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View additional product information for Mini Blot Module - FAQs (B1000)
30 product FAQs found
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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Similar to NuPAGE gels with storage temperatures of 4 to 25 degrees C.
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The shelf life for Bolt gels is similar to that for NuPAGE gels: 12 months from the date of shipment.
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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 most common power supplies from Thermo Fisher Scientific, Bio-Rad, and Hoefer are compatible. Also, power supply adapters are available for power supplies not designed for use with covered or non-retractable power leads. Thermo Fisher Owl branded gel tanks are not compatible
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Yes. While we would prefer that you use our devices, Bolt gels can also be transferred using devices from Bio-Rad, including the Mini Trans-Blot Cell, Trans-Blot SD Semi-Dry Transfer Cell, or Trans-Blot Turbo Transfer System.
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Yes. You can use the iBlot Dry Transfer System or XCell II Blot Module with SureLock tank and achieve similar transfer efficiency.
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The most common cause of abnormally high current is the transfer buffer. If the transfer buffer is too concentrated, this leads to increased conductivity and current. High current may also occur if Tris-HCl is accidentally substituted for the Tris base required in the transfer buffer. This will again result in low buffer pH and lead to increased conductivity and current and subsequently, overheating. We recommend checking the transfer buffer and its reagent components and re-diluting or remaking the buffer.
Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.
- 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.
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For proteins larger than 100 kDa, 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.
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Here are possible causes and solutions:
- Presence of air bubbles between the gel and the membrane preventing the transfer of proteins. Be sure to remove all air bubbles between the gel and membrane by rolling a glass pipette over the membrane surface.
- Expired or creased membranes used. Use fresh, undamaged membranes.
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The swirling and diffuse banding patterns are typical of molecules moving laterally before binding to the membrane during transfer. Here are possible causes and solutions:
- Poor contact between the gel and the membrane: The gel should be attached to the membrane through capillary action. To ensure that this happens, make sure that you roll over the surface of each layer of the gel/membrane sandwich with a glass pipette to ensure good contact between the gel and the membrane. It is helpful to use a disposable pipette to place some extra transfer buffer on the surface of each layer as the sandwich is being made. Also, the pads need to be fully saturated (push down with gloved hand when they are placed in transfer buffer to make sure there are no air bubbles.)
- Under-compression of the gel: The gel/membrane assembly should be held securely between the two halves of the blot module. Try adding another pad or replace any pads that have lost their resiliency with fresh ones.
- Over-compression of the gel: A good indication of over-compression is if the gel has been excessively flattened. In the event that the sandwich is over-compressed, remove enough pads so that the blotter can be closed without exerting excess pressure on the gel and membrane.
Note: The height of the uncompressed pads should be 0.5-1.0 cm above the level of the sealing gasket.
Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.
Here are possible causes and solutions:
- High ionic strength of the transfer buffer. Prepare the buffer as described in the manual.
- Power supply is operating at a current close to the current limit of the power supply. Use a power supply with higher limits.
Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.
Here are possible causes and solutions:
- Too short a transfer time: Increase the blotting time by 15 minute increments.
- Inappropriate gel type: Check the percentage of the gel used and switch to a higher percentage gel.
- Inappropriate amount of SDS: Add 0.01-0.02% SDS to the transfer buffer to facilitate migration of the protein out of the gel.
- Inappropriate methanol content: Decrease the amount of methanol in the transfer buffer.
Note: Higher molecular weight proteins usually do not transfer completely as compared to mid to low molecular weight proteins.
Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.
Here are possible causes and solutions:
- Too long a transfer tim: Shorten the transfer time by 15 minute increments.
- Inappropriate amount of SDS: Do not include any SDS in the transfer buffer.
- Inappropriate methanol content: Add additional methanol to the transfer buffer to increase the binding capacity of the membrane.
- Inappropriate gel type: Check the percentage of the gel used and switch to a higher percentage gel.
