Particles approximately 100–200 nm in size can be recovered.

Reference:
Particle size distribution of particles recovered from HEK293S cell culture supernatant

For cell culture supernatant samples, the recommended processing volume is 1–10 mL. Please note that the required centrifugation time may vary depending on the sample.

The amount of exosomes secreted varies depending on the cell type, so the amount of particles recovered may also vary depending on the cell type and culture conditions. As an example, we have recovered particles from 10 mL of HEK293S cell culture supernatant cultured under shaking conditions, with the following results:

Recovered protein amount: 131 μg
Recovered particle count: 2.5 × 10¹⁰ particles

Note: The protein amount was measured using the BCA assay, and the particle count was measured by nanoparticle tracking analysis.

We do not recommend using serum-containing medium. Serum used for cell culture, such as FBS, also contains exosomes. Therefore, exosomes recovered from serum-containing cell culture supernatant may include serum-derived exosomes, which could affect experimental results.

The recovered exosomes can be stored at 4°C for 1 to 6 months. For long-term storage, store them at −80°C. To minimize repeated freeze–thaw cycles, aliquot the exosomes before storage.

Reference:
Yoshioka, Y. and Ochiya, T. (2020). Exosome Experimental Guide. ISBN: 978-4-7581-2246-7.

Yes, a swing-bucket rotor can also be used. However, our internal validation has shown that the particle recovery efficiency when using a swing-bucket rotor is approximately half that achieved with a fixed-angle rotor.

When using a swing-bucket rotor, set the centrifugation conditions to 2,000 × g.

Yes, a sterile filtration filter with a pore size larger than 0.22 μm can also be used. However, pretreatment using a 0.22 μm sterile filtration filter is recommended, as centrifugation may take longer when using filters with larger pore sizes.

The recovery rate may vary depending on the sample.

As an example, the recovery rate was evaluated by re-purifying commercially available bovine milk-derived exosomes using ExoIsolatorⅡ DirectSpin. The result is shown below:

Particle recovery efficiency: 26.7%

Note: Please refer to the experimental examples in the instruction manual for details.