RNA in situ hybridisation
In situ hybridisation on frozen sections were carried out using 35S-labelled RNA probes. Briefly, 610 bp of mouse ADAM22 cDNA (position: 1351–1960 from initiating ATG) was cloned into the pBluescript II SK(+) or the pBluescript II KS(+) vector. Using these plasmids as templates, sense and antisense labelled RNA probes were generated by T7 RNA polymerase and [α35S]UTP. Frozen brain and spinal cord from 2 month-old C57BL/6 female mice were used in this analysis. Pretreatment, hybridisation, RNase treatment and washing was carried out following the protocol described in the literature [33]. Dehydrated slides were attached to imaging plates for 48 hours and the autoradiograms were analysed using a Bio-Image Analyser (BAS3000, Fuji Photo Film).