Wnt Signaling pp 119-125 | Cite as

Use of Primary Calvarial Osteoblasts to Evaluate the Function of Wnt Signaling in Osteogenesis

Part of the Methods in Molecular Biology book series (MIMB, volume 1481)


In vitro culture and genetic manipulation of primary calvarial cell cultures is a convenient and robust system to investigate gene function in osteoblast differentiation. We have used this system to study the functions of many genes in the Wnt signaling pathway within osteoblasts. Here, we describe a detailed protocol outlining the establishment and characterization of primary calvarial cells from mice carrying a conditionally inactivatable allele of the Wntless (Wls) gene (Wlsflox/flox). We previously used this approach to delete the Wntless gene by infecting with a Cre-expressing adenovirus, and to evaluate the effects of Wnt signaling loss on osteogenic potential in osteogenic medium with ascorbic acid. This detailed protocol is adaptable to use with any floxed allele.

Key words

Calvarial cell Osteogenic differentiation Adenovirus Alkaline phosphatase Alizarin red 


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Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  1. 1.Program for Skeletal Disease and Tumor Microenvironment and Center for Cancer and Cell BiologyVan Andel Research InstituteGrand RapidsUSA

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