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  1. No Access

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    Induction of PINK1/Parkin-Mediated Mitophagy

    PINK1/Parkin mitophagy is a key mechanism to contribute mitochondrial quality control, and the defects are thought to be a cause of those Parkinson’s disease onsets. Upon loss of mitochondrial membrane potenti...

    Shigeto Sato, Norihiko Furuya (2017)

  2. No Access

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    A Simple Method of Generating 3D Brain Organoids Using Standard Laboratory Equipment

    3D brain organoids are a powerful tool with prospective application for the study of neural development and disease. Here we describe the growth factor-free method of generating cerebral organoids from feeder-...

    Magdalena Sutcliffe, Madeline A. Lancaster (2017)

  3. No Access

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    A Cleanup Method for Mass Spectrometric Analysis of Sphingosine- and Ceramide-1-Phosphate in Blood and Solid Tissue Using a Phosphate Capture Molecule

    Cleanup technology and mass spectrometric determination of sphingosine-1-phosphate (S1P) using a phosphate capture molecule are shown. The protocol is rapid, requires neither thin-layer chromatography nor liqu...

    Jun-ichi Morishige, Ryouhei Yamashita, Tamotsu Tanaka, Kiyoshi Satouchi (2017)

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    The Use of Correlative Light-Electron Microscopy (CLEM) to Study PINK1/Parkin-Mediated Mitophagy

    In this chapter we describe the use of correlative light-electron microscopy (CLEM) to study, in cultured cells, the turnover of damaged mitochondria by PINK1/Parkin-dependent mitophagy. CLEM combines the adva...

    Chieko Kishi-Itakura, Folma Buss (2017)

  5. No Access

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    Human Intestinal Enteroids: New Models to Study Gastrointestinal Virus Infections

    Human rotavirus (HRV) and human norovirus (HuNoV) infections are recognized as the most common causes of epidemic and sporadic cases of gastroenteritis worldwide. The study of these two human gastrointestinal ...

    Winnie Y. Zou, Sarah E. Blutt, Sue E. Crawford, Khalil Ettayebi, Xi-Lei Zeng (2017)

  6. No Access

    Protocol

    Immunocytochemical Monitoring of PINK1/Parkin-Mediated Mitophagy in Cultured Cells

    Both PINK1 and parkin are the responsible genes (PARK6 and PARK2, respectively) for familial early-onset Parkinson’s disease (PD). Several lines of evidences have suggested that mitochondrial dysfunction would...

    Motoki Fujimaki, Shinji Saiki, Yukiko Sasazawa, Kei-Ichi Ishikawa, Yoko Imamichi (2017)

  7. No Access

    Protocol

    Sphingosine-1-Phosphate (S1P) Signaling in Neural Progenitors

    Sphingosine-1-phosphate (S1P) and its receptors are important in nervous system development. Reliable in vitro human model systems are needed to further define specific roles for S1P signaling in neural develo...

    Phillip Callihan, Mohammed Alqinyah, Shelley B. Hooks (2017)

  8. No Access

    Protocol

    Measuring Sphingosine-1-Phosphate: Protein Interactions with the Kinetic Exclusion Assay

    By directly detecting the ligand-free binding sites in a sample, the kinetic exclusion assay (KinExA®) provides a compelling alternative to SPR-based techniques for determining equilibrium dissociation constants ...

    Jonathan K. Fleming, Jonathan M. Wojciak (2017)

  9. No Access

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    Investigation of Yeast Mitophagy with Fluorescence Microscopy and Western Blotting

    Selective clearance of superfluous or dysfunctional mitochondria is a fundamental process that depends on the autophagic membrane trafficking pathways found in many cell types. This catabolic event, called mit...

    Sachiyo Nagumo, Koji Okamoto (2017)

  10. No Access

    Protocol

    Exploring MicroRNAs on NIX-Dependent Mitophagy

    The dysregulation of autophagy is implicated in many pathological disorders including infections, aging, neurodegenerative diseases, and cancer. Autophagy can be precisely controlled both transcriptionally and...

