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amyloid level in mice...HELP PLEASE!!!
Posted by: Nina-Ann (IP Hidden, New member, 1)
Date: June 8, 2005 08:03AM
Hi everyone!
We have just started working with mousemodels for Alzheimer´s Disease in our lab. I was told to analyse the level of amyloid-beta42 vs amyloid-beta40 in brain homogenates by elisa or western blot analysis. So I looked through the web and was just stunned by the amount of available antibodies (and their prizes ^.°). Has anyone of you experience with this typ of experiments and can propably help me to choose the right antibody or Kit? I would really appreciate it. Thanks Nina-Ann!!!
Re: amyloid level in mice...HELP PLEASE!!!
Posted by: femmeauburn (IP Hidden, Advanced member, 115)
Date: June 8, 2005 12:02PM
i have done quite a bit of elisa and western blot, but never using the specific markers you asked about. in general it is best to acquire antibodies that are monoclonal or affinity selected, as these will be the most specific. sadly these tend to cost a little more.
Re: amyloid level in mice...HELP PLEASE!!!
Posted by: femmeauburn (IP Hidden, Advanced member, 115)
Date: June 8, 2005 12:06PM
one other thing, if the amyloids you are interested in are named by their molecular weights (e.g. 42 is a protein of 42 kDA) it will be very difficult to distinguish between the two on western blotting as their weights are so very similar. to be able to do western blot, you need to obtain separate antibodies for both of the proteins which are specific for that protein only. that is there can be no cross-reaction between the two. cross-reaction will interfere with your analysis of 42 vs 40.
Re: amyloid level in mice...HELP PLEASE!!!
Posted by: 5'-ATCG (IP Hidden, Unregistered user, )
Date: June 8, 2005 03:40PM
Just to clarify 40/42 means amino acid residues not molecular weight, so they are A-beta (1-40) and A-beta (1-42) peptides.
It will be much better to do IHC on brain sections or FPLC/HPLC analysis. Both elisa and Western are tricky (especially homogenate) and depend on the quality of antibody on cross-reactivity, sensitivity, specificity...you may need to spend sometime and some money before find a good one (if you're lucky).
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