FIG. 2.

Levels of manganese superoxide dismutase (MnSOD) in amyotropic lateral sclerosis (ALS) patients. Human skin biopsy samples collected from the familial ALS (fALS) patients or from normal individuals were minced into small pieces in modified Eagle's medium (MEM) cell culture medium, and then transferred onto primary culture dish. After the tissues were attached to the flask, MEM was removed and supplemented with MEM containing 20% fetal bovine serum, 20 U/ml penicillin-streptomycin, 2 mM glutamine (Invitrogen). Cells at 80%–90% confluence were used for experiments. The cell lysates of the primary human skin fibroblasts obtained from ALS patients with R521G mutation in FUS gene or from normal individuals were subjected to 12.5% polyacrylamide native gel for MnSOD activity, which was then detected by staining the gel with nitro-blue-tetrazolium dye (A, top panel). The bands were densitometrically scanned and normalized with CuZnSOD (A, bottom panel). The cell lysates were also subjected to SDS-PAGE followed by Western blotting using MnSOD specific antibody to detect MnSOD protein levels (B, top panel). The protein levels were quantified by densitometric scanning of MnSOD bands followed by normalization with succinate dehydrogenase (SDHB) as an internal control (B, bottom panel). Each data point represents mean±SD of three independent samples and significant difference as compared to respective controls is indicated by **p<0.01. To see this illustration in color, the reader is referred to the web version of this article at www.liebertpub.com/ars