Monitoring Vesicular Trafficking in Cellular Responses to Stress - Part B

1st Edition, Volume 165 - July 1, 2021
Editors: Oliver Kepp, Lorenzo Galluzzi
Language: English
Hardback ISBN:
9 7 8 - 0 - 1 2 - 8 2 4 4 8 7 - 6
eBook ISBN:
9 7 8 - 0 - 1 2 - 8 2 4 4 8 8 - 3

Monitoring Vesicular Trafficking in Cellular Responses to Stress - Part B, Volume 165 in the Methods in Cell Biology series, highlights new advances in the field, with this new… Read more

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Monitoring Vesicular Trafficking in Cellular Responses to Stress - Part B, Volume 165 in the Methods in Cell Biology series, highlights new advances in the field, with this new release presenting coverage of different topics, including A novel tool for detecting lysosomal membrane permeabilization by high-throughput fluorescence microscopy, Exploring selective autophagy in Drosophila, Assessment of EGFP-Q74 degradation for the measurement of autophagic flux, Multimodal assessment of autophagy in mammalian cells with a novel, LC3-based tandem reporter, Multiplex quantification of autophagic flux by imaging flow cytometry, Monitoring autophagic flux in Caenorhabditis elegans using p62/SQST-1 reporters, High throughput screening for autophagy, and much more.

Cover image
Title page
Table of Contents
Copyright
Contributors
Vesicular trafficking in cellular responses to stress
Chapter 1: A novel tool for detecting lysosomal membrane permeabilization by high-throughput fluorescence microscopy
Abstract
1: Introduction
2: Material
3: Concluding remarks
Acknowledgments
Chapter 2: Exploring selective autophagy in Drosophila: Methods to identify Atg8-interacting proteins
Abstract
1: Introduction
2: iLIR database
3: GST pull-down assay with in vitro translated protein as input and 35S labeled methionine
4: GST pull-down assay with in vitro translated GST-tagged protein (bait) and his-tagged protein (prey)
5: Mosaic analysis
6: Concluding remarks
7: Notes
Acknowledgments
Chapter 3: Assessment of EGFP-Q74 degradation for the measurement of autophagic flux
Abstract
1: Introduction
2: Materials
3: Concluding remarks
Acknowledgments
Chapter 4: Multimodal assessment of autophagy in mammalian cells with a novel, LC3-based tandem reporter
Abstract
1: Introduction
2: Material
3: Equipment
4: Reagents
5: Autophagy LC3 HiBiT reporter cells culture and preparation for assay
6: Homogeneous, bioluminescence plate-reader assay for reporter levels
7: Fluorescence imaging for reporter subcellular localization in living cells
8: Antibody-free, protein blotting for LC3-I to LC3-II reporter conversion
9: Concluding remarks
Acknowledgments
Chapter 5: Multiplexed quantification of autophagic flux by imaging flow cytometry
Abstract
1: Introduction
2: Materials
3: Methods
4: Data analysis
5: Concluding remarks
6: Notes
Acknowledgments
Chapter 6: Monitoring autophagic flux in Caenorhabditis elegans using a p62/SQST-1 reporter
Abstract
1: Introduction
2: Materials
3: Concluding remarks
Acknowledgments
Chapter 7: High throughput screening for autophagy
Abstract
1: Introduction
2: Materials
3: Methods
4: Concluding remarks
Chapter 8: Determination of mitophagy by electron microscope
Abstract
1: Introduction
2: Materials
3: Methods
4: Benefits and pitfalls
5: Concluding remarks
6: Notes
Acknowledgments
Chapter 9: Quantification of intracellular ACBP/DBI levels
Abstract
1: Introduction
2: Material
3: Treatments
4: Intracellular ACBP/DBI measurements
5: Concluding remarks
6: Notes
Acknowledgments
Chapter 10: Immunoblot-based assays for assessing autophagy in the turquoise killifish Nothobranchius furzeri
Abstract
1: Introduction
2: Animal model
3: Treatments and common procedures
4: Concluding remarks
5: Notes
6: Materials
Chapter 11: Monitoring autophagy using super-resolution structured illumination and direct stochastic optical reconstruction microscopy
Abstract
1: Introduction
2: Material
3: Concluding remarks
Acknowledgments
Chapter 12: Monitoring selective autophagy of mitochondria using super-resolution microscopy
Abstract
1: Introduction
2: Material
3: Concluding remarks
Chapter 13: Monitoring autophagy-dependent ferroptosis
Abstract
1: Introduction
2: Monitoring autophagy-dependent ferroptosis using western blotting
3: Monitoring autophagy-dependent ferroptosis using the lipid peroxidation production assay kit
4: Monitoring autophagy-dependent ferroptosis using the lipid peroxidation probe
5: Monitoring autophagy-dependent ferroptosis using the ferrous ion (Fe2 +) probe
6: Concluding remarks
Conflict of interest
Chapter 14: Quantitative analysis of autophagy-related protein LC3B by quantum-dot-based molecular imaging
Abstract
1: Introduction
2: Materials and methods
3: Concluding remarks
Acknowledgments
Chapter 15: Study of microRNAs carried by exosomes
Abstract
1: Introduction
2: Exosome isolation
3: Exosomal RNA isolation and detection
4: Equipment and reagents required
5: Conclusions
Chapter 16: Simultaneous determination of intraluminal lysosomal calcium and pH by dextran-conjugated fluorescent dyes
Abstract
1: Introduction
2: Materials
3: Concluding remarks
4: Notes
Acknowledgments

