How many times do you reuse a primary antibody? - (May/25/2005 )
On an average how many time do you reuse your primary?and secondary? Does it depend on the company?? Also do you just stick it in the refrigerator or add some preservatives to keep it stable. Someone told me that adding sodium azide helps....
are you talking about antibodies???
if so it's single use for us.
i've recently known about micro-o-protect (0,05%) and that's equivalent as sodium azide, but as i mind the environment, i prefer not to use sodium azide which is VERY toxic...
generally i reuse antibodies several times (up to 5-6 times), but only in case of abundant proteins such p53 or TBP.
regarding case sensitive antibodies, i do single use.
I find that different antibodies have very different stabillity so it depends on the antibody. With one of my primary antibodies it is very definately once only in Western, with others I have reused for a couple of months without any reduction in sensitivity (tested against fresh antibody). To stop microbial contamination I add 0.1% Sodium Azide when I make up the antibody. Also I only reuse primary antibodies, secondary antibodies are comparatively cheap although I'm sure with the excess they are in you could reuse a couple of times.
Finally I don't reuse antibodies in histology only Western.
Hope this helps,
Yes Nick, I am talking about antibodies for westerns.
Thanks a lot ..scott , fred...
I reuse primary and secondary antibodies up to 3-4 times for western blotting, but not usually if they are older than about 2 weeks since dilution. I use western breeze kits from invitrogen, that say that you shouldn't use diluted kit solutions over a day old, but I have done very successfully up to a week old for general reagents.
I reused anti-His and anti-GST antibodies (Amersham) more than 4 times for Western Blot. I diluted antibodies in PBST and restored at 4oC for the next times.
i've re-used primary Ab over 5 times before, over the course of 2.5 months, seems to work fine for abundant proteins. for less abundant, 3 times max. also i add 0.01-0.02% NaN3 after the first use. never had a problem.