SDS PAGE: sample buffer 2-ME or DTT?? - (Oct/29/2009 )
one of recent observation in our lab:-we compared sample buffer(reducing) one having "beta-mercaptoethanol"(Sample buffer I) other with "DTT" (Sample Buffer II) as reducing agent.
all samples made prepared with Sample Buffer I (I.e. one with beta metcaptoethanol) picked up silver staining much better than smples prepared with sample buffer II
was wondering why is this effect?
which one is better reducing agent to be used in sample buffer???
DTT is a better reducing agent as it has 2 thiol groups (one more than BME). Both can be used in the sample buffers. BME has to be used more than DTT for the same action.
As far as picking of the silver stain concerns... i have not observed this but i have observed streaking of DTT present samples. The reason may be because DTT is more sensitive to storage conditions than is BME. DTT is generally stored in 4 degrees and the sample buffer should be stored in -20 or atleast should be frozen!!!
These are my obsevations. Any more suggestions or corrections are most welcome!!!