4X Laemmli Buffer¶
Recipe¶
| Component | Amount | Final (1X) Concentration |
|---|---|---|
| 1 M Tris-HCl, pH 6.8 | 12.5 mL | 62.5 mM |
| Glycerol | 20 mL | 10% |
| SDS | 4 g | 2% |
| Bromophenol blue | 10 mg | 0.005% |
| Milli-Q water | to 45 mL | |
| BME (2-mercaptethanol) | 5 mL (add fresh!) | 2.5% |
- Combine the above ingredients in a 50-mL conical tube, and mix with rotation for one hour.
- Store at room temperature.
- To make 10 mL of 4X Laemmli buffer, combine 9 mL with 1 mL BME.
- To make 10 mL of 2X Laemmli buffer, combine 4.5 mL with 0.5 mL BME, and 5 mL water.
- Buffer aliquots containing BME can be stored at -20 °C for several months. Concentrated buffer without BME can be stored at room temperature for at least a year. It may have to be resuspended with rotation before adding BME.
Usage¶
- Combine Laemmli buffer with cellular lysate or other protein-containing sample, to a final concentration of 1X (e.g. add 20 μL of 4X Laemmli buffer to 60 μL lysate, or add 60 μL of 2X Laemmli buffer to 60 μL lysate).
- Incubate at 100 °C for 5 minutes, or 70 °C for 10 minutes.
- Load 5-20 μL of sample to an SDS-PAGE gel for electrophoresis.
Reference¶
Modified from a Cold Spring Harbor protocol, and from the formulation given by Bio-Rad. This version has the original Tris-HCl concentration specified by Laemmli (1970), reduces final BME concentration to 2.5%, and calls for adding fresh BME to enable room temperature storage of concentrated buffer.
Last Udate¶
2025-10-16, jyelland