1× SDS Running Buffer
For this exercise, you will need to prepare 1 L of 1x SDS Running Buffer. The final
composition of this buffer is: 25 mM Tris, 200 mM Glycine pH 8.3, 0.1% (w/v) SDS
1. To a 1 L beaker, add 500 ml of distilled water. Place a magnetic stirring bar into
the beaker.
2. Calculate the amount of Tris base (MW 121.1) required give a final concentration of 25 mM. After checking the calculation with your demonstrator, weigh out the appropriate amount of Tris and add this to the beaker.
3. Calculate the amount of Glycine (MW 75.0) required give a final concentration of 200 mM. After checking the calculation with your demonstrator, weigh out the appropriate amount of Glycine and add this to the beaker.
4. Place the solution on the stirring plate until the above components have dissolved.
5. Calculate the volume of 10% (w/v) SDS to give a final concentration of 0.1% (w/v) and add to the beaker.
6. Using a graduated cylinder, adjust the volume to 1 L with distilled water.
7. Transfer the solution to a glass Schott bottle and check the pH using a pH strip.
Label the bottle appropriately
2. M = mols/L. You want 25 mM or 0.025 mols.
grams tris = mols tris x molar mass tris.
3. You want 200 mM or 0.2 mols.
grams glycine = mols glycine x molar mass glycine.
5. You have 10% stock solution of SDS. You want 0.1%. You want to dilute it 100 times and you want 1 L final solution so you take 10 mL of the stock and 10% x (10/1000) = 0.1%.
To prepare 1 L of 1x SDS Running Buffer with the given composition of 25 mM Tris, 200 mM Glycine pH 8.3, and 0.1% (w/v) SDS, follow these steps:
1. Start by adding 500 ml of distilled water to a 1 L beaker. Place a magnetic stirring bar into the beaker to facilitate mixing.
2. The first calculation involves determining the amount of Tris base (with a molecular weight of 121.1) required to achieve a final concentration of 25 mM. Check your calculation with your demonstrator, and then weigh out the appropriate amount of Tris. Add the Tris to the beaker.
3. Next, calculate the amount of Glycine (with a molecular weight of 75.0) required to achieve a final concentration of 200 mM. Again, check your calculation with your demonstrator, and then weigh out the appropriate amount of Glycine. Add the Glycine to the beaker.
4. Place the beaker on a stirring plate and allow it to stir until the Tris and Glycine have completely dissolved.
5. The next calculation involves determining the volume of 10% (w/v) SDS needed to achieve a final concentration of 0.1% (w/v). Check your calculation with your demonstrator. Once you have the correct volume, add the SDS to the beaker.
6. Using a graduated cylinder, adjust the volume to 1 L by adding distilled water. This step ensures that you have a final volume of 1 L for the buffer solution.
7. After preparing the buffer, transfer it to a glass Schott bottle for storage. Before sealing the bottle, use a pH strip to check the pH of the solution and ensure it matches the desired pH of 8.3. If necessary, adjust the pH by adding small amounts of acid or base as needed.
Finally, make sure to label the bottle appropriately, indicating that it contains 1x SDS Running Buffer and noting the pH.