I am Isolating mouse placental cells and immuno staining them for some nuclear proteins which are very rare. he anibody I use is monoclonal and I try to remove most of the decidual tissues before proceeding to digest the placenta. However, the non-specific binding is giving me a lot of problem while analyzing the endpoint of the experiment. Does anyone know how to reduce the non-specific binding when dealing with whole placental cells?
Clonality has nothing to do with your issue. If you use mouse AB on Mouse tissue, then you have a problem.
I use the following solutions:
http://www.dako.com/dist/ar38/p107160/prod_products.htm
Or
https://www.vectorlabs.com/catalog.aspx?prodID=1757
In case the biotin load of the sample is an issue.
Thanks all. I am using e18 placentas. I think its basically arising due to large percentage of mouse IgG to which the Ab binds. Still figuring out.
If it is not a autofluorescence problem, you could block this tissue with a variety of things, such as 0.05% fish skin gelatin in PBS, 3% BSA in PBS or add 1.5% of donkey normal serum. These blocking procedures are usually good to reduce non-specific binding.
Thanks Alexandre. I have tried all types of blocking. the last staining I did looks far better. hope I am getting there. Thanks
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