4°C Cooling | Air Cooling | Use and Operation
Want to know more about homogenization methods in general? Click here for an overview.
What is 4°C Cooling™?
4°C cooling uses a compartment filled with dry ice to keep samples near 4°C. When the instrument is in operation, cold air from the compartment is drawn into the instrument, cooling the samples. Some users prefer to pre-cool their Bullet Blenders before use. To pre-cool, fill the compartment with dry ice and run the instrument with no tubes inside for 5 minutes or until the “4°C” light comes on.
Which models have 4°C Cooling?
The 4ºC cooling feature comes on Gold models of the Bullet Blender— BB24-AU which homogenizes samples in 1.5 mL microcentrifuge tubes and BB50-AU which homogenizes samples in 50 mL tubes.
How much dry ice do I need to use 4°C Cooling?
It takes at least 2 lbs of dry ice for the instrument to achieve maximum cooling. We recommend filling the chamber with 3 lbs of dry ice- depending on how heavily the instrument is used, this should keep samples at 4°C for an entire day of homogenization.
Why is the 4°C light not lighting up on my Bullet Blender Gold unit?
Frost from condensation may be blocking the air passage. Remove the inner dry ice compartment and gently agitate it or replace the ice.
What will happen if I operate my Bullet Blender Gold unit with no dry ice?
When there is no dry ice in the compartment, you should operate the instrument with the lid of the dry ice compartment open so that the instrument can operate using Air Cooling™.
What is Air Cooling™?
Most models have Air Cooling™. These models exchange air with their surroundings; they do not have refrigeration units. Air Cooling™ effectively draws cold air into the instrument if you are using the Bullet Blender® in a cold room or a refrigerated cabinet. Air Cooling™ is also useful if you're running several sample cycles in a row or if your samples are extra sensitive to heating up. Normally, the samples in the Bullet Blender® only heat up a few degrees and stay quite cool. However, if you're running a second batch of samples soon afterward, they will heat up a little more because the Bullet Blender® is starting out slightly warmer. The Air Cooling™ keeps the Bullet Blender® cooler so that the first batch of samples heat up a little less and subsequent batches are kept just about as cool as the first batch.
Which models have Air Cooling™?
The Air Cooling™ feature comes on all models of the Bullet Blender® except the standard Bullet Blender for microcentrifuge tubes (BBX24). Gold models not running with dry ice can be run in Air Cooling™ mode if the dry ice compartment lid is left open to enable air exchange.
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Use and Operation:
How is the Bullet Blender different from other high-throughput homogenizers on the market?
The Bullet Blender is similar to other Bead Mill Style homogenizers in that it processes each sample in its own tube using homogenization beads. Therefore, there is no chance of cross-contamination and there are no parts that need cleaning after each sample.
A unique feature of the Bullet Blender is the way it agitates the sample tubes. Other bead-mill homogenizers vigorously shake the sample tubes, which requires a great deal of energy. The large motors required tend to make these homogenizers large and expensive: many are 2-3 times the cost of the Bullet Blender.
The Bullet Blender, on the other hand, uses a patented striking technology. Strikers impact the sample tubes. These bursts of energy force the sample tubes to oscillate vigorously, at a range of frequencies. We find that a range of frequencies is more effective than a single frequency. This energy-efficient approach requires a much smaller motor and avoids the need to clamp tubes into a holder, so the Bullet Blender is smaller, lighter, and less expensive than competitors' equipment while working just as well.
Also unique to the Bullet Blender, the sample tubes rapidly oscillate in a stream of rapidly flowing air. They are not clamped into a holder, preventing air exchange. Therefore, convective cooling is very effective and sample heating is minimal, especially in models with Air Cooling or 4ºC Cooling. The Bullet Blender can also be used in a cold room, unlike many of the other instruments on the market.
What types of tubes can be used?
Microcentrifuge models: The Bullet Blender® Storm and Gold models work with our RINO tubes or 1.5ml Eppendorf® Safe-Lock tubes. The Bullet Blender® and Bullet Blender® Blue work with standard polypropylene snap-cap microcentrifuge tubes though we strongly recommend 1.5 mL or 2.0 mL Eppendorf® Safe-Lock tubes. When using 2 mL tubes in the the Bullet Blender® or the Bullet Blender® Blue we recommend using a minimum of 2 tubes per homogenization cycle. The obsolete screw cap models (BBX24W and BBX24BW) use Axygen® and Corning® 1.5 ml screw cap tubes.
5mL tube models: The Bullet Blender® Storm 5 units work with 5ml Axygen® brand screw top transport tubes. We also offer the Bullet Blender 5 E that works with Eppendorf® 5 mL tubes. As these are different in size than the Axygen tubes, these 5mL models can handle one type of tube or the other, but not both in the same instrument.
