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.
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.
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.
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.
What is the maximum sample volume and concentration that may be loaded into the Invitrogen precast gel wells?
1-well, 1 mm: 700 µL (no more than 12 µg protein or 2.4 µg DNA per band)
2D-well, 1 mm: 400 µL or 1.5 mm: 600 µµL (no more than 12 µg protein or 2.0 µg DNA per band)
5-well, 1 mm: 60 µL (no more than 2 µg protein or 400 ng µg DNA per band)
9-well, 1 mm: 28 µL (no more than 0.5 µg protein or 100 ng DNA per band)
10-well, 1 mm: 25 µL or 1.5 mm: 37 µL (no more than 0.5 µg protein or 100 ng DNA per band)
12-well, 1 mm: 20 µL (no more than 0.5 µg protein or 100 ng DNA per band)
15-well, 1 mm: 15 µL or 1.5 mm: 25 µL (no more than 0.5 µg protein or 100 ng DNA per band)
17-well 1 mm: 15 µL (no more than 0.5 µg protein per band)
What percentage of stacking gel is in Invitrogen Tris-Glycine and NuPAGE Invitrogen gels? What is the length of stacking gel?
The stacking gel acrylamide concentration for Tris-Glycine and NuPAGE Bis-Tris gels is 4%. For NuPAGE Tris-Acetate gels it is 3.2%.
The stacking portion is approximately 8 to 9 mm long (it ends right above the first ridge on the cassette) on most Invitrogen protein gels (i.e., Tris-Glycine and NuPAGE gels).
What causes dumbbell- or barbell-shaped bands during protein electrophoresis?
Barbell-shaped bands are a result of loading too large a sample volume.
When a large sample volume is loaded, part of the sample tends to diffuse to the sides of the wells. When the run begins and the sample moves through the stacking portion of the gel, the sample will stack incompletely, causing a slight retardation of the portion of the sample that diffused to the sides of the wells.
This effect may be intensified in larger proteins, whose migration is more impeded in the low concentration acrylamide of the stacking gel.
To alleviate the problem, concentrate the protein and load a smaller volume. This gives a "thinner" starting zone.
What can cause "streaking forward" or "frowning" of samples on a SDS-PAGE gel? How can the results be improved?
Some potential causes are:
1) Re-oxidation of protein during run
2) Protein has highly hydrophobic regions where protein can exclude SDS.
Steps you can take to improve results:
1) Reduce samples right before loading, and add antioxidant to running buffer. Do not use samples that have been stored in reducing agent.
2) Load sample with 2X sample buffer instead of 1X.
3) Add SDS to upper chamber buffer: try 0.1, 0.2, 0.3, and 0.4% (don't go any higher than 0.4%)
Will NP-40 affect the migration of the samples in the SDS-PAGE gel?
Yes. All detergents and even phospholipids in cell extracts will form mixed micelles with SDS and migrate down into the gel.
They can also interfere with the SDS:protein binding equilibrium. Most of the nonionic detergents significantly interfere with SDS-PAGE.
We recommend that you keep the ratio of SDS to lipid or other detergent at 10:1 (or greater) to minimize these effects.
Will acetonitrile in my sample affect my electrophoresis run?
There shouldn't be any negative effects unless the percentage of acetonitrile reaches 40% or 50% of the sample volume.
At these concentrations, there is the possibility of the acetonitrile affecting the binding of SDS to the protein, which, in turns, affects the migration of the protein.
What is the concentration of SDS in Invitrogen gels?
There is no SDS in the gels. Denaturing conditions are created by using sample buffers and running buffers that contain SDS.
The benefit of not having SDS in the gels is that the gel can be used for both native and denaturing conditions.
Is it possible to run reduced and non-reduced samples on the same NuPAGE gel? Should the Antioxidant be used?
If the Antioxidant is omitted from the running buffer, it is possible to resolve reduced and non-reduced samples on the same gel, although the resolution may be lower. Furthermore, it is not recommended that the reduced and non-reduced samples be run side-by-side in adjacent lanes.
However, because of the neutral pH of the NuPAGE gels, the reducing agent (beta-mercaptoethanol or DTT) will not migrate through the gel with the protein the way it does in the basic environment of the Tris-Glycine gels. Instead, the reducing agent tends to remain at the top of the gel. For this reason, the NuPAGE Antioxidant is incorporated into the buffer in the upper buffer chamber. The antioxidant is able to migrate fully with the proteins and keep them reduced. As a result, it is possible that proteins prepared as non-reduced samples could become somewhat reduced during the electrophoresis run. This would result in smearing of the samples.
