Protocol
NuPAGE™ Bis-Tris Gels are precast polyacrylamide gels designed for optimal separation and resolution of small- to medium-sized proteins under denaturing gel electrophoresis conditions.
- To the aliquot of 5 µg proteins and peptides, add NuPAGE™ LDS Sample Buffer which is 4-times concentrated
Mix thoroughly on the vortex
Heat samples at 70°C for 10 minutes
Add 50 ml of 20X NuPAGE™ MOPS SDS Running Buffer to 950 ml of deionized water to prepare 1X SDS Running Buffer.
Prepare the gel, by removing the comb, and rinsing the gel wells three times using 1X Running Buffer.
Remove the white tape near the bottom of the gel cassettes.
Place the gels in the mini gel tank.
Fill the inner chamber with the 1X MOPS running buffer and wait until you are sure, that there are no leaks of the buffer.
Pour the running buffer to the lower chamber as well.
Load the appropriate volume of your samples in the appropriate wells.
Load your protein ladder in the appropriate well
If using MOPS Running Buffer, run for 50 minutes at 200 V constant
Pour the 1x MOPS over a sink into a bottle
Carefully get the gel out of the plastic frame and put it into a container.
Wash the gel thoroughly with water.
Pour enough of the staining reagent on top of the gel, so that the gel is covered and shake for 1h
Remove the reagent and wash the gel with water.