50mL tube models: The Bullet Blender 50-DX and 50-AU (DX and Gold models) work with 50 mL skirted (self-standing) polypropylene tubes, such as TPP® brand or Corning® brand. The slightly hazy polypropylene tubes are tougher and resist the striking better than the clearer tubes, which might crack at higher speeds. Use high quality tubes, rather than bargain brand tubes which are not as strong.
How do you set the Bullet Blender® to mix, disrupt, or homogenize different types of samples?
You can adjust the speed and the duration. Adjusting the speed affects how firmly the strikers impact the sample tubes, and thus how vigorously the sample tubes are forced to oscillate. The "speed" setting is a measure of homogenization power. A direct measure of speed would be misleading because the sample tubes are forced to oscillate at a range of frequencies. Adjusting the duration affects the total number of impacts. Homogenizing tough tissue requires longer durations at full "speed", while mixing or cell dissociation requires lower "speeds".
How easy is it to operate the Bullet Blender®?
Simply place up to 24 sample tubes in the holders, set the desired duration and speed, and push start. All the samples will be processed simultaneously, without any chance of cross contamination.
How much sample can be homogenized in each microcentrifuge tube?
General tips: Tubes should be filled no more than 75% full (50% for 50 mL tubes). The air volume inside the tube is critical to allow the sample to be ground up inside the tube, so if the tube is overfilled, the efficiency of the homogenization is reduced. A very high ratio of buffer to sample can also impair homogenization. A good general rule of thumb is to use one volume of beads and two volumes of buffer per volume of sample.
For microcentrifuge model Bullet Blenders, we recommend using 10-300 µL of tissue. Less than 10 µL will work, but it requires careful attention to the amount of beads and volume of homogenization buffer. Samples larger than 300 µL are not recommended. The 5 mL tubes can hold up to about 1 mL of sample tissue or pelleted cultured cells, and a total of 3.5 mL of sample, buffer, and beads combined. The 50 mL tubes can hold up to about 3.5 mL of sample tissue or pelleted cultured cells, and a total of 20 mL of sample, buffer, and beads combined. Note that for most wet organ tissue 1 mL≈1 g.
Do sample tubes need to be placed symmetrically?
No. There are no restrictions as to which holes to place your sample tubes in, however you may get better results if you space them apart as evenly as possible.
How do you retrieve the sample from the tubes?
After the Bullet Blender® stops, simply remove the tubes and draw off the liquid with a pipet. If a clear lysate is desired, the sample may be centrifuged to clarify the lysate prior to use in your applications. Tubes may be centrifuged with the beads in them for ease of handling.
Which beads should I use?
The protocols page contains suggested beads, speeds, and durations for processing many types of tissues and cells. The two basic rules for bead selection: (1) use denser beads for tougher samples, and (2) use beads of a comparable size to the size of your samples to maximize the effectiveness of collisions between beads and samples. For example, if you wanted to lyse mammalian cells with a soft membrane, use small glass beads. If you want to homogenize larger pieces of tough tissue, use large steel beads. "UFO" beads are excellent for homogenizing fibrous or resilient samples. You can also contact our technical support staff at firstname.lastname@example.org if you have any questions.
I am trying to process tough samples in the Bullet Blender® and the homogenization is incomplete. How can I improve my results?
There are several ways that you can increase the homogenization of your samples.
- Examine the shape of your samples- long thin samples will homogenize faster than cube-shaped or round samples.
- Try cutting your samples into smaller pieces.
- Ensure your tubes are not overloaded.
- If you are running many samples at once in the standard or Blue models of the Bullet Blender (BBX24 or BBX24B), try runing fewer tubes at a time. If a sample is extremely tough (e.g. mouse femur) you may only be able to achieve complete homogenization with 2-4 tubes at a time.
- If you are using the BB50-DX or BB50-AU, make sure that all of the sleeves are removed except for the sleeves containing tubes.
I previously owned another bead mill homogenizer. Can I use the same protocols that we developed for the other homogenizer with the Bullet Blender®?
No, the mechanism of action is different from all other homogenizers on the market. Our patented striking technology makes the Bullet Blender both highly efficient and effective. If you use another product's protocol with the Bullet Blender, you are very likely to experience different results. Please use one of our established protocols, or empirically determine a protocol that works best for your samples. We also offer free protocol development services.
Is it possible to homogenize tissue that has been frozen in liquid nitrogen?
It is possible to homogenize tissue that was frozen in liquid nitrogen. Immerse the frozen tissue in cold buffer and allow it to thaw, then treat it as you would any other sample. If the tissue was dried before it was frozen, you can pulverize the tissue by homogenizing with beads only (no buffer), then adding the cold buffer and running again to complete the homogenization. You cannot homogenize "wet" tissue while it is still frozen, as it will effectively be a block of ice and homogenization efficiency will be very poor.
What is the "Key-Switch Lock Out" option for Bullet Blenders?
This key switch option incapacitates the electronics, preventing others from operating your Bullet Blender. Thereby, others are discouraged from working in your area, using your supplies, and upsetting your experiments.
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Contact us if you have any more questions.