I am working on a project trying to detect a GFP in intestine stem cells. I tried Abcan 290 GFP antiboday staining but failed. It works once for endogenouse GFP signal on cryosection for the same mice. The problem came out when I try to repeat cryosection on different mice that I can' t got endogenous signal on cryosection. Would you guys recommend a successful protocal for cryosection? And a protocal for paraffin embedded section is also needed.
Hi Jun Yi, I know sometimes it is difficult to detect the GFP labelling, especially when you have a strong autofluorescence of the tissue. In that case I recommand to use the ABC-DAB technique. After incubation of the GFP-antibody (I use the rb GFP-Alexa488 form ThermoScientific ;#A21311 1:500 - 1:750), incubate the sections in a biotinylated secondary antibody (in my case b-Goat-Anti-Rabbit) for 90 min, then in ABC (Vector Labs) or streotavidin-POD for 90 min and finally in DAB (Diaminobenzidine; brown color) If you like you can counterstain your sections with Haematoxyline. This technique will help you to find out weather your mice are really transfected.
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