For some biofluids, the sample can be concentrated before application to qEV columns to enable larger volumes to be processed. The exact protocol may require modification, depending on the sample used and other preparation steps.
These are the types of filters that can be used:
- Merck's Microcon-30kDa Centrifugal Filter Unit to concentrate down to 0.1 mL, these require a standard lab microfuge up to 14,000 g
- 100 kDa cut-off Amicon Ultra 15 which is a 50 mL tube unit and requires a low speed bench centrifuge capable of up to 4000 g. This takes approximately 15 min/spin. This unit concentrates 15 mL down to approximately 0.5 mL
- 100kDa cut-off Centricon Plus-70 to concentrate up to 70 mL
- Amicon Stirred cells and Pellicon Tangential Flow devices, for larger volumes.
For Amicon filters, particles or proteins that are <100 kDa that can pass through the filter, as these are much smaller than EVs (for instance BSA which is a protien of 66 kDa and is 2-5nm in diameter). When concentrating a sample, the EVs are much larger than the filter that they are retained above the membrane and hence are concentrated; whereas the bulk fluid and proteins pass through and are removed.