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Tohoku University

15 ARTICLES PUBLISHED IN JoVE

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Biology

Identifying the Effects of BRCA1 Mutations on Homologous Recombination using Cells that Express Endogenous Wild-type BRCA1
Jeffrey Parvin 1, Natsuko Chiba 2, Derek Ransburgh 1
1Department of Biomedical Informatics, The Ohio State University, 2Departments of Molecular Immunology and Clinical Oncology, Tohoku University

We provide a method for testing BRCA1 variants in a tissue culture based assay for homologous recombination repair of DNA damage by depleting endogenous BRCA1 protein from a cell using RNAi and replacing it with a BRCA1 point mutant that contains a coding change.

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Immunology and Infection

Investigating Mast Cell Secretory Granules; from Biosynthesis to Exocytosis
Nurit P. Azouz 1,2, Mitsunori Fukuda 3, Marc E. Rothenberg 2, Ronit Sagi-Eisenberg 1
1Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel Aviv University, 2Division of Allergy and Immunology, Department of Pediatrics, Cincinnati Children's Hospital Medical Center, University of Cincinnati College of Medicine, 3Laboratory of Membrane Trafficking Mechanisms, Department of Developmental Biology and Neurosciences, Graduate School of Life Sciences, Tohoku University

The goal of the present protocol was to develop a method that will allow functional genomic analyses of mast cell secretion. The protocol is based on quantitative assessment of the release of a fluorescent reporter gene cotrasfected with the gene of interest and real time analyses of the secretory granule's morphology.

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Bioengineering

Photopatterning Proteins and Cells in Aqueous Environment Using TiO2 Photocatalysis
Hideaki Yamamoto 1,2, Takanori Demura 3, Kohei Sekine 3, Sho Kono 3, Michio Niwano 2,4, Ayumi Hirano-Iwata 2,5, Takashi Tanii 6
1Frontier Research Institute for Interdisciplinary Sciences, Tohoku University, 2CREST, Japan Science and Technology Agency, 3School of Fundamental Science and Engineering, Waseda University, 4Research Institute of Electrical Communication, Tohoku University, 5Graduate School of Biomedical Engineering, Tohoku University, 6Faculty of Science and Engineering, Waseda University

We describe a protocol for modifying cell affinity of a scaffold surface in aqueous environment. The method takes advantage of titanium dioxide photocatalysis to decompose organic film in the photo-irradiated region. We show that it can be used to create microdomains of scaffolding proteins, both ex situ and in situ.

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Medicine

Isolation and Profiling of MicroRNA-containing Exosomes from Human Bile
Ling Li 1, Klaus B. Piontek 1, Vivek Kumbhari 1, Masaharu Ishida 2, Florin M. Selaru 1
1Division of Gastroenterology and Hepatology, Department of Medicine, Johns Hopkins University School of Medicine, 2Department of Surgery, Tohoku University

Bile fluid is a valuable source of extracellular vesicles/exosomes that contain potentially important biomarkers. This protocol represents a robust method to isolate exosomes from human bile for further analyses including miRNA profiling.

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Developmental Biology

Immobilization of Caenorhabditis elegans to Analyze Intracellular Transport in Neurons
Shinsuke Niwa 1
1Frontier Research Institute for Interdisciplinary Sciences and Graduate School of Life Sciences, Tohoku University

Caenorhabditis elegans (C. elegans) is a good model to study axonal and intracellular transport. Here, I describe a protocol for in vivo recording and analysis of axonal and intraflagellar transport in C. elegans.

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Engineering

Growth and Electrostatic/chemical Properties of Metal/LaAlO3/SrTiO3 Heterostructures
Diogo Castro Vaz 1, Edouard Lesne 1,2, Anke Sander 1, Hiroshi Naganuma 1,3, Eric Jacquet 1, Jacobo Santamaria 1,4, Agnès Barthélémy 1, Manuel Bibes 1
1Unité Mixte de Physique CNRS/Thales, Université Paris-Saclay, 2Max Planck Institut für Mikrostrukturphysik, 3Department of Applied Physics, Tohoku University, 4Instituto de Magnetismo Aplicado, Universidad Complutense de Madrid

We fabricate metal/LaAlO3/SrTiO3 heterostructures using a combination of pulsed laser deposition and in situ magnetron sputtering. Through magnetotransport and in situ X-ray photoelectron spectroscopy experiments, we investigate the interplay between electrostatic and chemical phenomena of the quasi two-dimensional electron gas formed in this system.

