Fig. 6

Anti-SOD1^int antibody specifically detects pathological SOD1 in spinal cords of ALS-model mice. a, b SOD1 species recognized by a anti-SOD1^int and b anti-SOD1 (FL-154, Santa Cruz Biotechnology) antibody were quantified in the soluble fraction of the homogenates of lumbar spinal cord (red), cervical spinal cord (green), brainstem (blue), and cerebellum (gray) of G1H mice by sandwich ELISA. Three independent mouse samples at 30, 60, 100, 140, and 160 days of age were examined, and the averages were shown with error bars (standard deviation). ** (red and green) represents the P value less than 0.01 versus the data on lumbar and cervical spinal cords at 30 days of age, respectively. c Soluble disulfide-crosslinked SOD1 oligomers in mice were examined by Western blotting. Lumbar spinal cords of WT and G1H mice were homogenized in the presence of 100 mM iodoacetamide and 1% NP-40 and centrifuged at 20,000 x g for 30 min so as to prepare soluble supernatant. In the presence and absence of the reducing reagent, β-ME, the supernatant was then separated in a polyacrylamide gel by SDS-PAGE and probed by Western blot using anti-SOD1 antibody (FL-154, Santa Cruz Biotechnology). GAPDH was used as a protein loading control for Western blot