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PEG precipitation


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#1 ixsix

ixsix

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Posted 05 August 2004 - 07:29 PM

I combined cells and medium to purify aav with PEG 8000. In the final virus stork, lots of FCS was still detected by SDS. While the paper I looked for got quite pure virus(96%). AAV is resistant to chloroform.

Here is the flow chart. Does anyone have any opinion?

1 Homogenize cells by ultrasonic in start material(cells and medium combined)
2 10% chloroform and 1M nacl added
3 shaking 30 min
4 centrifuge and take the supernatant
5 add PEG 8000(10% w/v)
6 stand on the ice for an hour
7 centrifuge
8 collect the pellet and dissolve in pbs
9 add DNase I and RNase A for 30 min
10 add same volume of chloroform
11 centrifuge and take the supernatant




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