- Sample overloaded: Decrease the sample load.
- Finally, if using nitrocellulose membrane, switch to PVDF which has a higher binding capacity.
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It is possible that the gel/membrane sandwich was assembled in the reverse direction such that the proteins have migrated out into the buffer. Assemble the blot sandwich in the correct order using instructions provided in the manual.
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The most common cause of abnormally high current is the buffer. If the buffer is too concentrated, this leads to increased conductivity and higher current. High current may also occur if Tris-HCl was accidentally substituted for the Tris base required in the transfer buffer. Tris-HCl results in a low buffer pH and leads to increased conductivity and current, and, subsequently, overheating. Check the transfer buffer and its reagent components, re-dilute, or remake the buffer.
Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.
Here are possible causes and solutions:
- The buffer was accidentally made too dilute, therefore increasing resistance and thus lowering conductivity and current: Check the transfer buffer and its reagent components and then re-dilute it or remake it.
- The circuit is broken or impeded, as in the case of a corroded or broken electrode or malfunctioning power supply: Check the equipment.
- There is a leak in the blot module (this is indicated by a drastic decrease in current and in buffer volume within the module): Ensure that the inner buffer chamber is filled sufficiently so that the wells are covered with buffer.
- Tape at the bottom of the gel cassette was not removed: Double check that the tape on the bottom of the gel has been removed.
Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.
We recommend transferring for 60 minutes at 10 volts with nitrocellulose and 20 volts with PVDF. An optional faster transfer for 30-45 minutes at 15 volts can be used, but a slight loss of sensitivity may be observed.
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We recommend using 1x Bolt Transfer buffer with 10% methanol.
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No. However, we recommend adding water to the Mini Gel Tank to help dissipate any heat that may be generated during transfer.
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Typically, 200-250 mL of 1X transfer buffer is needed per module which is 2-4 times less than with other transfer devices.
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The Mini Blot Module is designed exclusively for the Mini Gel Tank. It will also fit in the Bolt Mini Gel Tank (discontinued as of December 31, 2014) but will not fit in the XCell SureLock Mini Cell or other vendors' electrophoresis tanks.
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Due to the universal electrode design, the Mini Blot Module fits on either side of the Mini Gel Tank.
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We recommend transferring a single gel per blot module.
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We recommend using 2 sponge pads to start with. If the sponge pads become thinner with extended use, then an additional sponge pad can be used.
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Here are the replacement parts we offer for the Mini Blot Module:
- Mini Blot Module Gasket, Cat. No. B1001
- Gel Knife, Cat. No. EI9010
- Bolt Tank Base, Cat. No. B4478640
- Gel Runner Tank, Cat. No. B4478641
- Mini Gel Tank Lid, Cat. No. A25944
- Cassette Clamp, Right, Cat. No. B4478592
- Cassette Clamp, Left, Cat. No. B4478593
- Power Supply Adapters, Cat. No. ZA10001
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We recommend rinsing the blot module with deionized water after use. To clean any residual build-up in the blot module, apply 50% nitric acid in deionized water to areas inside the blot module until residual build-up is removed. Once the build-up is removed, rinse the module at least three times in deionized water. Do not submerge the blot module or soak overnight in nitric acid.
Note: Use gloves when preparing the nitric acid solution.
Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.
Yes, we offer the Mini Blot Module (Cat. No. B1000), designed to be used with the Mini Gel Tank. This blot module will also work with the Bolt Mini Gel Tank (discontinued as of December 31, 2014). Please note that the Bolt Mini Blot Module (discontinued as of December 31, 2014) is also compatible with both the Bolt Mini Gel Tank and the Mini Gel Tank.
Bolt Bis-Tris Plus gels can also be transferred using the XCell SureLock Mini Cell, iBlot Dry transfer system, or using the Invitrogen Semi-Dry Blotter.
Find additional tips, troubleshooting help, and resources within our Protein Electrophoresis and Western Blotting Support Center.