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pre stained marker issues - (Mar/08/2010 )

Hello,

I am running a 5% gel and am seeing some issues- I transfected HEK293 cells and fibroblasts with my plasmid of interest. I obtained the cell lysates by incubating the cells with RIPA buffer, centrifugation and collecting supernatant.

When I run the gel, the pre stained marker is not separating at all. My protein size is 110KD. I only see the blue line from the samples on the gel but no clean separation of the marker.

Can anybody help me on this issue?

Thank you.

-xyz74-

Marker hasn't degraded has it? How much did you load? Have you used it before?

-bob1-

a 5% gel is neither very selective for a 110kDa protein nor for your markers.

you should use at least 7.5% but 10% would be better. a gradient (we like 5-15%) might be even better.

-mdfenko-