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).
Do you have a protocol for extracting proteins from polyacrylamide gels?
Do you have recommended protocols for copper or zinc staining of NuPAGE gels?
Copper or zinc staining is a rapid, sensitive method for detection of protein bands . ~10 ng reduced BSA on NuPAGE Bis-Tris gels can be detected with both the copper and zinc stain.
Copper Stain: Staining solution - 0.3 M CuCl2
After electrophoresis, remove the gel from the cassette and equilibrate the gel in 100 mL of 1X running buffer* for 15 minutes. Immerse the gel in 100 ml of 0.3 M CuCl2 solution for about 5 minutes (the protein band will appear as a negative stain with a blue background).
Zinc stain: Staining solution - 0.2 M Imidazole and 10 mM ZnCl2
After electrophoresis, remove the gel from the cassette and equilibrate the gel in 100 mL of 1X running buffer* for 15 minutes. Place the gel in 100 ml of 0.2M Imidazole solution for 10 minutes. Next immerse the gel in 100 ml of 10 mM ZnCl2 solution for about 5 minutes (the protein will appear as a transparent band with a white background).
*The 1X running buffer can be the buffer from the electrophoresis tank after run (MES, MOPS). However, for better contrast of the band, the 1X Tris-Glycine SDS running buffer is recommended.
Is it okay to use protein gels past their expiration date?
We do not recommend using gels past their expiration date because over time, the polyacrylamide hydrolyzes to acrylic acid and ammonia and this will affect the resolution of the proteins. Breakdown of polyacrylamide matrix is identified by:
- Ghost bands and doublets, seen first in the high molecular weight proteins
- Smiling of dye front across the gel, with bands in outer lanes becoming very slanted - proteins run slower there due to change in pH and pore size over time.
Can I run your protein gels overnight?
This is not really recommended, but it is always possible to increase run time by lowering the voltage of the run. In general, the relationships are linear - i.e., decreasing voltage by half will double the run time.
Do I need to increase the voltage when I run a 1.5 mm protein gel versus a 1.0 mm gel?
If you are running the gel at constant voltage, you do not need to increase the voltage regardless of the thickness of the gel.
Why do you recommend running your protein gels at constant voltage?
Using constant voltage allows the current and power to decrease during the run, providing a safety margin in case of a break in the system. Having lower power is a safety benefit and will also decrease the chances of experiencing an overheating of the gel. Further, the constant voltage setting does not need adjustment to account for differences in number or thickness of gels being run.
I ran my protein under native conditions on a Tris-Glycine gel. It has a pI that is higher than the pH of the Tris-Glycine transfer buffer. Can you offer some tips for transferring it?
- Increase the pH of Tris-Glycine transfer buffer to 9.2, allowing all the proteins below pI 9.2 to transfer towards the anode electrode.
- Use the Tris-Glycine transfer buffer and place a membrane on both sides of the gel. If there are any proteins that are more basic than the pH of the transfer buffer, they will be captured on the extra membrane placed on the cathode side of the gel. Both membranes can then be developed in the same manner.
- Prior to blotting, incubate the gel for 15 minutes in Tris-Glycine transfer buffer containing 0.1% SDS. The small amount of SDS will give the proteins enough charge to move unidirectionally towards the anode and in most cases, should not denature the protein. Proceed with the transfer using regular Tris-Glycine transfer buffer.
Do I need to add the chlorobutanol when making the 20X NuPAGE Transfer Buffer?
Chlorobutanol is used as a preservative in the NuPAGE transfer buffer and is not necessary for efficient transfer of proteins. You may prepare the buffer without chlorobutanol but keep in mind that the buffer will not be stable for long periods. We recommend using it within 2 weeks.
Can I transfer NuPAGE gels using Carbonate or CAPS transfer buffers?
We do not recommend using Carbonate or CAPS transfer buffers to transfer NuPAGE gels as the transfer efficiency will be badly compromised. Further, the high pH environment (>pH 9) of these buffers will make the NuPAGE Antioxidant non-functional.
Do you have any tips to improve transfer of high molecular weight proteins from NuPAGE gels?
To increase efficiency of transfer of high molecular weight proteins from NuPAGE gels, we recommend pre-equilibrating the gel in 2x NuPAGE Transfer buffer (without methanol) containing 0.02-0.04% SDS for 10 minutes before assembling the sandwich and then transferring using 1x NuPAGE transfer buffer containing methanol and 0.01% SDS.
