Hello, I would like to reduce some antibody (anti-mouse CD11c) using DTT, desalting it and then aliquot it and freeze it at -40C? Do you think the Ab will stay in a reduced form over time?
Thank you for your help, Fanny
Dear Fanny,
thats a really good question! If you completely remove the DDT and there is no redcing agent present, a fraction of the antibody will get oxidized again. And usually that is a larges fraction, if not almost all molecules depending on the mab.
You can prevent this by using iodoacetamid to block the reduced cysteins. Or you include a redox buffer in your aliquots, such as glutathion.
Be careful how u design the use of it. Medium is often quite oxidized and the mab will rapidly get oxidized in it.
I would recommend to store it at lower temperature.
As a tiny rule which I use:
the speed of chemical reactions double with each increase in 10 degrees.
So as example:
if you would thing that the reduced antibody would have maybe a half-life of one day at zero degress, each reduction of the temperature by 10 degrees this would double the half-life.
so at -10 the half-life would be 2 days,
at -20 it woud be 4 days,
at -30 then 8 days
and at -40 finally 16 days.
This half-life is just a guess to show an example.
ggod luck! Oli
Thank you very much Oliver. I really appreciate your reply. Fanny
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