Good concept, but to be widely worked with variety mo ab s for clear separation and proved .

I do not have experience using CIM Column for the separation of Ig. from cell culture supernatant. But the quality and quantity of IgG separated by protein A is still not beatable. If the manufacturer of CIM column claims higher quality and quantity of IgG,then give it a try on a small scale and use Protein A column as control to compare the results.

Good Luck,

Dr. Masoom Shah

Ph.D Microbiology

Auburn University Alabama, USA

The metal-chelate columns are useful only if the mAb is recombinant and expressed with the necessary tag; for naturally occuring IgG it is not useful. The benefits(claimed) of the metal–chelate CIM monolithic column are (1) less harsh elution conditions (reduced denaturation of IgG) and (2) higher flow rates with no chance of eluted Protein A in the mix.

Thank you Christopher,

So it is only suitable for mAb that expresses required tag. Good to know that.

Thanks again.

Dr. Masoom Shah

I agree with the opinion of Dr. Christopher Lloyd, metal-chelate CIM chromatography is a fundamentally different type of purification. And this way of purification of antibodies from supernatant, apparently, is not acceptable.

I agree with you, Mr.andreev

Thank you

I am so sorry I have no idea about your reaserch subject

@ Sergey Andreev ,

Why it is not acceptable ?

Do you know what are the bottleneck of mAb purification process where we are using Protein A chromatography wrt cost & quality ?

1. Protein A cost is very high which is always be a bottleneck small & mid sized bio pharmaceuticals for commercial production & doing it scale up.

2.Protein A require very harsh acidic condition ( less than pH 4) for elution which leads to aggregation.

Can anyone help me to understand , why we are finding alternative which is working good & why we are always want to improve our process wrt cost & quality ?

Why one has to find out alternative of Protein A chromatography although its works good ?

1. Protein A media cost is very high which is always be a bottleneck for small & mid sized bio pharmaceuticals for commercial production & its scale up.

2.Protein A require very harsh acidic condition ( less than pH 4) for elution which leads to aggregation & other structural conformational change ( Process related impurities) for removal of that you have to add one or two additional steps.

3. Protein A ligand is leaching in output which is one of regulatory constraints & you have to show your ligand leaching data , so additional cost adding in analysis.

4.Resin re-usability is also limiting.

I know many who are using Protein A chromatography , are disagree with my comments. But its fact. Many researcher are working , finding alternative for this & come up with good innovative ideas & research which might be next generation of cost effective MAb purification.I have showed here one of the alternative metal–chelate CIM monolithic columns.

Best Luck.

I prefere to use ProteinA, using the apropriete pH for Mab elution and them Ionic chromatography to polish!

the metal–chelate CIM you still see a lot of extra bands on SDS-PAGE after purification....

Very interesting topic you have started, however you have answered many of your questions.

Yes it is right for purification of IgG protein A is good choice but that doest mean we should never try for alternatives. Scientist all over the world are working on this aspect, however the problem is protein A has reached to such a position and has gained such as confidence that many people are not ready to change it, they are ready to work with its drawbacks. It is going to take longer time for alternative of protein A to replace it completely.

Regarding CIM monolithic columns i dont think they can replace protein A, because their are certain issues with CIM monolithic also. Based on my limited experience with such columns i can say they develop huge back pressure and many a times these columns gets chocked very fast.

We have developed such as alternative to protein A, that is called AbSep. It overcomes many of the protein A drawbacks such as use of harsh conditions for elution, ligand leaching, cost etc.

What is the essence of the AbSep?

Please go through the below given article to know about AbSep:

http://www.sciencedirect.com/science/article/pii/S0021967311001646

Thank you

Hi! Monika, nice paper, How do you get the PEG out of you sample? using the AbSep method?

Thanks Fernando, PEG can be removed using 5kDa or 10kDa membrane.