Sample Buffer, Laemmli 2× concentrate has been used as a sample buffer for denaturing and loading of protein samples in SDS-PAGE.
Laemmli sample buffer is especially formulated for protein sample preparation to be used in the Laemmli SDS-PAGE system. A protein sample is mixed with the 2X sample buffer (1:1) and heated in boiling water for 2-5 min. The 2-mercaptoethanol reduces the intra and inter-molecular disulfide bonds. The SDS detergent denatures the proteins and subunits and gives each an overall negative charge so that each will separate based on size. The bromophenol blue serves as a dye front that runs ahead of the proteins and also serves to make it easier to see the sample during loading. The glycerol increases the density of the sample so that it will layer in the sample well.
Solution contains 4% SDS, 20% glycerol, 10% 2-mercaptoethanol, 0.004% bromphenol blue and 0.125 M Tris HCl, pH approx. 6.8.
Recommended usage: add 1 volume gel loading solution to 1 volume sample and mix well.