Preparation of Amyloid-β Peptide

Synthetic human Aβ₄₂ peptide was prepared in two separate ways. Firstly the Aβ₄₂ lyophilised powder was dissolved in TFA and dried under a nitrogen stream. The remaining film was dissolved in 100% HFIP to a concentration of 1 mg/ml, sonicated 5 min in a bath sonicator and dried under a nitrogen stream. The HFIP treatment was repeated twice more and on final dissolving the peptide was dispensed into microcentrifuge tubes. After drying under a nitrogen stream, the peptide was further dried under vacuum for 1–2 h to give a clear film.
The second method was to take the original peptide and dissolve it in 10% (w/v) NH₄OH at 0.5 mg/ml. The peptide was incubated for 10 min at room temperature followed by sonication (5 min) and then dispensed (0.5 ml) into microfuge tubes. The NH₄OH was removed by lyophilisation to yield a salt free fluffy white peptide. All aliquots from both methods were then stored at −80°C. Immediately prior to use, the HFIP- and NH₄OH-treated and untreated Aβ₄₂ were dissolved in 60 mM NaOH and the concentration determined by absorbance at 214 and 280 nm using extinction coefficients of 76848 M^-1 cm^-1 or 1490 M^-1 cm^-1, respectively. In all cases the resuspended peptide was analysed by mass spectrometry and the peptide found to be unmodified with no trace of HFIP or NH₄OH.