antibody purification

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Big E
Big E's picture
antibody purification

Can I get protocols on the easiest, least expensive, and best methods for purifying monoclonal antibodies from hybridoma cell lines?

Germey
Germey's picture
That's and interesting way to

That's and interesting way to pose the question. Are you hoping for one protocol that meets all 3 requirments, or 3 different protocols? The fact of the matter is that there are several different routes to get to a purified antibody. The one that is best will depend on your specific needs as far as purity, activity, and time. Also a factor is batch size and yield requirements.
Probably the easiest is to use Protien A or Protein G, depending on the antibody's species of origin. The drawbacks are the cost of the media, and a fairly harsh elution that can affect the anitbody's activity.
You can use Ion exchange to get a fairly to highly pure antibody and use a less harsh elution.
If you have very large batches and do not need a high level of purity, you can do a salt based precipitation followed by a desalting step, either by size exclusion chromatography or an ultrafilter-based buffer exchange.
To try any of these methods in very small scale, as well as for some specific protocols, I recommend you visit www.vivascience.com (I am not affiliated with the company... but used to be).
Let me know if you want any more specific advice.
Cheers,
Jeremy

bwbrian
bwbrian's picture
If your cultures are grown

If your cultures are grown with bovine serum as a supplement, then you may want to consider Protein L instead of Protein A or G. Protein L doesn't bind to bovine antibodies, so you can purify your monoclonals quickly and get a nice pure prep. Protein L is more expensive than Protein A, but it's worth the extra cost to get a pure antibody in a single step.

Limin
Limin's picture
DaJo wrote:Can I get

DaJo wrote:

Can I get protocols on the easiest, least expensive, and best methods for purifying monoclonal antibodies from hybridoma cell lines?

Here is a cool web site for Ab purification. It has plenty of protocols.
http://www.protocol-online.org/prot/Molecular_Biology/Protein/Extraction___Purification/Antibody_Purification/index.html

manleyenoch
manleyenoch's picture
I will purify your antibodies

I will purify your antibodies in Ascites and antisera.
I will show you how to purify your antibodies.

<<<Moderator note: This is not very helpful. If you have an appropriate protocol, please post it here. If you're soliciting business, you're in the wrong forum! Cheers. -sam>>>