Antibody stability and additives - (May/21/2007 )
Hi Gang.
All of my reading has said that antibodies are very stable over long periods of time. I affinity purifiy my antibodies and then dialyze against 20mM NaPO4/28mM NaCl pH 7.4 and store in 0.2ml aliquots at -80.
When I want to use an aliquot I take one out of the -80 and store it at 4C until it is all used up. I typically use my antibodies for Westerns (work great) and Immunoppt (work sporadically).
Will I do any damage to my work if I add Na Azide (to 0.2%) and EDTA to 1mM to my antibody prep once I take it out of the -80?
I am thinking perhaps the erratic IP results might be a result of degradation of antibody over time.
Thanks
It would only affect your assay if you're using cells, since the azide is toxic to them. How long does it take you to go through one 200ul aliquot of antibody?
Could you add azide such that the final conc. is 0.02%. This is conc. we use for some of them.
Now we are actually storing the daily aliquot in -20C ( PI Insists). Seems to be working fine.
All of my reading has said that antibodies are very stable over long periods of time. I affinity purifiy my antibodies and then dialyze against 20mM NaPO4/28mM NaCl pH 7.4 and store in 0.2ml aliquots at -80.
When I want to use an aliquot I take one out of the -80 and store it at 4C until it is all used up. I typically use my antibodies for Westerns (work great) and Immunoppt (work sporadically).
Will I do any damage to my work if I add Na Azide (to 0.2%) and EDTA to 1mM to my antibody prep once I take it out of the -80?
I am thinking perhaps the erratic IP results might be a result of degradation of antibody over time.
Thanks
some Ab´s are not recommended to freeze, but adding glycerol make most freezable; use shock freezing;
we use lower NaN3 (0.001 %) but not for 2nd Ab if peroxidase labeled as NaN3 blocks peroxidase
Thanks for the replies.
I have tried storing at -20, but I thought that the regular freeze-that was adverly effecting the antibodies.
Generally I go thru 200ul in about 3 months time.
The antibodies have not been used such that they are placed on live cells.
I have thought about glycerol, but have rejected it because I don't know what effect it will have on western blots and IMP's.