My gel run is faster than normal with poor resolution. What could be causing this problem?
Here are possible causes and solutions:
- Buffers are too concentrated or incorrect. Check buffer recipe; dilute or re-make if necessary.
- Voltage, current or wattage is set at a higher limit. Decrease power conditions to recommended running conditions.
My gel will not run. Can you please help me troubleshoot?
It is important to determine whether the problem is with the power supply, the apparatus or the gel. Often, it helps to switch out the power supply or the lid to see if there is a faulty contact. Also, check to see whether the tape from the bottom of the gel cassette has been removed and whether the buffer core is damaged. Additionally, make sure there is sufficient buffer in the electrophoresis tank to cover the wells of the gel.
Which of your protein gels can I run using the Mini Gel Tank?
Our New Bolt Bis-Tris Plus Mini gels (Cat. No. NWxxxxxBOX), as well as our Invitrogen Mini gels and NuPAGE Mini gels can be run using the Mini Gel Tank. Please note that our original Bolt Bis-Tris Plus Mini gels (Cat. No. BGxxxxxBOX, discontinued as of December 31, 2014) can only be run in the Bolt Mini Gel Tank (discontinued as of December 31, 2014, and will be offered until inventory is depleted).
Which of your protein gels can I run using the XCell SureLock Mini-Cell?
Our New Bolt Bis-Tris Plus Mini gels (Cat. No. NWxxxxxBOX), as well as our Invitrogen Mini gels and NuPAGE Mini gels can be run using the XCell SureLock Mini-Cell.
I have set up my NuPAGE gel to run and have switched on the power supply, but my gel is not running and there is no voltage or current reading on the power supply. What is wrong?
*Double check that the tape on the bottom of the gel has been removed.
*Make sure that the gel(s) are oriented so that the taller sides of the cassette (with the printing) are facing the outside of the electrophoresis unit.
*Make sure that the inner buffer chamber is filled sufficiently so that the wells are covered with buffer. If the wells are not covered, check for leaks and reseal.
*Double check to see if there are any loose electrodes or connections on the Mini cell unit.
*Check the power supply unit.
I am running my NuPAGE gel using the XCell SureLock Mini Cell but the leads do not fit into my power supply. Can you please help?
You may purchase the ZOOM adapters, Cat. No. ZA10001 to help you connect your leads to the power supply.
I am trying to load my samples on a NuPAGE gel but am not able to see the sample wells. Can you please suggest some tips?
We recommend marking the cassette at the bottom of the wells with a marker pen prior to assembling the upper buffer chamber. Also, we recommend illuminating the bench area with a light source placed directly behind the XCell SureLock unit.
I am running my NuPAGE gel using the XCell SureLock Mini Cell, and the gel run is taking longer than usual. What could be causing this?
Here are possible causes and solutions:
1) Buffers are too dilute: Check buffer recipe; remake if necessary.
2) Upper buffer chamber is leaking: Make sure the buffer core is firmly seated, the gaskets are in place and the gel tension lever is locked.
3) Voltage is set too low: Set correct voltage.
I had problems transferring my larger molecular weight proteins from my NuPAGE gel. Can you please offer some suggestions?
For proteins larger than 100 kDa, we recommend pre-equilibrating the gel in 2X Transfer buffer (without methanol) containing 0.02-0.04% SDS for 10 minutes before assembling the sandwich and then transferring using 1X transfer buffer containing methanol and 0.01% SDS.
Can I use beta-mercaptoethanol instead of the NuPAGE Sample Reducing agent?
Although we recommend using the NuPAGE Sample Reducing agent for stability reasons, fresh, neat beta-mercaptoethanol can be substituted for the NuPAGE Sample Reducing Agent, with equivalent results. A final concentration of 2-5% beta-mercaptoethanol is usually sufficient to reduce the sample.
How much methanol do you recommend adding to the NuPAGE transfer buffer for transfer of NuPAGE Bis-Tris gels and NuPAGE Tris-Acetate gels?
We recommend adding 10% methanol to the NuPAGE transfer buffer for transfer of one gel and 20% methanol for the transfer of 2 gels.
Do you recommend adding the NuPAGE Antioxidant to the NuPAGE transfer buffer when I transfer proteins from NuPAGE Bis-Tris or NuPAGE Tris-Acetate gels?
