Monitoring cell lysis.  The MiniBeadbeater will disrupt over 90% of the cells in about 2-5 minutes of operation.  The homogenization procedure involves cell "cracking" action rather than high shear.  After homogenization, cell membranes may still appear to be intact when viewed under a microscope.  Therefore, to monitor the time course of cell breakage, rely on assay methods that measure intracellular constituents (e.g., OD260, enzyme activity, protein staining, PAGE).

Temperature control.  The homogenate will be warm after three minutes of "BeadBeating".  When isolating proteins, membranes or organelles, cooling may be necessary.  When isolating nucleic acids in aggressive extraction media containing phenol/chloroform, guanidinium salts, and/or detergents, temperature control is usually not necessary.  The easiest way to minimize heating is to start off with the vial, beads and buffer pre-chilled and by operating the MBB for one minute "ON" and then let the homogenate rest for one minute, cycling thus until homogenization is complete.  Consider placing the vials in ice water during the resting period.  Operating the MBB in the cold room does little to keep product cool.

Bead size selection.   The correct size beads are 0.1 mm dia. for bacteria, 0.5 mm dia. for yeast, and 1.0 mm dia. or 2.5 mm dia. for chopped-up plant or animal tissue.  While glass bead media is most commonly used, denser bead media is available for tough materials.   A list of bead media is located at our web site: http://www.biospec.com.  Click on Products, then on Beads.

Fill the two ml screw-cap vial at least 1/2 full (1/2-3/4 is okay) with beads and the rest of the volume with buffer and biomaterial.  Maximum biomass concentration during homogenization is 400 mg wet weight per ml of buffer.  The vial should be filled to the top to exclude as much air as possible when the vial cap is screwed on.  Note that dry beads can entrap air, especially when using 0.1 mm dia. beads.  You may have to invert the vial several times to wet the beads and then top-up the vial with more buffer.  Smaller vials can be used in the MBB using a special adapter available on request.

Insert the vial securely in the arm assembly.  Close the black safety cover.

Operate the BeadBeater for a total of three minutes by selecting a speed (the rabbit!) of 4800 rpm (48 on display) and a time (the clock!) of 180 seconds (18 on display).  At the end of the 3-minute run, the temperature of the homogenate will be about 25 deg C.  For cooling considerations, see comments above.

Most of the homogenate can be recovered by simply decanting after the beads settle to the bottom of the vial.  To recover the entire product, one can either wash the beads with buffer or, with beads 0.5 mm dia. or larger, simply plunge a micropipette to the bottom of the vial and sucking out all the liquid
 

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Last modified: 09/21/07.