I want to do MALDI-TOF mass for my protein. For its procedure I need to do bis tris electrophoresis of immunoprecipitated protein, then excise protein band. For running bis tris electrophoresis, I need to solve my protein in LDS sample buffer, but I think that would i replace SDS with LDS in this buffer? Is it too different for doing mass spect?
As far as I know SDS is subjected to precipitation more than LDS. So maybe you would see some undisolved particles in case of SDS.
Some people really think LDS is necessary, but it is not. We call the procedure Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE), not LDS-PAGE.
What you need in the loading dye is either SDS or LDS to denature your protein and give it a negative charge, glycerol to sink your samples into the well, DTT to reduce potential disulfide bonds in your protein, and a dye such as bromophenol blue to track migration of your protein into the gel.
Also, if your protein is reasonably pure, you can submit your immunoprecipitated protein in solution for Mass spectrometry analysis. Excising your protein from the gel is not necessary, neither is adding SDS or LDS.
thanks Adron Ung,
my sample isn't pure, so I have to excise the band of protein on bis tris gel. A suitable sample buffer for this type of gel is LDS buffer. Now I want to khow that can i replace SDS to LDS in this buffer?
LDS is used instead of SDS at low temperatures, as it does not precipitate (Delepelaire & Chua 1979).
You may replace LDS and SDS for PAGE. It is not applicable for mass spec whether lithium or sodium salt. Li-cor uses LDS in PAGE products and applications they recommend LDS instead of SDS due to enhanced solubility but also more applicable for plant proteomic research. For MS getting rid of all types of dodecyl salts and in-gel digestion by applying wash replicates after excising the band would be an efficient protocol.
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