OVERVIEW

Tris-Glycine-SDS running buffer is commonly used to resolve proteins in SDS-polyacrylamide gel. A concentrated buffer is usually prepared that is diluted to working concentration at the time of use. Here we show a procedure to prepare 10x Tris-glycine-SDS running buffer. It is also called Laemmli electrophoresis running buffer.

To prepare 1L of 10x Tris-glycine-SDS running buffer using 20% SDS solution, first 30.3 g Tris base and 144 g Glycine are dissolved in 700 ml of deionized water, and subsequently 50 ml of 20% SDS solution is added and the final volume of solution is adjusted to 1L.

REQUIREMENTS

Reagents and solutions
Tris base (Molecular Weight: 121.14) (e.g., Sigma-Aldrich #T1503)
Glycine (Molecular Weight: 75.07) (e.g., Sigma-Aldrich #G8898)
20% Sodium dodecyl sulfate (SDS) solution
Deionized water

* Tris base (CAS Number: 77-86-1) and Trizma® base (CAS Number: 77-86-1) are the same chemicals.

Equipment and disposables
Beaker of suitable size (use a 2 L beaker to prepare 1 L solution)
Magnetic stirrer and beads
Spatula
Weighing balance
Measuring cylinder

Composition: 
Running buffer composition (10x)
0.25 M Tris
1.92 M Glycine
1.0% SDS
pH 8.3

Running buffer composition (1x)
25 mM Tris
192 mM Glycine
0.1% SDS
pH 8.3

Objective
Preparation of 1L of 10x SDS-PAGE Running Buffer

PREPARATION

Use personal protective equipment (lab coat, gloves, goggles, etc) for your safety and follow the guidelines of your institute.

Step 1: Weigh out 30.3 g Tris base and 144 g Glycine and transfer to a 2L beaker.

Step 2: Add 750 ml of deionized water and stir until all reagents dissolve completely.

Step 3: Add 50 ml of 20% SDS solution and adjust the volume to 1L using deionized water.
Transfer all liquid from the beaker to a 1L measuring cylinder and add 50ml of 20% SDS solution. Then add deionized water to the measuring cylinder to adjust the final volume to 1L.

Note: The pH of the solution should be 8.3 without any adjustment.

Storage: 
The solution can be stored at room temperature for a year.