Hi all,
I tried using 15% gel SDS for running vertical SDS-PAGE gel electrophoresis at conditions: 100V, then 120V after loading sample is lining.
I found out that the loading sample/dye (mixing with 5x loading dye in ratio 1(dye):5(total) and boiled at 95°C for 5 min) ran down much faster than the prestained protein marker does; the dye is near at bottom of gel while prestained protein marker is still at half of the gel only. When stained with Ponceau S, loading protein running down only half of the membrane.
I usually use 10% gel and stop at the dye front are a few cm above bottom of the gel. The dye and the protein are running about the same speed.
My question is whether loading dye's running down quicker than protein loading sample. Don't they suppose to run down together? What are the causes?
Thank you for your answers
The dye front generally moves faster compared to your proteins. I don't see what the problem is, if your bands correspond to the markers.
If your protein marker is prestained you can use it has reference soyou run the complete gel an your proteins dont fallout.
check the components of loading dye.If possible prepare a fresh dye and run the samples.
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