Virus purification using 20% sucrose cushion centrifugation

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Virus purification using 20% sucrose cushion centrifugation

Postby thomson90 » Mar 22 2018 3:22 am

Hi, for those with experience, can you please help me check if anything wrong with my protocol?

- 900ul virus supernatant (clarified from cell debris by low speed centrifugation and 0.45um filter)
- layer on top of 100ul 20% sucrose in a 1.5ml tube
- spin max speed (13200rpm) with normal tabletop centrifuge machine for 4h (we do not have ultracentrifuge machine in our lab)
- pipet out supernatant, remove sucrose
- air dry the tube upside down in cabinet for 20 min
- resuspend in 100ul PBS
- recover in RT for 10 min

Take 10ul pelleted virion + 2.5 ul 4X Laemli buffer and run SDS-PAGE for western blot detection

thanks
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