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Published TCIMAIL newest issue No.197
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We know that when researching exosomes, the extraction of high-purity, intact exosomes from your samples is of utmost importance. That's why TCI offers ready-to-use solutions optimized for exosome isolation from serum and cell culture supernatant.
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Example of Exosome Isolation from Human Serum Using E1553
- Centrifuge human serum sample at 3,000 x g for 15 minutes at 4 °C.
- Transfer the supernatant containing clarified serum to a new tube.
- Add Exosome Isolation Reagent [for Serum] [E1553] to the serum and mix via pipetting until homogeneous. Use 20 μL of E1553 per 100 μL serum.
- Incubate for 30 minutes at 4 °C.
- Centrifuge at 10,000 x g for 10 minutes at 4 °C.
- Remove the supernatant carefully without disturbing the pellet (containing exosomes).
- Resuspend pellet in 1x PBS. Use 25-50 μL of PBS per 100 μL of starting serum.
Figure 1. Size distribution of isolated exosomes using E1553
The particle size of exosomes was measured by dynamic light scattering. The pellet was found to be highly enriched in human serum-derived exosomes.
Figure 2. Quantification of isolated exosomes using E1553 by ELISA
An ELISA plate was coated with anti-CD63 antibody. Plates were then incubated with resuspended pellet and supernatant adjusted to the same protein concentration.
After washing, detect was carried out using anti-CD9 antibody. As can be shown, exosomes expressing CD9 and CD63 on the surface can be collected at high yields.
Example of Exosome Isolation from Cell Culture Supernatant Using E1601
- Collect culture supernatant of MCF7 cells and centrifuge at 3,000 x g for 15 minutes at 4 °C.
- Transfer the supernatant to a new tube.
- Add Exosome Isolation Reagent [for Cell Culture Media] [E1601] to the supernatant and mix via pipetting until homogeneous. Use 500 μL of E1601 per 1 mL supernatant.
- Incubate at 4 °C overnight.
- Centrifuge at 10,000 x g for 1 hour at 4 °C.
- Remove the supernatant carefully (pellet is often invisible).
- Resuspend pellet in 1x PBS. Use 25-50 μL of PBS per 1 mL of starting media.
Figure 3. Size distribution of isolated exosomes using E1601
The particle size of exosomes was measured by dynamic light scattering. The pellet was found to be highly enriched in MCF7 cell-derived exosomes.
Figure 4. Quantification of isolated exosomes using E1601 by ELISA
An ELISA plate was coated with anti-CD63 antibody. Plates were then incubated with diluted pellet and undiluted supernatant.
After washing, detect was carried out using anti-CD9 antibody. As can be shown, exosomes expressing CD9 and CD63 on the surface can be collected at high yields.
Figure 5. Western blotting of isolated exosomes using E1601
Proteins contained in the pellet were transferred to a PVDF membrane after non-reducing electrophoresis.
CD9, a surface marker for exosomes, was detected using mouse anti-CD9 antibody.