Amyloid-Beta Oligomerization with APOE Isoforms

Aβ₄₂ peptide (American Peptide Company), HiLyte^™ Fluor 488-Aβ₄₂, or HiLyte^™ Fluor 555-Aβ₄₂ (AnaSpec) were used for all Aβ oligomerizations and injections^{27 (link)}. Under a fume hood, 0.1 mg Aβ peptide was dissolved in ice cold 1,1,1,3,3,3-Hexafluoro-2-Propanol (HFIP, Fluka) then vortex for a few seconds. The solution was dried with a gentle stream of nitrogen to obtain a peptide film at the bottom of the vial. Prior to use, the film was resuspended in anhydrous dimethyl sulfoxide (DMSO) to form a 5 mM solution, sonicated in a water bath for 10 min and diluted in sterile phosphate-buffered saline (PBS) to a final concentration of 100 μM. HiLyte^™ Fluor 488- or 555-Aβ or Aβ₄₂ peptide and E3 or E4 native particles were combined, vortexed, held under oligomer forming conditions (room temperature) for 24 h at a final concentration of 50 μM Aβ and 5 μM APOE and stored at −20 ^oC until use (abbreviated AβE3 or AβE4). The same molar concentration (10:1) of scrambled Aβ (AnaSpec) was dissolved in vehicle, combined with isolated E3 or E4 particles, held under oligomer forming conditions and stored at −20 ^oC until use (abbreviated scrAβE3 or scrAβE4). scrAβE3 or scrAβE4 was utilized as negative controls throughout the subsequent experiments. Furthermore, Aβ₄₂ peptide and scrambled Aβ was incubated with PBS vehicle as negative control for APOE particles.