Novex™ 4 to 12% Tris-Glycine Plus, 1.0 mm, Midi Protein Gels, 20-well, 10 gels (1 box) - FAQs

View additional product information for Novex™ Tris-Glycine Plus Midi Protein Gels, 4 to 12%, 1.0 mm - FAQs (WXP41212BOX, WXP41226BOXA, WXP41226BOX, WXP41220BOX, WXP41220BOXA, WXP41212BOXA)

42 product FAQs found

Can Tris-Glycine Plus Midi Gels be stored at room temperature?

No. They must be stored at 4 degrees C.

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Do Tris-Glycine Plus Midi Gels require any specific buffers?

Tris-Glycine Plus Midi Gels can use standard Tris-Glycine sample and running buffers. If you want to run under native conditions, we recommend using sample and running buffers that do not contain SDS (or our Native Tris-Glycine premade buffers).

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What in the chemistry changed to allow for a longer shelf life for Tris-Glycine Plus Midi Gels?

We were able to optimize shelf life of the gels through a proprietary gel formulation change. Unfortunately, we are unable to provide specific details on the chemical changes.

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What is the shelf life of Tris-Glycine Plus Midi Gels?

Tris-Glycine Plus Midi Gels have a shelf life of up to 1 year depending upon gel percentages. The minimum shelf life of these gels is at least 6 months from date of manufacture.

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Can Tris-Glycine Plus Midi Gels be run in the Bio-Rad Criterion gel running tank?

Yes, Tris-Glycine Plus Midi Gels can be run in the Bio-Rad Criterion tank with the help of adapters. If you are interested in running these gels in the Bio-Rad Criterion tank, we recommend buying the versions of these gels that come with adapters.

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With the launch of Tris-Glycine Plus Midi gels, will the current Tris Glycine Midi Gels be discontinued?

Yes, the current Tris-Glycine Midi Gels will be discontinued on December 31 2017.

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I stained Tris-Glycine gels with Colloidal Blue stain and noticed that the background was higher in low acrylamide percentage Tris-Glycine gels compared to high acrylamide percentage Tris-Glycine gels. Why is this and how can I resolve it?

Background is generally higher in gels with less than 10% acrylamide percentage due to penetration and trapping of colloids within the large pores of these gels. Excess background may be reduced by incubating the gel in 25% methanol solution until a clear background is obtained. Be aware that the dye will also be partially removed from the bands and that prolonged incubation in >25% methanol will result in complete destaining of protein bands and background.

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I used the Colloidal Blue staining kit for staining my Tris-Glycine gels and got very high background. Can you please offer some tips to reduce the background?

It may be possible to reduce background by using the protocol provided for NuPAGE Invitrogen Bis-Tris gels/Small Peptides. This protocol incorporates an extra fix step to remove excess SDS, which can act as an anti-colloidal agent and lead to higher background. The low pH of the staining solution will fix the gel, but not as fast as the pre-fix step specified in the NuPAGE protocol.

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I have set up my Tris-Glycine 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.

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I am running my Tris-Glycine 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

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I am trying to load my samples on a Tris-Glycine 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.

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I am running my Tris-Glycine 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 volatage

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I ran my protein samples on a Tris-Glycine gel and the samples stopped running in the bottom portion of the gel whereas the marker ran fine. Why did this happen?

This can happen if Tris-Glycine gels are run using NuPAGE Running buffer containing Antioxidant. Please make sure that the correct Tris-Glycine Running buffer is used with Tris-Glycine gels.

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The protein bands in some of my gel lanes are irregular or wavy? What would have caused this problem?

This could be due to:

*Debris in the well
*High salt in the sample (make sure that the salt concentration does not exceed 50-100 mM)
*Running buffer issue
*Gel casting error

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I am seeing a very wavy and uneven dye front with my samples. Can you please help me troubleshoot?

This could be due to a gel polymerization issue combined with incorrect sample preparation (final sample dilution less than 1X). Please try a different lot of the same gel and make sure that the sample is correctly prepared.

