Soluble oligomers of Aβ1-42 (synthesized by China Peptides Co., Ltd.) were prepared as previously described [26 (link)]. Aβ was dissolved in hexafluoroisopropanol and dried in a speed-vacuum centrifuge. Before use, the aliquots were dissolved in dimethyl sulfoxide (DMSO) and then diluted in PBS to a final concentration of 400 μM/mL. The solution was allowed to oligomerize for 24 h at 4 °C.
Aβ1 42
Manufactured by ChinaPeptides
Sourced in China
Aβ1–42 is a peptide product offered by ChinaPeptides. It is a 42-amino acid fragment of the amyloid-beta protein. This peptide is commonly used in research applications.
Automatically generated - may contain errors
Lab products found in correlation
Appswe, by Jackson ImmunoResearch (1 mentions)
Ps1m146v, by Jackson ImmunoResearch (1 mentions)
Taup301l, by Jackson ImmunoResearch (1 mentions)
Baicalin, by Chengdu Must Bio-Technology (1 mentions)
Wogonoside, by Chengdu Must Bio-Technology (1 mentions)
Milli q integral water purification system, by Merck Group (1 mentions)
Baicalein, by Chengdu Must Bio-Technology (1 mentions)
Nativepage sample buffer, by Thermo Fisher Scientific (1 mentions)
Annexin 5 staining kit, by BD (1 mentions)
Ez link nhs lc lc biotin, by Thermo Fisher Scientific (1 mentions)
Pvdf membrane, by Pall Corporation (1 mentions)
α lac, by Merck Group (1 mentions)
Ovalbumin (ova), by Merck Group (1 mentions)
Bovine serum albumin (bsa), by Merck Group (1 mentions)
9 protocols using aβ1 42
1
Preparation of Soluble Aβ Oligomers
2
Alzheimer's Disease Mouse Model Characterization
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Aβ1–42, Aβ1–42NT, Aβ1–42DM, Aβ1–42I, Aβ1–42Cl, and Aβ1–42NF (>95%, lyophilized powder) are synthesized by China Peptides (Shanghai, China). Thioflavin T (ThT), 4,4′-dianilino-1,1′-binaphthyl-5,5′-disulfonic acid (Bis-ANS) are obtained from Sigma–Aldrich (St. Louis, MO, USA). All other reagents are in the analytical grade. The female triple transgenic model mice of AD carrying human gene mutants APPswe, PS1M146V, and tauP301L are purchased from the Jackson Laboratory (Bar Harbor, ME, USA).
3
Preparation of Amyloid-Beta Copper Aggregates
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Aβ1–40 and Aβ1–42 were purchased from ChinaPeptides Co., Ltd. The peptides were dissolved in hexafluoroisopropanol (HFIP) and stored at –20 °C as the stock solution. Before use, the solvent HFIP was evaporated under a gentle stream of nitrogen and Aβ was re-dissolved in HEPES buffer (20 mM, pH 7.4). For preparing Aβ40–Cu aggregates, 100 μM Aβ40 was incubated with an equimolar amount of CuCl2 at 37 °C for 8 h and then diluted to the corresponding concentration before use. For preparing Aβ42–Cu aggregates, 100 μM of Aβ42 was incubated with an equimolar amount of CuCl2 at 37 °C for 4 h and then diluted to the corresponding concentration before use. The Aβ–Cu species were dialyzed to remove any free Cu before the click reaction.
4
Baicalin, Wogonoside, and Baicalein Effects
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Baicalin, wogonoside and baicalein (≥98% purity, HPLC) were obtained from Chengdu MUST Bio-technology Company Ltd. (Chengdu, China). SB with the place of origin in Ji Lin province of China was purchased from Beijing Tong Ren Tang Zhuhai Pharmacy Co., Ltd. (Guangdong, China). 3-(4,5-dimethylthiazol-2-yl)-2,5-dimethyltetrazolium bromide (MTT) and ThT were purchased from Sigma (St. Louis, MO, United States). Milli-Q water was prepared by the Milli-Q integral water purification system (Millipore, Billerica, MA, United States) in our laboratory. Acetonitrile was purchased from Anaqua Chemicals Supply (Houston, TX, United States). Aβ (1–42) was obtained from the China Peptides Co., Ltd. (Shanghai, China). EZ-Link NHS-LC-LC-Biotin was obtained from Thermo Scientific Waltham (MA, United States). Super Streptavindin (SSA) biosensors were purchased from FortéBIO, PALL Life Sciences (Port Washington, NY, United States). The Annexin V staining kit was purchased from BD Biosciences (San Jose, CA, United States). The native PAGE Bis-Tris Gel (4–16%), running buffer (20X) and the native PAGE sample buffer were obtained from Invitrogen (Carlsbad, CA, United States). The PVDF membrane was obtained from PALL Life Sciences (Port Washington, NY, United States).
