Promoter activity evaluation by EYFP - (Nov/27/2009 )

Hi, everyone.
I want to evaluate promoters in vivo by means of using EYFP as a reporter gene. We don't have a fluorimeter in our lab, however we have a VersaDoc system (BIORAD). Is it possible to use this system for that purpose? Do you have any protocol or reference that could help me?

Thanks,
Fernando

Hi Fernando,
I never heard of using a Versadoc to do live imaging. You did not say what animal but i am assuming you mean in mice--true?

We just got a new Biorad imager and when the rep comes to demonstrate it i can ask them if it can be used for in vivo imaging, but i don't think so. If you are imaging gene expression in mice, however, you will have to set up an anesthesia device to keep them immobilized for the 10 min or so that it takes to form an image. We used a luciferase reporter and Xenogen imaging system to monitor gene expression in mice but that has a set-up for anesthesia built into it.

You could call BioRad's tech support and ask them or you could just try it and see if the EYFP is visible.

Let me know what happens.

Best wishes,
Ergo

Hi Fernando,

me too I wonder about the possibilities of versadoc. In fact, it's true the opposite: with bioluminescence imaging workstations it is also possible to image western blots with high sensitivity (you acquire the HRP-luminescence of the secondary instead to develop the film).

Why EYFP? let's try infrared fluorescent protein (even if my feelings is that is still better luciferase at least in vivo)

Eventually, i.p. injections of ketamine-xilazine can be used as surrogate anesthethic if the isofluorane apparatus is not available.

best

96well
--my blog on reporter genes----