When running SDS- Page on monoclonal antibodies I am noticing on the non-reduced samples, that the slight leading band that precedes the monomer band is presenting with equal intensity to the monomer band when using the NuPage 4X LDS sample buffer. When using the 2X SDS sample buffer, the monomer band presents as expected with the main band with a slight band beneath it. This leads me to believe that it has something to do with the dye interaction between the lithium/sodium in the sample buffer and the coomassie blue. I'm curious as to what about the LDS causes the bands to present in this manner?
These results have been compared to GXII profiles of the mAb in question.
Any thoughts or insight would be appreciated.
Carsten: I think the different intensities of the faster band, ie, stronger in LDS and weaker in SDS may be attributed to the fact that lithium is a stronger cation than sodium. Thus, LDS is a more potent detergent than SDS, and can have a potent denaturation effect on your mAb. If you still suspect an interaction of LDS or SDS with CB, then just run your sample in SDS buffer, wash away SDS in an SDS-free buffer, followed by LSD buffer prior to staining with CB. Good luck.
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