The absence of ADAM22 protein in the Adam22 -/- mice was confirmed by Western blot analysis. Briefly, the cerebellum was isolated from a P14.5 mouse of each genotype and homogenized with a Polytron homogenizer in cell lysis buffer (50 mM Tris-HCl, pH 7.5, 100 mM NaCl, 1% NP-40, Complete protease inhibitors [Roche Diagnostics]). After removal of cell debris by centrifugation, the supernatant was separated on 10% SDS-PAGE, and transferred to a nitrocellulose membrane. The blot was then incubated with polyclonal antibody against the cytoplasmic domain of ADAM22 (at 1:1,000). Bound antibodies were visualized with horseradish peroxidase-labelled second antibody and a ECL-plus chemiluminescence detection system (Amersham Biosciences Corp.).