Why should some antibodies not be aliquoted? - (Oct/31/2007 )
Hello,
Does anyone know why some antibodies (e.g. anti-phospho-AMPK Thr172) are not supposed to be aliquoted according to their datasheet?
Thanks!
not aliquoted or not frozen?
Hello Dominic,
I use anti-phospho-AMPK Thr 172 to detect my protein of interest (phosphoryltaed AMPK).
It says on the datasheet not to aliquote it. I understand aliquoting helps us 'maintain' that stability of the antibody if we use it numerous times.
I didn't aliquot my antibody. I take the vial in and out of the fridge all the time. But now I;m starting to think that it's better to aliquote it since my western blot results are getting more and more strange (multiple bands started to appear, suggesting antibody's starting to be partly denatured).
So,, I was wondering why it says "do not aliquote" in the 1st place.
Thanks in advance for any help!
One more thing!!
Eventhough I'm looking at phosphorylation, the antibody datasheet also encourages using milk as blocking agent.
Anyone has experience working with this Ab?
I don't know why they write do not aliquote. maybe they simply mean you don't need to aliquote?
About milk and antiphospho antibodies : if the antibody is directed against a specific phosphorylated protein, then milk is not a problem, because this protein is not in the milk. But if it's an antibody directed against any phosphorylated protein like 4G10 or PY20, then the antibody can recognize the phosphorylated proteins in the milk and you should avoid milk as blotting agent.
i agree with missele - you only really need to aliquot if you are avoiding freeze thaw cycles, but you said fridge not freezer so you should be ok leaving it in the origional bottle - i doubt this is the cause of your weird results.
dom
maybe it has to do with maintaining the concentration of the Ab? perhaps in small volumes it is more likely that the Ab will be adsorbed to the tube thus changing the concentration? Perhaps the original tube is somehow treated or this is taken into account so they suggest leaving the Ab in the original tube and not aliquotting in order to prevent adsorbtion...
HTH
HTH
or evaporation (increasing concentration)?
Does anyone know why some antibodies (e.g. anti-phospho-AMPK Thr172) are not supposed to be aliquoted according to their datasheet?
Thanks!
I think it is negative propaganda to accelerate consumption; I always aliquot and freeze any Ab; if I do not know the buffer I add glycerol and increase ionic strength
I agree with the Bearer.
If there are several users of the same antibody, I prefer to aliquote, to avoid a big contamination.