Monitoring Vesicular Trafficking in Cellular Responses to Stress

1st Edition, Volume 164 - June 30, 2021
Editors: Oliver Kepp, Lorenzo Galluzzi
Language: English
Hardback ISBN:
9 7 8 - 0 - 1 2 - 8 2 3 5 4 4 - 7
eBook ISBN:
9 7 8 - 0 - 1 2 - 8 2 3 5 4 5 - 4

Monitoring Vesicular Trafficking in Cellular Responses to Stress, Volume 164 in the Methods in Cell Biology series, highlights new advances in the field, with this new volume pre… Read more

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Monitoring Vesicular Trafficking in Cellular Responses to Stress, Volume 164 in the Methods in Cell Biology series, highlights new advances in the field, with this new volume presenting interesting chapters on a variety of timely topics. Each chapter is written by an international board of authors.

Cover image
Title page
Table of Contents
Copyright
Contributors
Vesicular trafficking in cellular responses to stress
Chapter 1: Monitoring TFEB translocation
Abstract
1: Introduction
2: Disposable material
3: Equipment
4: Reagents
5: Cell culture and passage
6: Immunofluorescence
7: Concluding remarks
8: Notes
Acknowledgment
Chapter 2: Monitoring lipophagy in kidney epithelial cells in response to shear stress
Abstract
1: Introduction
2: Materials
3: Methods
4: Conclusion
5: Notes
Acknowledgments
Chapter 3: Live cell imaging of LC3 dynamics
Abstract
1: Introduction
2: Material
3: Methods
4: Concluding remarks
5: Notes
Chapter 4: Autophagy assessment in circulating leukocytes
Abstract
1: Introduction
2: Material
3: Induction of autophagy by starvation in mice and blood collection
4: Leukocytes isolation, fixation and permeabilization
5: Sample preparation and immunostaining
6: Image flow cytometric acquisition and data analysis
7: Notes
8: Concluding remarks
Acknowledgments
Chapter 5: Measuring lysosomal size and frequency by electron microscopy
Abstract
1: Introduction
2: Material
3: Workflow
4: Concluding remarks
Notes
Acknowledgment
Chapter 6: Autophagic flux assessment by immunoblot
Abstract
1: Introduction
2: Material
3: Concluding remarks
4: Notes
Chapter 7: Assessing autophagic flux in yeast
Abstract
1: Introduction
2: GFP liberation assay
3: ALP activity assay
4: Concluding remarks
5: Notes
Chapter 8: Multiple analysis of mitochondrial metabolism, autophagy and cell death
Abstract
1: Introduction
2: Cell death assays
3: Autophagy monitoring
4: Mitochondrial network analysis
5: Real time bioenergetic profile analysis
Notes
Acknowledgments
Chapter 9: Quantitative determination of mitophagy by fluorescence microscope
Abstract
1: Introduction
2: Material
3: GFP-LC3 and MitoTracker red approach
4: Concluding remarks
Acknowledgments
Chapter 10: Quantifying phagosomal HOCl at single immune-cell resolution
Abstract
1: Introduction
2: Protocol
3: Summary
Acknowledgments
Chapter 11: The intracellular metabolome of starving cells
Abstract
1: Introduction
2: Materials
3: Methods
4: Quality control
5: Concluding remarks
6: Notes
Chapter 12: Quantitative determination of autophagy flux by probes
Abstract
1: Introduction
2: Material
3: Concluding remarks
4: Notes
Chapter 13: Assessment of mammalian endosomal microautophagy
Abstract
1: Introduction
2: Biochemical procedures for the assessment of eMI activity in mammals
3: Image-based procedures for assessment of eMI activity in cultured cells
4: Criteria to identify a protein as potential eMI substrate
5: Concluding remarks
Acknowledgments
Chapter 14: Measuring oxidation within LC3-associated phagosomes that optimizes MHC class II restricted antigen presentation
Abstract
1: Introduction
2: Materials
3: Protocol I: Measurement of the oxidation level within LAPosomes in human macrophages
4: Protocol II: Antigen presentation assay using whole blood CD4+ T cells
5: Concluding remarks
6: 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