Lidstrom: SDS-PAGE
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Gel Prep
- Clean cover plate and thicker spacer plate 75 mM
- Soap and Water
- Ethanol
- DI water
- Dry plates
- Setup one spacer plate and one cover plate in each gel holder
- The cover plate goes on the side of the spacer plate with the spacers in order to create a small gap between the plates
- Put the gel holder into the casting stand
Pour the Resolving Gel
- Mix components of the amounts in the Gel Mix link. Mix in the order listed.
- Gel Mix Recipe
- Don't add APS/TEMED until ready to pour
- Use pipette to put gel mix into the gap between the plates
- Carefully layer 50%EtOH 50% ddH2O on top of the gel to prevent the top of the gel from drying out
- Let dry for an hour
Pour the Stacking Gel
- Mix components of the amounts in the Gel Mix link. Mix in the order listed.
- Gel Mix Recipe
- Don't add APS/TEMED until ready to pour
- Use pipette to put gel mix into the gap between the plates
- Insert the comb being careful not to trap any bubbles
- Let dry for an hour
Sample Prep
- 10-20 ug protein
- Load 12-15 uL, absolute max 20 uL for big comb
- Denature protein by mixing with sample buffer, BME and boiling