NuPAGE™ Transfer Buffer (20X)

DTT is not stable, so it must be added and the reduction performed just prior to loading your samples.

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Precipitation of the LDS or SDS at 4 degrees C is normal. Bring the buffer to room temperature and mix until the LDS/SDS goes into solution. If you do not want to wait for it to dissolve, you can store the sample buffer at room temperature.

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While they are both Bis-Tris based gels, the chemistries are very different since Bolt gels are optimized for western blotting. Another key difference is the wedge well design of the Bolt gels, which allows larger sample volumes to be loaded.

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The neutral operating pH of the NuPAGE Gels and buffers provides following advantages over the Laemmli system:
-Longer shelf life of 8-12 months due to improved gel stability
-Improved protein stability during electrophoresis at neutral pH resulting in sharper band resolution and accurate results (Moos et al, 1998)
-Complete reduction of disulfides under mild heating conditions (70 degrees C for 10 min) and absence of cleavage of asp-pro bonds using the NuPAGE LDS Sample buffer (pH > 7.0 at 70 degrees C)
-Reduced state of the proteins maintained during electrophoresis and blotting of the proteins by the NuPAGE Antioxidant
Please refer to the following paper: Moos M Jr, Nguyen NY, Liu TY (1988) Reproducible High Yield Sequencing of Proteins Electrophoretically Separated and Transferred to an Inert Support. J Biol Chem 263:6005-6008.

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Yes. The NuPAGE transfer buffer works with Tris-Glycine gels. Proteins are subjected to a more neutral pH, and the absence of glycine in the NuPAGE transfer buffer makes protein sequencing of proteins extracted from the gels much easier. While protein transfer is generally more efficient when using the NuPAGE transfer buffer, remember that the other benefits of the NuPAGE system (band sharpness, better resolution, sample stability) only apply if the proteins have been separated under these conditions prior to transfer (i.e., on a NuPAGE gel).

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