optimize b-catenin topflash assay using wnt3a-CM

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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.