Materials and Reagents

1. NIH3T3 cells or HEK293 cells
   
2. Purified active Wnt3a proteins (ATCC, catalog number: CRL­2647 TM)
   
3. Fetal bovine serum (FBS)
4. DMEM (high glucose) (Life Technologies, Invitrogen^TM, catalog number: 11965 )
   
5. TransIT-2020 (Mirus Bio LLC, catalog number: MIR5404 )
   
6. TCL/LEF-Firefly luciferase expression construct (Addgene, catalog number: 12456 )
   
7. pRL-SV40-Renilla luciferase expression construct
   Note: The pRL-SV40-Renilla luciferase expression construct is made at Dr. Beachy’s Laboratory.
   
8. Dual-Luciferase @ Reporter Assay System (Promega Corporation, catalog number: E1960 )
   
9. Growth media (see Recipes)
   
10. Wnt3a-conditioned media (see Recipes)
    
11. Serum-deprived media (see Recipes)
    

Equipment

1. T75 culture flask (BD Biosciences, Falcon^®, catalog number: 353135 )
   
2. 24-well tissue culture plate (BD Biosciences, Falcon^®, catalog number: 353047 )
   
3. 96-well flat-bottom plate (Corning, Costar^®, catalog number: 3915 )
   
4. Centrifuge (Eppendorf, model: 8810R )
   
5. Water bath 
6. CO₂ incubator
   
7. Berthold Luminometer (Berthold Tecnologies, model: Centro XS LB960 )
   

Procedure