Use the filter-centrifugation method to consistently increase the concentration of your sample. Unlike simple centrifugation, filter-centrifugation gives consistent results.
In addition, filter-centrifugation is more gentle for the bacteria. The force required to drive liquid through the membrane filter is relatively subtle.
There are too many pitfalls that each lead to inconsistent results.
For dilute bacterial samples, the pellet will be brittle and very difficult to see with the naked eye. When you remove the supernatant, you may remove (parts of) the pellet as well.
Repeat as necessary to increase the concentration further.
All our vials are 15 mL conical centrifuge tubes. They fit inside standard centrifuges.
The pellet may be difficult to see
The pellet may be transparent and difficult to see with the naked eye. In turn, there is a high risk that you unintentionally remove bacteria from your sample when you try to remove the supernatant.
How to resuspend bacteria
The centrifugation process puts the bacteria close together. In turn, the bacteria may form clumps (aggregates). This is something to avoid. See Break up clumps and chains for more information.
Here is a short how-to guide for proper resuspension:
Resuspend the pellet in diluent. Use an empty vial for this purpose. Target the 10 mL mark.
Coarse disaggregation. Use a pipette to break apart the cells. Repeatedly aspirate and dispense the liquid.
Fine disaggregation. Use one of the mechanical disaggregation methods (See Disaggregate mechanically). For example:
Ultrasound bath (20 khz, 5 min)
Vortex with 3 mm glass beads. Add beads to increase the liquid level by 1 mL and vortex (1 min, max RPM. Decant liquid to empty vial before you measure on BactoBox®.
