6X Laemmli buffer

This post was written by admin on March 5, 2009
Posted Under: Buffers & recipes, Western blot

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

#1 
Written By tengku on August 24th, 2009 @ 4:46 am

Hi, Yes, you are totally right. 5ul of protein sample + 1ul of 6X Laemmli buffer. Hope it works!

#2 
Written By admin on August 24th, 2009 @ 10:27 am

Hi! I was trying to solve SDS, but it seems, that it don’t want to be solved :) Please, any suggestions. Thanks in advance

#3 
Written By Lev on September 17th, 2009 @ 5:59 am

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!

#4 
Written By hui chen on September 17th, 2009 @ 9:32 am

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.

#5 
Written By admin on September 17th, 2009 @ 11:22 am

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!

#6 
Written By admin on September 17th, 2009 @ 11:26 am

Add a Comment

required, use real name
required, will not be published
optional, your blog address