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Hebrew University of Jerusalem

10 ARTICLES PUBLISHED IN JoVE

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JoVE Journal

Fluorescence in situ Hybridizations (FISH) for the Localization of Viruses and Endosymbiotic Bacteria in Plant and Insect Tissues
Adi Kliot 1,2, Svetlana Kontsedalov 1, Galina Lebedev 1, Marina Brumin 1, Pakkianathan Britto Cathrin 1, Julio Massaharu Marubayashi 1, Marisa Skaljac 1,3, Eduard Belausov 4, Henryk Czosnek 2, Murad Ghanim 1
1Department of Entomology, Volcani Center, 2Institute of Plant Sciences and Genetics in Agriculture, Robert H. Smith Faculty of Agriculture, Food and Environment, Hebrew University of Jerusalem, 3Department of Applied Sciences, Institute for Adriatic Crops and Karst Reclamation, 4The Institute of Plant Sciences, Volcani Center

We describe here a simple fluorescence in situ hybridization (FISH) method for the localization of viruses and bacteria in insect and plant tissues. This protocol can be extended for the visualization of mRNA in whole mount and microscopic sections.

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Neuroscience

Slice It Hot: Acute Adult Brain Slicing in Physiological Temperature
Lea Ankri 1, Yosef Yarom 1, Marylka Y. Uusisaari 1
1Department of Neurobiology, Hebrew University of Jerusalem

In this paper we show a method for preparing acute brain slices in physiological temperature, using a conventional physiological solution without special modifications for the cutting (such as adding sucrose) and without intracardial perfusion of the animal before slice preparation.

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Medicine

A Brain Tumor/Organotypic Slice Co-culture System for Studying Tumor Microenvironment and Targeted Drug Therapies
Emily J. Chadwick 1, David P. Yang 1, Mariella G. Filbin 2, Emanuele Mazzola 3, Yu Sun 1, Oded Behar 1,4, Maria F. Pazyra-Murphy 1, Liliana Goumnerova 5, Keith L. Ligon 6, Charles D. Stiles 1, Rosalind A. Segal 1
1Department of Cancer Biology, Dana-Farber Cancer Institute, 2Department of Pediatrics, Children's Hospital, 3Department of Biostatistics & Computational Biology, Dana-Farber Cancer Institute, 4Department of Developmental Biology and Cancer Research, Hebrew University of Jerusalem, 5Department of Neurosurgery, Children's Hospital, 6Center for Molecular Oncologic Pathology, Department of Medical Oncology, Dana-Farber Cancer Institute

Many types of human brain tumors are localized to specific regions within the brain and are difficult to grow in culture. This protocol addresses the role of tumor microenvironment and investigates new drug treatments by analyzing fluorescent primary brain tumor cells growing in an organotypic mouse brain slice.  

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Biology

High Resolution Quantification of Crystalline Cellulose Accumulation in Arabidopsis Roots to Monitor Tissue-specific Cell Wall Modifications
Yulia Fridman 1, Neta Holland 1, Rivka Elbaum 2, Sigal Savaldi-Goldstein 1
1Faculty of Biology, Technion-Israel Institute of Technology, 2Smith Institute of Plant Sciences and Genetics in Agriculture, Hebrew University of Jerusalem

Crystalline cellulose is an important constituent of the plant cell wall. However, its quantification at a cellular resolution is technically challenging. Here, we report the use of polarized light technology and root cross sections to obtain information of cell wall composition at a spatiotemporal resolution.

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Genetics

Genome-wide Determination of Mammalian Replication Timing by DNA Content Measurement
Yishai Yehuda 1, Britny Blumenfeld 1, Dan Lehmann 2, Itamar Simon 1
1Dept. of Microbiology and Molecular Genetics, IMRIC, Faculty of Medicine, Hebrew University of Jerusalem, 2The Core Research Facility, IMRIC, Faculty of Medicine, Hebrew University of Jerusalem

We describe here a relatively fast and simple approach for mapping genome-wide mammalian replication timing, from cell isolation to the basic analysis of the sequencing results. A genomic map of a representative replication program will be provided following the protocol.

