6X Laemmli buffer
Laemmli is a sample buffer to use in western blot. 6X sample buffer is added to each protein sample and is boiled or heated for 5-10 minutes.
Recipe to prepare 10 ml:
- 1.2gr SDS (sodium dodecyl sulfate)
- 6mg bromophenol blue
- 4.7ml glycerol
- 1.2ml Tris 0.5M pH6.8
- 2.1ml dwater
warm it a little bit and shake it till everything is dissolved. It takes time.
- add 0.93gr DTT.
Wait until it is completely dissolved.
Aliquote and keep frozen at-20ºC.
Reader Comments
Hi, can you please clarify the dilution ratio of 6xLaemmli buffer with protein sample? My understanding is 5:1 = Protein sample:buffer. Is that right?
thanks
Hi, Yes, you are totally right. 5ul of protein sample + 1ul of 6X Laemmli buffer. Hope it works!
Hi! I was trying to solve SDS, but it seems, that it don’t want to be solved
Please, any suggestions. Thanks in advance
Hi, you want to make 10ml 6X Laemmli buffer, but From your receipe, the total volume just 8ml. What should add more? Just whater? Thanks!
Hi Hui Chen, thanks for your comment. When you add the SDS, bromophenol and DTT, the volume increases to about 10ml. You won’t need to add more water.
Hello Lev, It takes a little bit to dissolve. What I normally do is to weight the SDS and put it together with the water in a flask, throw a magnet in it and leave it shaking on a stirring hot plate. Meanwhile I weight the rest of the things in the recipe and add them to the flask progressively. The plate doesn’t need to be very hot, just a litlle bit warm to help SDS dissolve. Don’t let it boil. I hope it works!