what is the role of glycine in antibody purificiation??? - (Jul/25/2005 )
In many antibody purification protocols, glycine is broadly used. Would someone kindly tell me what's the role of glycine and whether i can use some other substitutes? Thanks.
is the glycine used as a amino-acid for decreasing non-specific adsorption of Abs or proteins to the matrix...
I believe glycine is used to elute bound antibodies in an acidic condition. I forget what concentration but the glycine I use is around pH 3.5.
Typically people catch the eluted fractions in some Tris (pH very high) to immediately neutralize it.
There are other ways to elute antibody of course.
As with any column, before using it, typically I think people run all their different buffers through it first to get rid of anything non-specific that make come off with the buffers alone.
Harlowe and Lane published a great lab manual "Antibodies: A laboratory manual" that will give you both detailed protocols and explanations for what the different buffers etc do, especially in regards to antibodies.
Another great source is the Current Protocols in Immunology (or other fields). Most of these are available on line now. You may need to click througha f ew links to find the full-text. The best part about these protocols is that they feature a commentary at the end of each, explaining the critical parameters and *why* things might or might not work.
Yes I agree with Flickers
I think Glycine is used simply because it buffers well in the acidic (pH 2-3) range, which is necessary for antibody elution.