# Disaggregate mechanically ## Vortex with glass beads

Protocol: 5 mL sample + 1 mL [3 mm glass beads](https://help.sbtinstruments.com/encyclopedia/item-register/consumables/glass-beads-3-mm-50-g). 3 min at max speed. ## Bead beating

Protocol: 5 mL sample + 1 mL [3 mm glass beads](https://help.sbtinstruments.com/encyclopedia/item-register/consumables/glass-beads-3-mm-50-g). 1 min at max speed. ## Ultrasound bath

* Use 37-80 KHz cavitation frequency as lower frequency ultrasound (typically 20khz) results in aggressive cavitation that may induce lysis. * Use glass vessels for optimal propagation of ultrasound as the conventional plastic test tubes dampens ultrasound. * Make sure ultrasound bath is clean and that vessel is placing at the recommended water level and position in the bath * Typical protocol: 5 mL sample, 5 min @40 KHz in a glass vessel. If the ultrasound bath is equipped with a sweep functionality, use this. ## Ultrasound probe

* Typically, the rod-type sonicator probes are more aggressive than the above bath types, but amplitude, wattage and frequency parameters can be tailored. * Use [probes for small volumes](https://bandelin.com/en/shop/sonopulse-ultrasonic-homogenisers/sonopuls-hd-5020-homogeniser/) and higher frequencies than 20khz. * Cool sample in ice bath when performing homogenization * Typical protocol: 1mL sample on ice. Example: 10 cycles of 30 sec pulses at 20 watts, with 30 sec idle time. ## Rotor/stator blender

* Handheld rotor/stator equipment, e.g. [tissueruptor](https://www.qiagen.com/at/spotlight-pages/ias/automated-qpcr-workflow/detection/tissueruptor-ii/) or ultra-turrax types. Preferentially choose disposable probes to minimize contamination * Walk-away platforms like the [tube drive](https://www.ika.com/en/Products-Lab-Eq/Dispersers-Homogenizer-csp-177/ULTRA-TURRAX-Tube-Drive-cpdt-3646000/) and the DT-20M tubes are sealed to minimize aerosol formation. * Add antifoam agents, e.g. 1 µL Sigma antifoam 204 to 5 mL of culture * Typical protocol: Add 0.005-0.01% antifoam. 5-10mL sample. Homogenize for e.g. 2-5 min at max speed. ## Syringe homogenization

Protocol: Passage 200 µL sample through 27G cannula 20 times.