Yes, we recommend adding the NuPAGE Antioxidant to the NuPAGE transfer buffer for enhanced blotting results with reduced proteins in order to maintain the reduced state of the proteins throughout the run.
What stains do you recommend for NuPAGE gels?
NuPAGE gels are compatible with any of the standard Coomassie staining procedures. The protocols that are accelerated by heat are preferable as the heat serves as a “fix” for proteins, especially smaller peptides. NuPAGE gels are also compatible with most silver staining protocols. They are also compatible with copper or zinc staining, and fluorescent stains.
Can I use NuPAGE Bis-Tris gels with NuPAGE MOPS or MES Running Buffer prepared without SDS for electrophoresis under native conditions?
We do not recommend using NuPAGE Bis-Tris gels with NuPAGE MOPS or MES Running Buffer prepared without SDS for electrophoresis under native conditions. This buffer system may generate excessive heat, resulting in poor band resolution. Further, the protein of interest may not migrate very well in a neutral pH environment if it is not charged.
How are Bolt Bis-Tris Plus gels different from NuPAGE Bis-Tris gels?
While they are both Bis-Tris based gels, the chemistries are very different as Bolt Bis-Tris Plus gels have been optimized for western blotting. Another key difference is the enabling wedge well design for Bolt Bis-Tris Plus gels, which allows larger sample volume load.
What are the main advantages of NuPAGE gels over Invitrogen Tris-Glycine gels?
NuPAGE gels have the following advantages over Tris-Glycine gels:
*Higher stability and longer shelf life: NuPAGE Bis-Tris gels and NuPAGE Tris-Acetate gels have a lower operating pH (pH 7 for NuPAGE Bis-Tris gels and pH 8.1 for NuPAGE Tris-Acetate gels) than Invitrogen Tris-Glycine gels (pH 9.5). At basic pH, polyacrylamide hydrolyzes to polyacrylic acid and ammonia whereas at neutral pH, this hydrolysis is slower. Hence, NuPAGE gels have higher stability and longer shelf life than Invitrogen Tris-Glycine gels (12 months at 4-25 degrees C for NuPAGE Bis-Tris gels and 8 months at 4 degrees C for NuPAGE Tris-Acetate gels vs 4-8 weeks at 4 degrees C for Tris-Glycine gels).
*Better resolution of proteins due to:
- Reduced undesired chemical modifications: Free acrylamide alkylates proteins at basic pH (8.5 to 9.0). It targets sulfhydryl cysteines and amine groups at the N-terminus and on lysines. This modification does not happen at pH below 8. Hence, proteins run on NuPAGE gels undergo fewer of these undesired chemical modifications than those run on Tris-Glycine gels.
- Reduced hydrolysis of proteins: Heating of Tris-Glycine sample buffer (pH 6.8) results in a drop in pH, causing Asp-Pro cleavage of proteins. High temperature and longer duration of heating/boiling increase the rate of this cleavage resulting in multiple peptide bands of decreased intensity. At 100 degrees C, the pH drops as low as pH 4.3. On the other hand, NuPAGE LDS sample buffer (pH 8.5) drops to pH 8.1 when heated to 70 degrees C, avoiding this cleavage.
*Faster run times: 35-50 min for NuPAGE Bis-Tris gels and 1 hour for NuPAGE Tris-Acetate gels vs 90 min for Tris-Glycine gels
How does the operating pH for Tris-Glycine gels differ from that for NuPAGE Bis Tris and NuPAGE Tris-Acetate gels?
The operating pH for Tris-Glycine gels is 9.5; the operating pH for NuPAGE Bis-Tris gels is 7 and for NuPAGE Tris-Acetate gels is 8.1.
Can I run Mini gels with 10 cm gel cassettes using a Bolt Mini Gel Tank?
To run Mini gels with 10 cm gel cassettes using a Bolt Mini Gel Tank (without replacement of 10.5 cm cassette clamp cam handles with 10 cm cassette clamp cam handles), please use the instructions provided on Page 22 of the manual (https://tools.thermofisher.com/content/sfs/manuals/mini_gel_tank_man.pdf).
Note: For optimal results, to run 10 cm cassette Mini gels with a Bolt Mini Gel Tank, one should replace the black 10.5 cm cassette clamp cam handles on the Bolt Mini Gel Tank with gray 10 cm cassette clamp cam handles (Cat. No. A26732). Instructions for replacement of the cam handles can be found on Page 20 of the manual (http://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-gel-electrophoresis/protein-gel-electrophoresis-chamber-systems/mini-gel-tank/resources-upgrading-bolt-mini-gel-tank.html) or in this video (https://www.youtube.com/watch?v=1FtiX8Skllw).
