optimize b-catenin topflash assay using wnt3a-CM  1 reply [Last
post] Joined: 10/06/2011  What cell lines are effective for
topflash reporting induced with wnt3a-cm?  What is a reasonable
fold increase in transcription under optimal CM and time
conditions?   rnajdi (not verified) Hello,In my experience, I've
In my experience, I've found that HEK293 cells are perfect for
reporting TopFlash activity in response to Wnt-conditioned
media. I have not tried Wnt3A-conditioned media yet, but
Wnt1-conditioned media added for 24h on 293 cells results in
about a 50 to a 100-fold increase in reporter activity. It is
important to mix the conditioned media with fresh media before
adding it to the cells (1:1 ratio). Adding conditioned media
alone results in lower activation. I have also tried adding
purified Wnt3a (from Stem RD) on 293 cells. In this case,
incubation with Wnt3a for 4h results in a 30-fold increase. I
hope this helps.