Fig. 2. NRADD expression in mouse tissues. (A) Characterization of an anti-NRADD antibody. 293 cells were transfected with full length p75^NTR and NRADD FLAG-tagged at the N-terminus. Cell lysates from N2a or transfected 293 cells were analysed by three identical western blots probed with anti-NRADD, antigen blocked anti-NRADD and anti-FLAG antibodies. The N2a and 293 lanes were exposed to film for 5 minutes and the transfected 293 lanes were exposed for 30 seconds. (B) Mouse tissues were homogenized and separated into membrane (M) and soluble (S100) fractions. An aliquot of each fraction was deglycosylated in vitro with PNG-F or Endo H. Deglycosylation converted the fuzzy mNR band into a darker dglNR band. sNR indicates the small NRADD isoform. N.S. indicates a non-specific band in the brain lysates that was not blocked by preincubation of the antibody with antigen.