Analysis of autophagy deficiency and cytotoxicity in autophagy-deficient human

被引：0

作者：

Korsgen, Miriam E. ^{[1
]}

Sun, Congxin ^{[1
]}

Seranova, Elena ^{[1
,4
]}

Zatyka, Malgorzata ^{[1
]}

Astuti, Dewi ^{[1
]}

Kataura, Tetsushi ^{[2
]}

Barrett, Timothy ^{[1
,3
]}

Korolchuk, Viktor I. ^{[2
]}

Sarkar, Sovan ^{[1
]}

机构：

[1] Univ Birmingham, Inst Canc & Genom Sci, Inst Biomed Res, Coll Med & Dent Sci, Birmingham B15 2TT, England

[2] Newcastle Univ, Biosci Inst, Fac Med Sci, Newcastle Upon Tyne NE4 5PL, England

[3] Birmingham Womens & Childrens Hosp, Dept Endocrinol, Steelhouse Lane, Birmingham B4 6NH, England

[4] NMN Bio Ltd, Birmingham B19 2HN, England

来源：

STAR PROTOCOLS | 2023年 / 4卷 / 03期

基金：

英国生物技术与生命科学研究理事会; 英国医学研究理事会; 英国惠康基金; 日本学术振兴会;

关键词：

STEM-CELLS; NEURODEGENERATION;

D O I：

10.1016/j.xpro.2023.102529

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Autophagy, a catabolic process governing cellular and energy homeostasis, is essential for cell survival and human health. Here, we present a protocol for generating autophagy-deficient (ATG5-/-) human neurons from human embryonic stem cell (hESC)-derived neural precursors. We describe steps for analyzing loss of autophagy by immunoblotting. We then detail analysis of cell death by luminescence-based cytotoxicity assay and fluorescence-based TUNEL staining. This hESC-based experimental platform provides a genetic knockout model for undertaking autophagy studies relevant to human biology.For complete details on the use and execution of this protocol, please refer to Sun et al. (2023).1

引用

页数：18

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