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Developmental Biology

Fluorimetric Techniques for the Assessment of Sperm Membranes
Alisa Komsky-Elbaz 1,2, Zvi Roth 1,2
1Animal Sperm Research Center, Department of Animal Sciences, Robert H. Smith Faculty of Agriculture, Food and Environment, Hebrew University of Jerusalem, 2Department of Animal Sciences, Robert H. Smith Faculty of Agriculture, Food and Environment, Hebrew University of Jerusalem

Here, we present methodologies to evaluate spermatozoan membrane integrity, a cellular feature associated with sperm fertilization competence. We describe three techniques for the fluorimetric assessment of sperm membranes: simultaneous staining with specific fluorescent probes, fluorescence microscopy, and advanced sperm-dedicated flow cytometry. Examples of combining the methodologies are also presented.

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Biology

Inducing Apical Periodontitis in Mice
Elisheva Goldman 1,2, Eli Reich 1, Itzhak Abramovitz *2, Michael Klutstein *1
1Institute of Dental Sciences, Faculty of Dental Medicine, Hebrew University of Jerusalem, 2Department of Endodontics, Hadassah Ein Kerem Medical Center

Here, we present a protocol to locally induce apical periodontitis in mice. We show how to drill a hole in the mouse's tooth and expose its pulp, in order to cause local inflammation. Analysis methods to investigate the nature of this inflammation, such as micro-CT and histology, are also demonstrated.

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Biology

Investigating Cardiac Metabolism in the Isolated Perfused Mouse Heart with Hyperpolarized [1-13C]Pyruvate and 13C/31P NMR Spectroscopy
David Shaul 1,2, Gal Sapir 1,2, Naama Lev-Cohain 1, Jacob Sosna 1, J. Moshe Gomori 1, Rachel Katz-Brull 1,2
1Department of Radiology, Hadassah Medical Organization and Faculty of Medicine, Hebrew University of Jerusalem, 2The Wohl Institute for Translational Medicine

We describe an experimental setup for administrating hyperpolarized 13C-labeled metabolites in continuous perfusion mode to an isolated perfused mouse heart. A dedicated 13C-NMR acquisition approach enabled the quantification of metabolic enzyme activity in real-time, and a multiparametric 31P-NMR analysis enabled the determination of the tissue ATP content and pH.

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Developmental Biology

Using Ex Vivo Live Imaging to Investigate Cell Divisions and Movements During Mouse Dental Renewal
Abinaya Sundari Thooyamani *1, Elias Shahin *2, Sanako Takano 1, Amnon Sharir 2, Jimmy K. Hu 1,3
1School of Dentistry, University of California Los Angeles, 2The Institute of Biomedical and Oral Research, Faculty of Dental Medicine, Hebrew University of Jerusalem, 3Molecular Biology Institute, University of California Los Angeles

Ex vivo live imaging is a powerful technique for studying the dynamic processes of cellular movements and interactions in living tissues. Here, we present a protocol that implements two-photon microscopy to live track dental epithelial cells in cultured whole adult mouse incisors.

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Biology

Intracameral Injection in Rats with Low Risk of Adverse Effects
Oriel Ratzon *1, Mor Schlesinger *1, Keren Ben-Yaakov 1, Ortal Zaks 1, Ziv Rotfogel 1,2, Arie L. Marcovich 1,2, Avital Eisenberg-Lerner 1
1Ophthalmology Research Laboratory, Kaplan Medical Center, 2Department of Ophthalmology, Kaplan Medical Center, Faculty of Medicine, Hebrew University of Jerusalem

This protocol describes a technique for intracameral injection in rats using a central corneal incision and a long tunnel into the anterior chamber. This injection method minimizes the risk of inducing inadvertent tissue damage and thereby improves precision and reproducibility.

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