Immunoblot Analysis of Protein Targets

Immunoblot analysis was performed as previously described (44 (link)). Antibodies were used as follows: anti-USP25 (Abcam, Cambridge, MA, USA; ab187156, 1:1000), anti-APP (Millipore, Billerica, MA, USA; MAB348, 1:1000), anti-Iba1 (Wako Pure Chemical, Osaka, Japan; 016-20001, 1:500), anti-GFAP (glial fibrillary acidic protein) (Cell Signaling Technology, Danvers, MA, USA; 3670, 1:1000), anti–β-III-tubulin (Abcam, ab18207, 1:1000), anti-HA (hemagglutinin) (Sigma-Aldrich, St. Louis, MO, USA; H6908, 1:1000), anti-ubiquitin (Santa Cruz Biotechnology, Santa Cruz, CA, USA; sc-8017, 1:500), anti–β-actin (Xmbcss, Xiamen, China; bc001, 1:2000), and horseradish peroxidase (HRP)–conjugated secondary antibodies (Thermo Fisher Scientific, 31430 or 31460; 1:3000).

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Zheng Q., Li G., Wang S., Zhou Y., Liu K., Gao Y., Zhou Y., Zheng L., Zhu L., Deng Q., Wu M., Di A., Zhang L., Zhao Y., Zhang H., Sun H., Dong C., Xu H, & Wang X. (2021). Trisomy 21–induced dysregulation of microglial homeostasis in Alzheimer’s brains is mediated by USP25. Science Advances, 7(1), eabe1340.

Publication 2021

anti-USP25 anti-APP anti-Iba1 anti-GFAP anti–β-III-tubulin anti-HA (hemagglutinin) anti-ubiquitin horseradish peroxidase (HRP)–conjugated secondary antibodies

Actin Antibodies Glial fibrillary acidic protein Hemagglutinin Horseradish peroxidase Immunoblot analysis Tubulin Ubiquitin

Corresponding Organization : Xiamen University

Other organizations : Xizang Minzu University, Children’s Hospital of Fudan University Xiamen Branch, Tsinghua University, First Affiliated Hospital of Xiamen University

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Variable analysis

independent variables

Antibodies used in immunoblot analysis

dependent variables

Protein expression levels

control variables

Immunoblot analysis protocol as previously described (44 (link))

controls

Positive control: None specified
Negative control: None specified

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