Oliver Kepp

Dr. Oliver Kepp graduated from Humboldt University Berlin and obtained a PhD degree in Molecular Biology. Today he is a tenured researcher at INSERM and a Co-director of the BioCell automation platform at GRCC. Oliver and his team investigate several aspects of tumor immunogenicity, focusing on systems cell biology approaches. He is also a co-founder of Samsara therapeutics and was appointed associate director of the European academy of tumor immunology in 2019. Oliver received the prestigious Clarivate Highly Cited Researcher Award four years in a row since 2018 and has recently been ranked among the top biology and biochemistry scientists in France (#5; www.research.com)

Affiliations and expertise

Equipe labellisée par la Ligue contre le cancer, Université de Paris, Sorbonne Université

Lorenzo Galluzzi

Lorenzo Galluzzi is Assistant Professor of Cell Biology in Radiation Oncology at the Department of Radiation Oncology of the Weill Cornell Medical College, Honorary Assistant Professor Adjunct with the Department of Dermatology of the Yale School of Medicine, Honorary Associate Professor with the Faculty of Medicine of the University of Paris, and Faculty Member with the Graduate School of Biomedical Sciences and Biotechnology of the University of Ferrara, the Graduate School of Pharmacological Sciences of the University of Padova, and the Graduate School of Network Oncology and Precision Medicine of the University of Rome “La Sapienza”. Moreover, he is Associate Director of the European Academy for Tumor Immunology and Founding Member of the European Research Institute for Integrated Cellular Pathology. Galluzzi is best known for major experimental and conceptual contributions to the fields of cell death, autophagy, tumor metabolism and tumor immunology. He has published over 450 articles in international peer-reviewed journals and is the Editor-in-Chief of four journals: OncoImmunology (which he co-founded in 2011), International Review of Cell and Molecular Biology, Methods in Cell biology, and Molecular and Cellular Oncology (which he co-founded in 2013). Additionally, he serves as Founding Editor for Microbial Cell and Cell Stress, and Associate Editor for Cell Death and Disease, Pharmacological Research and iScience.

Affiliations and expertise

Assistant Professor of Cell Biology in Radiation Oncology, Department of Radiation Oncology, Weill Cornell Medical College, NY, USA