This is an instructional video on how to set up and run an SDS-PAGE gel. The following are all the materials, chemicals, and instruments used in this experiment.

Start by preparing your protein samples. Typically, this involves mixing them with a loading buffer containing SDS and a reducing agent such as beta-mecaptoethanol to denature the proteins and break disulfide bonds, otherwise called the Lambly buffer. Heat the sample mixture at 95 degrees Celsius for five minutes to further denature the proteins.

Shortly spin the samples to collect everything at the bottom of the reaction tube.

Assemble the stored SDS-PAGE gel in the chamber with the comb facing on the inside part.

Since here we only run one gel, a dummy plate is used to stabilize the chamber.

Fill up the chamber with SDS-PAGE running buffer. Use the lines on the chamber as reference. They indicate how much the chamber should be filled up with buffer depending on the number of gels you are running.

Once the gel is fully submerged in buffer, carefully remove the comb. One by one load the samples. If the loading dye works properly, the proteins will accumulate at the bottom of the well. One well is reserved for the latter.

Close the chamber by paying attention to how it is connected to the power supply. Turn on the power supply and set the correct parameters to start the separation of the proteins. Allow the gel to run for 20 to 40 minutes, again depending on the size of the proteins you are analyzing.

Be careful not to prolong the run since the proteins can pass outside of the gel. You can confirm that the gel is running by looking if there are small bubbles formed at the bottom of the chamber. Once the proteins are close to the bottom of the gel indicated by the blue line, stop the run and turn off the power supply. Remove the cover of the chamber and disassemble the gel.

Usually for further analysis, the stacking part of the gel is removed as shown in the video.

In this experiment, we stained the gel with quick kamasi dye, so the resolving part of the gel was transferred to a clean plastic container. Pour kamasi to cover the gel and place it on a shaker for 10 to 15 minutes.

Once blue bands are visible, the gel can be analyzed for proteins of interest at the respective sizes.

To destain the gel, collect the dye, add distilled water and leave the gel on the shaker overnight.

This way you have managed to successfully separate protein samples on an SDS-PAGE gel.