    Wen Li, Hao Chen, Shupeng Li, Guanghong Lin, Du Feng (2017)

  11. No Access

    Protocol

    Intestinal Crypt Organoid: Isolation of Intestinal Stem Cells, In Vitro Culture, and Optical Observation

    The isolation and culture of intestinal stem cells (ISCs) was first demonstrated in the very recent decade with the identification of ISC marker Lgr5. The growth of ISCs into crypt organoids provides an in vit...

    Yun Chen, Chuan Li, Ya-Hui Tsai, Sheng-Hong Tseng (2017)

  12. No Access

    Protocol

    Quantum Dot Toolbox in Membrane Neurotransmitter Transporter Research

    Quantum dot-based fluorescence techniques enable multi-scale molecular profiling ranging from real-time single molecule dynamics to expression trends in million-cell populations. In comparison to currently ava...

    Lucas B. Thal, Danielle M. Bailey, Oleg Kovtun, Sandra J. Rosenthal (2017)

  13. No Access

    Protocol

    Monitoring Mitochondrial Changes by Alteration of the PINK1-Parkin Signaling in Drosophila

    Mitochondrial quality control is a key process in tissues with high energy demands, such as the brain and muscles. Recent studies using Drosophila have revealed that the genes responsible for familial forms of ju...

    Tsuyoshi Inoshita, Kahori Shiba-Fukushima, Hongrui Meng, Nobutaka Hattori, Yuzuru Imai (2017)

  14. No Access

    Protocol

    Mitophagy in Yeast: A Screen of Mitophagy-Deficient Mutants

    Mitochondrial autophagy (mitophagy) is a process that selectively degrades mitochondria via autophagy. Recent studies have shown that mitophagy plays an important role in mitochondrial homeostasis by degrading...

    Kentaro Furukawa, Tomotake Kanki (2017)

  15. No Access

    Protocol

    Maintenance of Human Embryonic Stem Cells by Sphingosine-1-Phosphate and Platelet-Derived Growth Factor

    Human embryonic stem cells (hESCs) have historically been cultivated on feeder layers of primary mouse embryonic fibroblasts (MEF) in a medium supplemented with fetal calf serum (FCS). However, serum contains ...

    Raymond C. B. Wong, Martin F. Pera, Alice Pébay (2017)

  16. No Access

    Protocol

    Detection of Hypoxia-Induced and Iron Depletion-Induced Mitophagy in Mammalian Cells

    Mitochondrial quality and quantity are not only regulated by mitochondrial fusion and fission but also by mitochondria degradation. Mitophagy, an autophagy specific for damaged or unnecessary mitochondria, is ...

    Shun-ichi Yamashita, Tomotake Kanki (2017)

  17. No Access

    Protocol

    Assessment of Mitophagy in iPS Cell-Derived Neurons

    Aberrant mitochondrial function is associated with many neurological diseases. Mitophagy is a key mechanism for the elimination of damaged mitochondria and maintenance of mitochondrial homeostasis. Induced plu...

    Kei-Ichi Ishikawa, Akihiro Yamaguchi, Hideyuki Okano, Wado Akamatsu (2017)

  18. No Access

    Protocol

    MitoPho8Δ60 Assay as a Tool to Quantitatively Measure Mitophagy Activity

    Mitophagy, a selective type of macroautophagy (hereafter referred to as autophagy), specifically mediates the vacuole/lysosome-dependent degradation of damaged or surplus mitochondria. Because this process reg...

    Zhiyuan Yao, Xu Liu, Daniel J. Klionsky (2017)

  19. No Access

    Protocol

    Flow Cytometer Monitoring of Bnip3- and Bnip3L/Nix-Dependent Mitophagy

    Mitochondria are organelles with numerous vital roles in cellular metabolism. Impaired or damaged mitochondria are degraded in autophagolysosomes in a process known as mitophagy. Given the fundamental role of ...

    Matilda Šprung, Ivan Dikic, Ivana Novak (2017)

  20. No Access

    Protocol

    Monitoring Mitophagy During Aging in Caenorhabditis elegans

    Mitochondria constitute the main energy-producing centers of eukaryotic cells. In addition, they are involved in several crucial cellular processes, such as lipid metabolism, calcium buffering, and apoptosis. ...

    Nikolaos Charmpilas, Konstantinos Kounakis, Nektarios Tavernarakis (2017)

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