Aβ25-35 is not recommended to be dissolved directly in triple-distilled water, and here are the reasons (see bolded notes). Below is a reference protocol for Aβ preparation and aging, along with some common issues you may encounter:

1. Aβ Aging Protocol (references [1–4])

(1) Monomerization

• Dissolve Aβ42 in HFIP (1 mg/mL, incubate at room temperature ≥1 h) to generate monomers.
• Evaporate HFIP under vacuum to form a transparent peptide film at the tube bottom. The film can be stored frozen at −20 °C or below.

Potential issues:

• TFE can be used as an alternative to HFIP [2, 4, 5].
• Water-based solutions are not recommended: dissolving Aβ42 directly in ddH₂O produces a mixture of monomers, oligomers, protofibrils, and fibrils [2].
• If neither HFIP nor TFE is available, Aβ can be dissolved directly in DMSO and aged; in 100% DMSO, Aβ exists mainly as monomers, though short fibrils may form over time [2].
• Another option is to sonicate Aβ42 (0.5 mg/mL) in 10% NH₄OH before aging [6].
• As a last resort, direct dissolution in water followed by biological experiments is possible, but not recommended as the aggregation state of Aβ will be uncertain [2].

(2) Resuspension

• Before use, resuspend HFIP-treated Aβ42 film in DMSO to 5 mM, producing a uniformly dispersed monomer solution.

Potential issues:

• Do not replace DMSO with H₂O; solubility is poor [2].
• Low-concentration NaOH can also dissolve Aβ42 [4, 6], then dilute with PBS for aging.

(3) Aging

• To form oligomers: dilute the resuspended solution with F12 medium [2,3] or sterile PBS [1,4] to 100 µM, incubate at 4 °C for 24–48 h.
• To form fibrils: dilute with 10 mM HCl and incubate at 37 °C for 24 h.

Potential issues:

• Neutral/weakly basic pH (e.g., 7.4) favors oligomer formation, whereas acidic conditions favor fibril formation [1, 2, 4].
• Aggregation state depends on incubation time: 100 µM Aβ42 at 4 °C for 1 week can form fibrils [2].
• Temperature also influences aggregation: 100 µM Aβ42 at 37 °C produces fibrils within hours [1, 2].

2. Other Notes

• Solubility: Aβ dissolves quickly in HFIP. If some aggregates remain after 1 h, they may be insoluble deposits; remove by centrifugation or extend incubation [2].
• Storage: It is best to store Aβ as lyophilized powder or HFIP-prepared peptide films at −20 °C or −80 °C. Use aged solutions freshly; do not store as stock solutions.
• Aβ40 and Aβ25-35: Their aging protocols are similar to Aβ42, but typically require longer incubation times [2, 3, 7].

References:

1. Ahmed, Mahiuddin, et al. Nature structural & molecular biology 17.5 (2010): 561-567.

2. Stine, W. Blaine, et al. Journal of Biological Chemistry 278.13 (2003): 11612-11622.

3. Stine, W. Blaine, et al. Alzheimer's Disease and Frontotemporal Dementia: Methods and Protocols (2011): 13-32.

4. WO2007005359A1

5. Zhang-Haagen, Bo, et al. PloS one 11.2 (2016): e0150267.

6. Ryan, Timothy M., et al. PeerJ 1 (2013): e73.

7. Hu, Yueqiang, et al. Human & Experimental Toxicology 40.12_suppl (2021): S347-S358.