MOESM5 of Wild-type Cu/Zn-superoxide dismutase is misfolded in cerebrospinal fluid of sporadic amyotrophic lateral sclerosis

Additional file 5: Figure S4. Almost complete depletion of C4F6-reactive SOD1 from CSF by immunoprecipitation. C4F6-crosslinked magnetic beads were first incubated with the CSF of ALS2 (40 μL) at 4 °C for 24 h, and the solution was collected as the “unbound” fraction in 1st IP. The remaining magnetic beads were then treated with 10 μL of 100 mM citrate buffer at pH 3.1, and the eluate was collected as the “bound” fraction in 1st IP. The unbound fraction (40 μL) in the 1st IP step was further treated with C4F6-crosslinked magnetic beads at 4 °C for 24 h, and the solution was collected as the unbound fraction in 2nd IP. The remaining magnetic beads were then treated with 10 μL of 100 mM citrate buffer at pH 3.1, and the eluate was collected as the bound fraction in 2nd IP. Ten microliter of the CSF (input) as well as the unbound and bound fractions in 1st IP and 2nd IP steps were analyzed by Western blotting with FL-154 antibody. The full-length and truncated SOD1 proteins were indicated with filled and open arrows, respectively.