Fig. 6.
Expression of clusterin in lumbar spinal cord of symptomatic mice transgenic for mutant (G93A) SOD1 in comparison to other markers of disease progression. Mice were assessed symptomatic by abnormal hindlimb splay reflex. WT SOD1 transgenic mice were analyzed as a control for overexpression of SOD1. Shown are both low magnification images and higher magnification of specific regions. 10 μm coronal cryostat sections from symptomatic aged mice (>140 days postnatal) were immunolabeled with antibodies to a and c b and d SOD1, b and c a and c clusterin (SC- 6420), d the small heat shock protein, αB-crystallin, e the microglial marker, Iba1, and f the astroglial marker, GFAP. Conditions for fixation and immunolabeling were optimized for each antibody (see Table 1). In a, b, d, e, and f, biotinylated secondary antibodies were used and developed using the peroxidase method. In c, fluorescent secondary antibodies were used. Arrows in a d point to inclusion bodies in sections from SOD1G93A mice immunolabeled by anti-SOD1, a pathological hallmark. b d Clusterin antibody strongly labeled inclusions in sections of lumbar spinal cord from these mice; c d labeling of sections with both antibodies revealed coexistence of clusterin and SOD1 in the same inclusions. d Inclusions positive for αB-crystallin were also present (arrows in d d). Antibodies against Iba1 (e) and GFAP (f) revealed activation of microglia and astrocytes, respectively, in SOD1G93A sections, a known pathological feature of disease in these mice. Characterization of abnormalities observed at different stages of disease are presented in Table 2