NuPAGE™ LDS Sample Buffer (4X), 10mL - FAQs

View additional product information for NuPAGE™ LDS Sample Buffer (4X) - FAQs (NP0008, NP0007)

18 product FAQs found

我已经使用NuPAGE LDS样品缓冲液制备了蛋白质样品。我能否煮沸样品？

可以煮沸蛋白质样品，因为煮沸不会损害缓冲液中的任何成分。我们之所以推荐将样品在70℃加热10分钟，而非煮沸样品，是因为较低的温度可减少蛋白质水解。

为什么NuPAGE样品缓冲液中使用LDS，而不是SDS？

NuPAGE 4X样品缓冲液的pH比普通Laemmli样品缓冲液高很多，它们的pH分别为8.5和6.8。在pH 8.5时，加入4X缓冲液所需浓度的SDS，会产生沉淀。此外，LDS（十二烷磺酸锂）更稳定，用于变性同样有效。

NuPAGE缓冲液应如何保存？

NuPAGE缓冲液应保存在4–25℃。

Can I prepare my protein sample with the reducing agent and store it for future use?

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

Find additional tips, troubleshooting help, and resources within our Protein Gel 1D Electrophoresis Support Center.

My LDS or SDS sample buffer precipitates when stored at 4 degrees C. Can I warm it up? Can I store it at room temperature?

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.

Find additional tips, troubleshooting help, and resources within our Protein Gel 1D Electrophoresis Support Center.

How are Bolt gels different than NuPAGE gels?

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.

Find additional tips, troubleshooting help, and resources within our Protein Gel 1D Electrophoresis Support Center.

What is the advantage of NuPAGE Gels over regular Tris-Glycine gels?

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.

Find additional tips, troubleshooting help, and resources within our Protein Gel 1D Electrophoresis Support Center.

What is the composition of the NuPAGE LDS Sample Buffer and what is the recipe for the 4X formulation?

The composition of the 1X NuPAGE LDS Sample Buffer is as follows:

141 mM Tris base
106 mM Tris HCl
2% LDS
10% Glycerol
0.51 mM EDTA
0.22 mM SERVA Blue G
0.175 mM Phenol Red
pH 8.5


To prepare 10 mL of 4 X NuPAGE LDS Sample Buffer, dissolve the following reagents to 8 mL ultrapure water:

Tris HCl 0.666 g
Tris Base 0.682 g
LDS 0.800 g
EDTA 0.006 g
Glycerol 4 g
SERVA Blue G (1 solution) 0.75 mL
Phenol Red (1 solution) 0.25 mL

Mix well and adjust the volume to 10 mL with ultrapure water. Store at +4. The buffer is stable for 6 months when stored at +4°C.

Find additional tips, troubleshooting help, and resources within our Protein Gel 1D Electrophoresis Support Center.

Can a protein sample be boiled in NuPAGE LDS Sample Buffer? Are there any components that may be destroyed by boiling?

You may boil the protein sample, although we recommend heating at 70°C for 10 min instead. The buffer components are not damaged during boiling. Heating rather than boiling will produce sharper protein bands, and this may be due to reduced protein hydrolysis at the lower temperature.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

What is the pH of the NuPAGE LDS Sample Buffer (4X) at 70°C?

The temperature coefficient (ΔpKa/degree C) for Tris is -0.031/degree C. Therefore, if the pH of the NuPAGE LDS Sample Buffer is 8.5 at 25°C, then its pH at 70°C is 8.5 + (-0.031 X 45) = 7.1. Similarly, our Tris Glycine Sample Buffer at 85°C would have a pH of 4.94, at 100°C its pH would be 4.5.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

Can beta-mercaptoethanol (BME) be used rather than DTT as the reducing agent in the NuPAGE LDS Sample Buffer?

Either BME or DTT can be used in the NuPAGE LDS Sample Buffer.

Make sure that a fresh solution of BME is used. FINAL concentration:

DTT 50-100 mM

BME 2-5%

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

What is the recipe for traditional Laemmli Buffer?

The Laemmli buffer or 2X SDS Buffer is composed of the following: 100 mM Tris HCl , pH 6.8, 200 mM dithiothreitol, 4% SDS, 0.2% bromophenol blue, 20% glycerol. 2X SDS gel loading buffer lacking dithiothreitol can be stored at room temperature. Dithiothreitol should then be added, just before use.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

Why is LDS (lithium dodecyl sulfate) used in the 4X NuPAGE sample buffer instead of SDS?

SDS in a 4X sample buffer concentrate tends to precipitate from solution and to make the solution viscous and difficult to pipette. The LDS is much more soluble.

Find additional tips, troubleshooting help, and resources within our Protein Gel 1D Electrophoresis Support Center.

Can I use CTAB rather than SDS in my sample buffer?

No, CTAB will not work with any of our gels except for the NuPAGE Tris-Acetate gels. To use CTAB, you would need to use a running buffer of 50 mM acetic acid and 50 mM beta-alanine in equal concentrations. You would also need to switch the electrodes. Since CTAB is a cationic detergent, this would establish conditions for running a basic protein towards the anode (into the gel).

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

Are the NuPAGE buffers/components (i.e., sample buffer, running buffer, and antioxidant) compatible with Bolt gels?

Yes, NuPAGE buffers/components are compatible with Bolt gels.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

I have prepared my protein sample using NuPAGE LDS Sample buffer. Can I boil my samples?

It would be okay to boil the protein sample as none of the buffer components are damaged upon boiling. The reason we recommend to heat the sample at 70 degrees C for 10 minutes and not boil the sample is because protein hydrolysis is reduced at lower temperatures.

Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.

Why is LDS used in the NuPAGE Sample buffer instead of SDS?

The pH of the NuPAGE 4X Sample buffer is much higher than that of the usual Laemmli Sample buffer: 8.5 vs. 6.8. At this pH (8.5), SDS will precipitate when added at the concentration needed for a 4X buffer. On the other hand, LDS (lithium dodecyl-sulfate) is more soluble and is just as effective for denaturation.

Find additional tips, troubleshooting help, and resources within our Protein Gel 1D Electrophoresis Support Center.

How should I store the NuPAGE buffers?

NuPAGE buffers can be stored at 4-25 degrees C.

Find additional tips, troubleshooting help, and resources within our Protein Gel 1D Electrophoresis Support Center.