Additional resources can be found here (https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-gel-electrophoresis/protein-gel-electrophoresis-chamber-systems/mini-gel-tank/resources-upgrading-bolt-mini-gel-tank.html).
How do you recommend transferring Midi gels?
Midi gels can be transferred using:
*iBlot Dry Blotting System in conjunction with Transfer Stacks
*Invitrogen Semi-Dry Blotter for simultaneous transfer of up to 2 Midi-gels
*Thermo Scientific Power Blotter for simultaneous transfer of up to 2 Midi gels
*Thermo Scientific G2 Fast Blotter (will be discontinued as soon as we exhaust current inventory).
Will NP-40 affect the migration of my protein samples?
All detergents, or even phospholipids in cell extracts, will form mixed micelles with SDS and migrate down into the gel. They can also interfere with the SDS:protein binding equilibrium. Most of the non-ionic detergents, including NP-40, are the worst at interfering with SDS-PAGE. The rule of thumb is to keep the ratio of SDS to lipid or other detergent at 10:1 or greater to minimize these effects.
Do your Invitrogen protein gels contain any carbohydrates and are they suitable for carbohydrate analysis?
All Invitrogen protein gels contain sucrose as a density-adjusting agent to facilitate pouring of the gel. Protein samples run on Invitrogen gels would be contaminated with large amounts of sucrose. Thus, Invitrogen gels are not recommended for this application.
What is the material used for making your Invitrogen precast gel plastic cassettes?
The cassettes are made of a styrene copolymer.
Can I recycle your Invitrogen precast gel plastic cassettes?
We do not recommend recycling our plastic cassettes because they have a chemical coating on them that may produce toxic fumes when melted and potentially cause contamination.
What is the difference between Invitrogen Mini and Midi gel formats?
Midi gels are wider than Mini gels and hence have a larger number of wells to accommodate additional samples in one gel. An experiment from a Mini gel can be easily scaled-up to a Midi gel of the same gel chemistry.
Midi gels:
*NuPAGE Bis-Tris, NuPAGE Tris-Acetate, & Invitrogen Tris-Glycine: Gel dimensions are 13cm x 8.3cm and Cassette dimensions are 15cm x 10.3cm.
Mini gels:
*NuPAGE Bis-Tris, NuPAGE Tris-Acetate, & Invitrogen Tris-Glycine: Gel dimensions are 8cm x 8cm and Cassette dimensions are 10cm x 10cm.
*New Bolt Bis-Tris Plus (Cat. No. NWxxxxxBOX): Gel dimensions are 8cm x 8.3cm and Cassette Dimensions are 10cm x10cm.
*Original Bolt Bis-Tris Plus (Cat. No. BGxxxxxBOX): Gel dimensions are 8cm x 8.3cm and Cassette Dimensions are 10cm x 10.5cm.
What are the dimensions of your precast protein gels?
All of our Invitrogen precast protein gels (NuPAGE gels, Bolt Bis-Tris Plus gels, and Novex gels) are available in Mini format. Our Mini gel dimensions are 8 cm x 8 cm and the cassette dimensions are 10 cm x 10 cm.
Our NuPAGE Bis-Tris, NuPAGE Tris-Acetate, and Novex Tris-Glycine Plus gels are also available in the wider Midi format. Our Midi gel dimensions are 8 cm x 13 cm and the cassette dimensions are 10 cm x 15 cm.
Are your precast protein gels available in Mini and Midi formats?
All our Invitrogen protein gels are available in Mini format. Certain gel chemistries (NuPAGE Bis-Tris, NuPAGE Tris-Acetate, and Invitrogen Tris-Glycine gels) are also available in the wide Midi format.
Note that Bolt Bis-Tris gels are not available in the Midi format and our Thermo Scientific Precise precast gels are only available in Mini format.
When running two protein gels, do I need to double the voltage?
If you are running the gels at constant voltage, you do not need to increase the voltage regardless of the number of gels. However, the resulting current and wattage observed will multiply linearly with the number of gels. Keep in mind that the expected total current for your gels should not exceed the current limit of the power supply, or else the current will plateau and the run will slow down. (For example: Recommended constant voltage for running a NuPAGE Bis-Tris gel with MES Buffer is 200 V, with a starting current of 110-125 mA/gel and end current of 70-80 mA/gel. If the power supply has a current limit of 500 mA, the maximum number of NuPAGE Bis-Tris gels that can be run at one time with full power is 500 mA/125 mA = 4 gels. Any additional gels will decrease the current per gel and increase the run time.
