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Viral isolation kit QIA buffer substitute


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#1 Trof

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Posted 07 April 2020 - 06:17 AM

Hi all, I think this may interest people around the glove as many labs steuggle with SARS-CoV-2 diagnostics..

Here we use QIAGEN viral RNA kit, but we seemed to be limited (among others) by lysis buffer AVL used in a robot.
It should be guanidium salt something, but I'm trying to find exact composition or lysis buffer (virus inactivating) that would be compatible with Q columns.
It's not about price anymore.. it is about availability.
Anyone having the same problem?
Thanks..

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#2 bob1

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Posted 07 April 2020 - 12:43 PM

Yep, same problem world-wide. Have a look at the MSDS; it tells you that this is indeed a guanidinium salt solution. I think from memory there is a buffer (tris probably), some EDTA and a couple of other things, but I don't know the exact composition.

 

As far as I can tell, the columns are the same for pretty much every kit and the buffers are just based off the old manual RNA extractions that we performed 20 years ago. If you have access to a recent copy of Sambrook's Molecular Cloning, it'll probably tell you explicitly how to make these. Unfortunately I only have a 1989 version and only Vol. 1 and 2, so I can't even look it up for you.



#3 hobglobin

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Posted 08 April 2020 - 09:18 AM

Unfortunately openwetware also does not have this one, not sure if there is an equivalent one listed (never used RNA kits). 

Sambrook's Molecular Cloning you can get more or easily as pdf for free (but with the usual legal issues ph34r.png)



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#4 Trof

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Posted 08 April 2020 - 10:32 AM

Yeah, checked Openwetware :/
I actualy do have Molecular Cloning 3rd edition (with usual legal issues), but it only has solutions for phenol-chlorophorm and ultra-centrifugation mRNA isolation (Chapter 7.10 or something), but I would need something more compatible with silica-based isolation.

Found this:
https://jcm.asm.org/.../3/495.full.pdf

(GuSCN, Tris pH6.4, EDTA and TritonX-100)

But main problem is, we do not have much time nor spare columns to optimize various GuSCN versions, when it is not even sure, how long we can even use Q columns and other consumables (and since we use a robot, changing just column in not option).
It may be, we run out of other stuff just a bit after we run out of AVL. There is a state-wide effort to optimize magnetic-bead isolation instead, because those can be manufactured in university labs in our country.


Our country has a serious deficiency in lighthouses. I assume the main reason is that we have no sea.

I never trust anything that can't be doubted.

'Normal' is a dryer setting. - Elizabeth Moon





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