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Bioengineering

Microhoneycomb Monoliths Prepared by the Unidirectional Freeze-drying of Cellulose Nanofiber Based Sols: Method and Extensions
Zheng-Ze Pan 1,2, Hirotomo Nishihara 3, Wei Lv 1, Cong Wang 1,2, Yi Luo 1,2, Liubing Dong 1,2, Houfu Song 1,4, Wenjie Zhang 2, Feiyu Kang 1,2,4, Takashi Kyotani 3, Quan-Hong Yang 1,4,5
1Engineering Laboratory for Functionalized Carbon Materials and Shenzhen Key Laboratory for Graphene-based Materials, Graduate School at Shenzhen, Tsinghua University, 2School of Materials Science and Engineering, Tsinghua University, 3Institute of Multidisciplinary Research for Advanced Materials, Tohoku University, 4Tsinghua-Berkeley Shenzhen Institute (TBSI), Tsinghua University, 5School of Chemical Engineering and Technology, Tianjin University

Here, we present a general protocol to prepare a variety of microhoneycomb monoliths (MHMs) in which fluid can pass through with an extremely low pressure drop. MHMs obtained are expected to be used as filters, catalyst supports, flow-type electrodes, sensors and scaffolds for biomaterials.

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JoVE Journal

Imaging FITC-dextran as a Reporter for Regulated Exocytosis
Ofir Klein 1, Amit Roded 1, Koret Hirschberg 2, Mitsunori Fukuda 3, Stephen J. Galli 4, Ronit Sagi-Eisenberg 1
1Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel Aviv University, 2Department of Pathology, Sackler Faculty of Medicine, Tel Aviv University, 3Laboratory of Membrane Trafficking Mechanisms, Department of Developmental Biology and Neurosciences, Graduate School of Life Sciences, Tohoku University, 4Departments of Pathology and of Microbiology and Immunology and Sean N. Parker Center for Allergy and Asthma Research, School of Medicine, Stanford University

Here we detail a method for live cell imaging of regulated exocytosis. This method utilizes FITC-dextran, which accumulates in lysosome-related organelles, as a reporter. This simple method also allows distinguishing between different modes of regulated exocytosis in cells that are difficult to manipulate genetically.

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Immunology and Infection

Assays for the Specific Growth Rate and Cell-binding Ability of Rotavirus
Syun-suke Kadoya 1, Daisuke Sano 1,2
1Department of Civil and Environmental Engineering, Tohoku University, 2Department of Frontier Science for Advanced Environment, Graduate School of Environmental Studies, Tohoku University

Here we present two protocols, one for measuring the specific growth rate and the other for the cell-binding ability of rotavirus using the plaque assay and RT-qPCR. These protocols are available for confirming the differences in phenotypes between rotavirus strains.

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Immunology and Infection

Application of Consistent Massage-Like Perturbations on Mouse Calves and Monitoring the Resulting Intramuscular Pressure Changes
Naoyoshi Sakitani *1, Takahiro Maekawa *1, Kumiko Saitou 1,2, Katsuhiko Suzuki 3, Shuhei Murase 1,4, Masakuni Tokunaga 1, Daisuke Yoshino 5, Keisuke Sawada 6, Atsushi Takashima 7, Motoshi Nagao 1, Toru Ogata 1, Yasuhiro Sawada 1,8
1Department of Rehabilitation for Motor Functions, National Rehabilitation Center for Persons with Disabilities, 2Graduate School of Sport Sciences, Waseda University, 3Faculty of Sport Sciences, Waseda University, 4Department of Orthopaedic Surgery, Graduate School of Medicine, The University of Tokyo, 5Frontier Research Institute for Interdisciplinary Sciences, Tohoku University, 6University of Cincinnati College of Medicine, 7Department of Assistive Technology, National Rehabilitation Center for Persons with Disabilities, 8Department of Clinical Research, National Rehabilitation Center for Persons with Disabilities

Here we describe the protocols for applying defined mechanical loads to mouse calves and for monitoring the concomitant intramuscular pressure changes. The experimental systems that we have developed can be useful for investigating the mechanism behind the beneficial effects of physical exercise and massage.

