I already check protein expression level on SDS-PAGE gel.
For checking I already used CBB stainning.
But I'm not sure it was my target protein or not.
So on this gel, I want to do transfer to membrane using same gel.
Is that possible to detect band after coommasie stainning?
Or should I have to do destaining step?
Which Coomassie reagent did you use? If you stained the gel in 40 % MeOH 10 % Acetic acid then the transfer will be fairly inefficient. You will observe better efficiency if you stained with the colloidal form of Coomassie (G-250) with lower concentrations of acid/alcohol.
Better still, re-run your samples and transfer onto PVDF/Nitro membrane. Stain the membrane in Ponceau-S then partially destain (in plain water) to visualise the proteins and note the marker/ladder position (or use coloured markers). Take a photo then fully destain the membrane and procede with the immunodetection. You could also run a duplicate gel and stain that in coomassie blue for comparison.
- Badges
- Science method