β-Amyloid Synthesis, Purification, and Conformation Characterization

Human β-amyloid (1–42) (Aβ_1-42), provided by Dr. Charles Glabe (University of California at Irvine), was synthesized by fluoren-9-ylmethoxy carbonyl chemistry using a continuous flow semiautomatic instrument as described previously (Burdick et al., 1992). The peptide was reconstituted in filter-sterilized water at a concentration of 1 mM after which an equal volume of 2 × TBS (0.033 M Tris, 0.267 M NaCl) was added (final concentration 500 µM Aβ). After 20 to 24 hr at 4 ℃ to allow fibril formation, aliquots were frozen for future use. The peptide conformation was analyzed by circular dichroism (CD) to confirm β-sheet conformation. Briefly, after using 1 × TBS as a blank, 200 µl of the peptide at 50 µM was run on a Jasco J-720 CD spectrometer and read from 200 nm to 250 nm with a step resolution of 0.5 nm and a scan speed of 20 nm/min. Four scans were acquired and averaged to generate the CD spectra of the peptide (Supplemental Figure S1; Li et al., 2004).