Ribolysor apparatus

For this experiment, the number of brains analyzed per group was as follows: WT CTR: $n=4$ ; WT T: $n=4$ ; J20 CTR: $n=5$ ; and J20 T $n=6$ (Figure 1C, Study 1). Hemi-brains were homogenized as described previously (Lafon et al. 2018 (link)). Briefly, frozen left hemispheres were homogenized in 20% wt/vol in a homogenization buffer [ $140\text{\hspace{0.17em} mM}$ potassium chloride (KCl), $10\text{\hspace{0.17em} mM}$ sodium hydrogen phosphate ( ${\mathrm{Na}}_2{\text{HPO}}_4$ ), $1.7\text{\hspace{0.17em} mM}$ monopotassium phosphate ( ${\text{KH}}_2{\text{PO}}_4$ ), and $1\text{\hspace{0.17em} mM}$ ethylenediaminetetraacetic acid (EDTA)] containing protease inhibitors (Complete Ultra, Roche) and phosphatases inhibitors (PhosStop™, Sigma). Homogenization was performed in microbead-containing tubes on a Ribolysor apparatus (Biorad). Samples were aliquoted and immediately frozen at $-80°\mathrm{C}$ until use. For the ${\mathrm{A}\mathrm{\beta }}_{1–42}$ assay, 2% sodium dodecyl sulfate (SDS) was added to brain homogenates before ultracentrifugation at $\mathrm{196,000}\times g$ (Beckman Optima TL100 centrifuge) for 30 min at 4°C. Supernatants obtained were used to quantify soluble fractions of human ${\mathrm{A}\mathrm{\beta }}_{1–42}$ peptides with an enzyme-linked immunosorbent assay kit (Invitrogen). Results were normalized according to their protein concentration determined by the bicinchoninic acid (BCA) method protein assay (Pierce Biotechnology).