5
Purification and Characterization of Amyloidogenic Proteins
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α-Lac, alcohol dehydrogenase (ADH), BSA, CS, OVA, and CPK were purchased from Sigma-Aldrich. Purified human SOD was a gift from L. McAlary (Illawarra Health and Medical Research Institute, University of Wollongong, Australia). Aβ1-42 was purchased from China Peptides (Shanghai, China) or Australian Biosearch (Perth, Australia). ccβω, a modified version of the ccβ peptide (43 (link)) that has an additional tryptophan at its N terminus, was purchased from GenScript (Piscataway, NJ, USA). The ccβ peptide was purpose-designed to provide an experimentally convenient model system to study amyloid formation (43 (link)); the addition of the N-terminal tryptophan increases the rate at which the peptide forms amyloid and allows determination of the concentration of the peptide in solution by measurement of its absorbance at 280 nm (36 (link)). Human plasma CLU was purified by immunoaffinity chromatography of plasma prepared from human blood donated by Wollongong Hospital (Wollongong, Australia), as described before (44 (link)). Purified hTTR and a stable monomeric mutant of hTTR that carries F87M and L110M mutations and is unable to form a tetramer (mTTR) were produced as previously described (13 (link), 14 (link)).
6
Amyloid-beta Fibril Preparation
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Aβ1–42 fibrils were prepared as previously described (Jekabsone et al., 2006). Briefly, 1 mg of Aβ1–42 (China Peptides, Shanghai, China) was dissolved in 100 µL of phosphate-buffered saline (PBS) and then cultivated at 37°C for 7 days to accelerate aggregation into fibrous peptides. Hyp powder (98% purity; Chengdu ManSiTe Biotechnology, Sichuan, China) was dissolved in PBS and diluted to 8 mM.
7
Preparation of Amyloid-Beta Peptides
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Wild-type full-length Aβ1–42 and reverse control Aβ42–1 (ChinaPeptides, Shanghai) were prepared at a nominal concentration of 100 µM by dissolving known weights of peptides in mild alkali (0.1% ammonium hydroxide) in milliQ water to avoid isoelectric precipitation. Both solutions were then centrifuged at 100,000 × g for 3 h, which readily pellets fibrils and protofibrils, and the upper 75% of the supernatant taken. An aliquot of the supernatant was reserved to estimate the relative peptide concentration using the micro BCA protein assay (Thermo-Fisher Scientific Life Science Research Products, Rockford, IL). Then the concentrations of the remaining supernatants of Aβ1–42 and reverse control Aβ42–1 were adjusted to 75 µM stock solutions and stored at −80 °C until required.
8
Intracerebral Injection of Amyloid-Beta
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Saline (vehicle) and Aβ1-42 (ChinaPeptides, Shanghai, China) in saline (25 µM) was delivered into the lateral ventricle of mice at the rate of 0.5 µl/min for 40 min (500 pmol/mouse) via a microinjection canula as described previously [16 (link)].
9
Preparation of Amyloid-Beta Aggregates
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Aβ25–35 and Aβ1–42 (ChinaPeptides Co., Shanghai, China) were prepared as previouly described22 (link),54 (link),55 (link). The Aβ25–35 was dissolved in sterile double-distilled water (3 mM) and incubated at 37 °C for 4 days to form “aged” Aβ25–35. To form Aβ1–42 fibrils, Aβ1–42 was dissolved in double-distilled water (10 mM) and incubated at 37 °C for 24 h. To prepare Aβ1–42 oligomers, Aβ1–42 was dissolved in dimethyl sulfoxide (DMSO, St. Louis, MO, USA) (5 mM) and incubated at 4 °C for 24 h. Aβ aggregates were diluted to the final concentration with phosphate buffer saline (PBS) just before each experiment.
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