab but no dice. The problem seems to be the new tank design. Has anybody else had this issue with the newer tetra tank?
Alternatively, is there something I'm doing wrong? Why would one tank work better than the other?
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Kaioshin
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Posted 20 January 2011 - 08:18 PM
etodem, on 19 January 2011 - 09:25 PM, said:
Hi,
We use the bio-rad precast gels for SDS-PAGE and the mini-protean tetra setup for running and transferring them. We don't do too many westerns but every now and then.
My issue is with the transfer. We use both an older 3 cell tank mini trans-blot module as well as a newer tetra tank mini trans-blot module. The older 3 cell tank transfers very well and after 90 min at 100V, RT, there are no markers left in the gel and mostly everything has transferred to the membrane. However, with the newer tetra tank, even after 90, or even 120min at the same voltage, there are always high MW markers still left in the gel. We've remade the buffers, added filter paper to increase contact, even bought a new core electrode assembly and tried a new power source from a neighbor lab but no dice. The problem seems to be the new tank design. Has anybody else had this issue with the newer tetra tank?
Alternatively, is there something I'm doing wrong? Why would one tank work better than the other?
We use the bio-rad precast gels for SDS-PAGE and the mini-protean tetra setup for running and transferring them. We don't do too many westerns but every now and then.
My issue is with the transfer. We use both an older 3 cell tank mini trans-blot module as well as a newer tetra tank mini trans-blot module. The older 3 cell tank transfers very well and after 90 min at 100V, RT, there are no markers left in the gel and mostly everything has transferred to the membrane. However, with the newer tetra tank, even after 90, or even 120min at the same voltage, there are always high MW markers still left in the gel. We've remade the buffers, added filter paper to increase contact, even bought a new core electrode assembly and tried a new power source from a neighbor lab but no dice. The problem seems to be the new tank design. Has anybody else had this issue with the newer tetra tank?
Alternatively, is there something I'm doing wrong? Why would one tank work better than the other?
Does your power supply have any way to measure Amps? Check that and see if it varies between the two transfer methods, it could be that the older setup pulls more amps and thus transfers better/faster.
Additionally, for minigels, if you haven't been, you can add 0.02-0.1% SDS to the transfer buffer to aid in transfer.
