avidin, streptavidin, neutravidin ??? - confused!! (Feb/24/2010 )

Hi guys, wasn't too sure where to post this so decided that general should be okay.

I'm looking to buy some fluorescently labelled streptavidin to detect biotinylated antibodies, and got myself confused between Avidin, Streptaviding, and Neutravidin. As far as I can see they all do the same, so not too sure how to decide which one to buy (other than the price).

So, can anyone tell me a) What’s the actual difference, What’s the best option? Any thoughts will be greatly appreciated.
I want to use them for western blot, in case that helps making a decision.

Thanks!!!!

almost a doctor on Feb 24 2010, 10:03 AM said:

I can't tell you the difference, but I can offer a little advice. I'm currently using a streptavidin/HRP conjugate along with a primary and biotin-labeled secondary to detect proteins in whole cell lysates. If you're doing anything with a heterogeneous mix of proteins from pro- or eukaryotes, be prepared for the possibility that the streptavidin conjugate will bind endogenously biotinylated proteins in your lysate. I've recently learned the way around that is to block the membrane normally, then also block with avidin/streptavidin/neutravidin, then follow up with a biotin block to cover all the biotin-binding sites of the avidin so the biotinylated antibody doesn't stick to them. It will add a little bit of time to the procedure, as well as a little cost as I've found that avidin/streptavidin/neutravidin are not exactly cheap.

If you think this could be an issue, there is a protocol from Pierce: http://www.piercenet.com/files/TR0016-Bloc...nous-biotin.pdf

Of course, if you're working with a purified protein, this shouldn't be a problem.

neutravidin is made from avidin (removal of carbohydrate, near neutral pI). binding is more specific than with avidin.

streptavidin has a greater affinity for biotin so it is bound tighter than with avidin (less chance of the complex falling apart over a range of harsh conditions).