Can I run reduced and non-reduced protein samples on the same gel?
We do not recommend running reduced and non-reduced protein samples on the same gel, especially in adjacent lanes, since the reducing agent may have a carry-over effect on the non-reduced samples if they are in close proximity.
Can I store my reduced protein samples for later use?
We do not recommend storing reduced protein samples for long periods of time even if they are frozen because reoxidation of the sample may happen during storage, causing inconsistent results.
What is the ratio of acrylamide:bisacrylamide and percentage of cross-linker in your Invitrogen precast gels?
*Tris-Glycine gels (except 4% Tris-Glycine gels) have a 34.5:1 Acrylamide:bisacrylamide and 2.6% Crosslinker.
*4% Tris-Glycine gels have a 76:1 ratio Acrylamide:bisacrylamide and 1.3% Crosslinker.
What is the percentage of the stacking gel in your Invitrogen precast protein gels?
The percentage of the stacking gel is 4% in most of our gels including the Bolt Bis-Tris Plus gels. The NuPAGE Tris-Acetate gels contain a 3.2% stacking gel.
Do your Invitrogen precast protein gels contain a stacking gel?
Our Invitrogen precast protein gels contain a stacking gel that is ~8 to 9 mm long (it ends right above the first ridge on the cassette). The manufacturing method used results in an interface between the stacking and resolving gels that is not visually detectable.
What are the recommended sample loading volumes and protein loading amounts for your precast protein gels?
*Tris-Glycine and Invitrogen Tricine Mini gels: see here (http://tools.thermofisher.com/content/sfs/manuals/electrophoresisguide_man.pdf), Page 8
*NuPAGE Tris-Acetate and NuPAGE Bis-Tris Mini gels: see here (http://tools.thermofisher.com/content/sfs/manuals/nupage_tech_man.pdf), Page 10
*Bolt Bis-Tris Plus Mini gels: see here (http://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-gel-electrophoresis/protein-gels/bolt-bis-tris-gels.html)
*Thermo Scientific Precise Tris-HEPES gels: see here (https://tools.thermofisher.com/content/sfs/manuals/MAN0011499_Precise_Protein_Gels_UG.pdf), Page 1
*Midi gels (Invitrogen Tris-Glycine, NuPAGE Bis-Tris and NuPAGE Tris-Acetate): see here (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/novex_midigel_man.pdf), Page 4
*Thermo Scientific Precise Tris-Glycine gels: see here (https://tools.thermofisher.com/content/sfs/manuals/D25MAN0011814_Precise_TrisGlycine_Gels_UG.pdf), Page 1
Do your precast protein gels contain SDS?
Our precast protein gels do not contain SDS but they can be run under denaturing conditions when used with the appropriate denaturing running buffer.
Note: NuPAGE Bis-Tris gels, Bolt Bis-Tris Plus gels, and Thermo Scientific Precise Tris-HEPES gels cannot be run under native conditions; they can only be run under denaturing conditions.
*Invitrogen Tris-Glycine gels: For Native electrophoresis, use Invitrogen Tris-Glycine Native Running Buffer. For Denaturing electrophoresis, use Invitrogen Tris-Glycine SDS Running Buffer
*NuPAGE Tris-Acetate gels: For Native electrophoresis, use Invitrogen Tris-Glycine Native Running Buffer. For Denaturing electrophoresis, use NuPAGE Tris-Acetate SDS Running Buffer
*NuPAGE Bis-Tris gels: For Denaturing electrophoresis, use NuPAGE MOPS-SDS Running Buffer or NuPAGE MES-SDS Running Buffer
*Bolt Bis-Tris Plus gels: For Denaturing electrophoresis, use Bolt MOPS SDS Running Buffer or Bolt MES SDS Running Buffer
*Thermo Scientific Precise Tris-Glycine gels: For Native electrophoresis, use Tris-Glycine SDS Running Buffer without SDS added. For Denaturing electrophoresis, use Tris-Glycine SDS Running Buffer.
*Thermo Scientific Precise Tris-HEPES gels: For Denaturing electrophoresis, use Tris-HEPES SDS Running Buffer.