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I am seeing a faint, artifact doublet band at ~60 kDa in all my lanes. This band seems to be getting darker the longer I stain the gel. What could be causing this?

Possible cause:

*Excess reducing agent (beta-mercaptoethanol)
*Skin protein contaminants (keratin)

Remedy:

*The addition of iodoacetamide to the equilibration buffer just before applying the sample to the gel has been shown to eliminate these artifact bands.
*Use new electrophoretic solutions and wear gloves when handling and loading the gel. This issue is more common when highly sensitive stains are used.

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I loaded different protein samples in each well but I see the same protein band in several neighboring lanes. What could have happened?

Possible cause:

*Carry-over contamination of sample from one well into neighboring wells due to loading error
*Contaminated running buffer
*Gel casting error: malformed wells

Remedy:

*Use a gel loading tip to load wells
*Reduce the sample volume
*Do not delay while loading wells
*Do not delay after the run, as proteins can diffuse horizontally; a full well left next to an empty well would eventually contaminate the empty well over time.

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My protein bands appear to be skewed or distorted. What is the problem?

Possible cause:

*Poor polymerization around sample wells
*High salt concentration in sample
*Uneven gel interface
*Excessive pressure applied to the gel plates when the gel is placed into the clamp assembly
*Uneven heating of the gel
*Insoluble material in the gel or inconsistent pore size throughout the gel
*Air bubble during the run

Remedy:

*Remove excess salt/other material by dialysis, Sephadex G-25 or any other desalting column or using an Amicon concentrator.
*Either use a cooled apparatus or reduce the current at which electrophoresis is performed.

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My gel seems to be lifting off the cassette. What could be causing this?

Gel lifting off the cassette can be caused by:

*Expired gels that are degrading
*Improper storage of gels
*Too much heat accumulating during the electrophoresis run due to excessive current
*Insufficient polymerization of the polyacrylamide

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I am seeing a faint shadow, or "ghost" band below a normal and expected protein band? What could be the potential issue?

Ghost bands are usually attributed to a slight lifting of the gel from the cassette, which results in the trickling down of some sample beyond its normal migration point. It then accumulates and appears as a faint second band.

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My protein bands in the outer lanes of the gel show a "smiling" effect. Can you please help me troubleshoot?

"Smiling" bands may be the result of the acrylamide in the gel breaking down, leaving less of a matrix for the proteins to migrate. We recommend checking to ensure that the gels have not been used past their expiration date.

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I see dumbbell or barbell shaped bands after protein electrophoresis. What could be causing this?

Barbell shaped bands are a result of loading too large of 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 incompletely stack causing a slight retardation of the portion of the sample that diffused to the sides of the wells. This effect may be intensified for larger proteins, whose migration is more impeded in the low concentration acrylamide of the stacking gel. To alleviate the problem, we recommend concentrating the protein and loading a smaller volume. This gives a "thinner" starting zone.

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Why do I get streaking forward or "frowning" of one of my samples on my protein gel?

Here are possible causes and solutions:

1) Sample overload: Do not overload samples
2) Addition of reducing agent that is not fresh: Reduce samples right before loading and do not use samples that have been stored in reducing agent
3)Re-oxidation of the protein during the run: Add antioxidant to the running buffer if you are running NuPAGE gels
4) Presence of highly hydrophobic regions where the protein can exclude SDS: Load the sample with 2X sample buffer instead of 1X sample buffer
5) Excess salt in the sample: Precipitate and reconstitute in lower salt buffer
6) Not enough SDS in the sample: Add SDS to the upper buffer chamber (try 0.1%, 0.2%, 0.3% and 0.4% SDS)

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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

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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.

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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).

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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.

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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.

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What is the material used for making your Invitrogen precast gel plastic cassettes?

The cassettes are made of a styrene copolymer.

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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.

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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.

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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.

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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.

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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.

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Which protein standard do you recommend using with gels run under native conditions?

We recommend using the NativeMark Unstained Protein Standard, Cat. No. LC0725.

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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.

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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.

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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.

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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.

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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.

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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

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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.

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