Preparation of Sample
Use 0.1 mm beads for bacteria, 0.5 mm beads for yeast, fungi, and tissue culture cells and 1.0 mm beads for plant and animal tissue. Up to 400 mg wet wt of biomass can be disrupted per ml of extraction media. If you have solid tissue, pre-chop it with a single-edge razor blade or scalpel.**
** Tissue sample sizes over a few tens of milligrams should be prechopped into pieces less than 1 mm in cross-section. If your sample is already harvested and frozen, do not thaw. This is especially important if you are isolating nucleic acids. Rather, Cryopulverize the sample. For additional information on this latter method, see https://www.biospec.com/products/biopulverizer?taxon_id=28.
Fill screw-cap vials at least 1/2 full (1/2 - 3/4 is okay) with beads. BioSpec has a Flip and Fill (https://biospec.com/products?keywords=702L) to make loading a few 96-well microplates easy and accurate. For loading numerous deep-well microplates, see Cat. No. 617L and Cat. No 1117. Bead size and type should be matched to the biomaterial you are processing (see https://www.biospec.com/beads-guide-lines/loading-beads-in-vials-or-microplates for details). Then add your extraction media and cells, being sure to fill the microtubes or wells almost to the top. This will exclude as much air as is practical from the microtubes or wells and minimize foaming during beadbeating.
We highly recommend using screw-cap microtubes with o-ring seals. Snap-top microtubes can leak and release sample aerosol during shaking. Be sure there are no beads on the threads of the microtubes or on the lip area of the microplate(s) when sealing the microtube or microplate(s). If sealing microplates with a flexible mat, firmly press the mat or film over the entire top surface of the microplate. BioSpec offers a manual mat roller to make sealing easy and secure. Adhesive film does not maintain a good seal on microplates during high energy shaking in a MiniBeasdbeater-96.
Even some microplates and their matching mats or sealing films perform better than others when using our high-energy MBB-96. BioSpec has tried microplate sealing techniques using mats made by several different manufacturers in an effort to find those that provide the integrity needed to prevent sample leakage from well to well or, in the case of using steel bead media, rupture of the bottoms of the plate wells during the beadbeating process. Only Porvair Sciences (Wales, UK) deep-well microplate products 219030 (2 ml) sealed with EVA polymer sealing mats 219004, both distributed by BioSpec Products, have passed our rigorous leakage and breakage tests.
Operating the Mini-Beadbeater-96
1) Insert microtubes into the tube holder and distribute the tubes symmetrically with at least, four tubes on each corner of the tube holder. Place the loaded rack of microtubes or sealed microplate(s) into the chamber holder, top side toward you. If you are using 1 ml microplates you can stack two at a time. Screw down the two black plastic knobs until the top stainless steel retainer plate is in contact with the microtube caps or top of the microplate(s). Tighten the knobs snugly but not overly tight. Finally, firmly screw down the white nylon wing nuts and close the black safety cover.
2) Turn on the 'on-off' switch located on the front of the MBB-96 cabinet and select the cell disruption time (0-5 min). Typical cell disruption times are 3 minutes.
Notes: If you are working with heat-sensitive material such as native proteins, consider pre-cooling your beads and sample, homogenizing for 1 minute, then removing the vials or plate and cooling in an ice-water mixture for 1 minute. Cycle for a total 'on' time of three minutes. Cooling is not needed when extracting nucleic acids in nucleic acid isolation media. Operating an MBB-96 in a cold room is not recommended.
Cold air does not prevent sample warming during beadbeating. Do not operate the MBB-96 in a cold room. Special electronics in the unit will prevent the machine from starting.
Alternatively, a pre-chilled aluminum vial rack (https://biospec.com/products/aluminum-vial-rack-2ml?taxon_id=4) can help control sample warming.
Operate the MBB-96 with the black plastic hood lowered over the chamber. This prevents the user accidentally coming in contact with the shaker during operation and will trap anything should it break free. Do not bypass the safety features of the lid.
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