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JoVE Journal

Evaluation of Photosynthetic Behaviors by Simultaneous Measurements of Leaf Reflectance and Chlorophyll Fluorescence Analyses
Kaori Kohzuma 1
1Graduate School of Life Sciences, Tohoku University

We describe a new technical approach to study photosynthetic responses in higher plants involving simultaneous measurements of chlorophyll a fluorescence and leaf reflectance using a PAM and a spectral radiometer for the detection of signals from the same leaf area in Arabidopsis.

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Chemistry

Self-Assembly of Hybrid Lipid Membranes Doped with Hydrophobic Organic Molecules at the Water/Air Interface
Xingyao Feng 1, Teng Ma 2, Daisuke Tadaki 1, Ayumi Hirano-Iwata 1,2
1Research Institute of Electrical Communication, Tohoku University, 2Advanced Institute for Materials Research, Tohoku University

We report a protocol for producing a hybrid lipid membrane at the water/air interface by doping the lipid bilayer with copper (II) 2,9,16,23-tetra-tert-butyl-29H,31H-phthalocyanine (CuPc) molecules. The resulting hybrid lipid membrane has a lipid/CuPc/lipid sandwich structure. This protocol can also be applied to the formation of other functional nanomaterials.

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Behavior

A Common Marmoset Model of Mother-Infant Intervention for Breastfeeding Disorders in the Presence of Paternal Inhibition and Maternal Neglect
Mayuko Yoda 1,2, Yoshimasa Kamei 3, Hayato Sakurai 4, Hiroko Kakei 4, Ting Tao 1,2,4, Hideo Yamanouchi 4, Tetsuya Kunikata 4, Masanori Hariyama 5, Ricki Colman 6, Mamiko Koshiba 1,4,5
1Graduate School of Sciences and Technology for Innovation, Yamaguchi University, 2Faculty of Health Sciences and Nursing, Juntendo University, 3Obstetrician, Medical Department, Saitama Medical University, 4Department of Pediatrics, Saitama Medical University, 5Graduate School of Information Sciences, Tohoku University, 6Department of Cell and Regenerative Biology and Wisconsin National Primate Research Center, University of Wisconsin, Madison

Here, we present a nonhuman primate model of mother-infant intervention for breastfeeding disorders in the presence of paternal inhibition and maternal neglect. The mate model video complements education to support primate and human caregivers with respect to infants with breastfeeding problems such as pain.

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Behavior

Four Temporary Waterslide Designs Adapted to Different Slope Conditions to Encourage Child Socialization in Playgrounds
Zhudi Hua 1, Ting Tao 1, Risa Akita 1, Tomofusa Akita 1, Yoshiaki Hayakawa 1, Masanori Hariyama 2, Hayato Sakurai 3, Ricki Colman 4, Mamiko Koshiba 1,2,3
1Graduate School of Sciences and Technology for Innovation, Yamaguchi University, 2Graduate School of Information Sciences, Tohoku University, 3Pediatrics, Saitama Medical University Hospital, 4Department of Cell and Regenerative Biology, University of Wisconsin-Madison

Early life social learning is enhanced by interactions with effectively designed environments. Four events were held at different city parks using inexpensive, temporary waterslides to stimulate social learning. This study describes the prototypes used and the evaluation of the children's interactions.

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Developmental Biology

Fluorescent In Situ Hybridization and 5-Ethynyl-2'-Deoxyuridine Labeling for Stem-Like Cells in the Hydrozoan Jellyfish Cladonema pacificum
Sosuke Fujita 1, Erina Kuranaga 1, Masayuki Miura 2, Yu-ichiro Nakajima 2
1Graduate School of Life Sciences, Tohoku University, 2Graduate School of Pharmaceutical Sciences, The University of Tokyo

Here, we describe a protocol for visualizing stem-like proliferating cells in the jellyfish Cladonema. Whole-mount fluorescent in situ hybridization with a stem cell marker allows for the detection of stem-like cells, and 5-ethynyl-2'-deoxyuridine labeling enables the identification of proliferating cells. Together, actively proliferating stem-like cells can be detected

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