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Medical University of Graz

10 ARTICLES PUBLISHED IN JoVE

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Medicine

Oral Biofilm Analysis of Palatal Expanders by Fluorescence In-Situ Hybridization and Confocal Laser Scanning Microscopy
Barbara Klug 1,2, Claudia Rodler 1, Martin Koller 3, Gernot Wimmer 3, Harald H. Kessler 2, Martin Grube 4, Elisabeth Santigli 1
1Department of Orthodontics and Maxillofacial Orthopedics, Medical University of Graz, 2Institute of Hygiene, Microbiology and Environmental Medicine, Medical University of Graz, 3Department of Prosthodontics, Restorative Dentistry, Periodontology and Implantology, Medical University of Graz, 4Institute of Plant Sciences, Karl-Franzens-University Graz

We present a protocol for structural and compositional analysis of natural oral biofilm from orthodontic appliances with in situ hybridization (FISH) and confocal laser scanning microscopy (CLSM). Oral biofilm samples were collected from palatal expanders, scraping acrylic-resin flakes off their surface and referring them for molecular processing.

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Biology

Application of Genetically Encoded Fluorescent Nitric Oxide (NO•) Probes, the geNOps, for Real-time Imaging of NO• Signals in Single Cells
Emrah Eroglu 1, Rene Rost 1, Helmut Bischof 1, Sandra Blass 1, Anna Schreilechner 1, Benjamin Gottschalk 1, Maria R. Depaoli 1, Christiane Klec 1, Suphachai Charoensin 1, Corina T. Madreiter-Sokolowski 1, Jeta Ramadani 1, Markus Waldeck-Weiermair 1, Wolfgang F. Graier 1, Roland Malli 1
1Institute of Molecular Biology and Biochemistry, Medical University of Graz

This manuscript presents protocols for the application of novel genetically encoded nitric oxide (NO•) probes (geNOps) to monitor single cell NO• fluctuations in real-time using fluorescence microscopy. The Ca2+-triggered NO• formation on the level of individual endothelial cells was visualized by combining geNOps with a chemical Ca2+ sensor.

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Medicine

Oral Biofilm Sampling for Microbiome Analysis in Healthy Children
Elisabeth Santigli 1, Martin Koller 2, Barbara Klug 1
1Division of Oral Surgery and Orthodontics, Department of Dental Medicine and Oral Health, Medical University of Graz, 2Division of Preventive and Operative Dentistry, Periodontology, Prosthodontics and Restorative Dentistry, Department of Dental Medicine and Oral Health, Medical University of Graz

Changes in the oral microbiome throughout childhood are of growing interest. Comparison of different microbiome studies reveals a lack of standardized sampling protocols. Limited space makes sampling the sound subgingival sulcus of children challenging. Paper point sampling is presented here in detail as the method of choice for this area.

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Cancer Research

Target Cell Pre-enrichment and Whole Genome Amplification for Single Cell Downstream Characterization
Shukun Chen *1, Amin El-Heliebi *1, Julia Schmid 1, Karl Kashofer 2, Zbigniew T. Czyż 3, Bernhard Michael Polzer 3, Klaus Pantel 4, Thomas Kroneis 1,5, Peter Sedlmayr 1
1Institute of Cell Biology, Histology and Embryology, Medical University of Graz, 2Institute of Pathology, Medical University of Graz, 3Fraunhofer Institute for Toxicology and Experimental Medicine ITEM, 4Department of Tumor Biology, University Medical Center Hamburg-Eppendorf, 5Sahlgrenska Cancer Center, University of Gothenburg

This protocol is to recover and prepare rare target cells from a mixture with non-target background cells for molecular genetic characterization at the single-cell level. DNA quality is equal to non-treated single cells and allows for single-cell application (both screening based and targeted analysis).

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Medicine

Echocardiographic Measurement of Right Ventricular Diastolic Parameters in Mouse
Bakytbek Egemnazarov 1, Grazyna Kwapiszewska 1,2, Leigh M. Marsh 1
1Ludwig Boltzmann Institute for Lung Vascular Research, 2Department of Physiology, Otto Loewi Research Center, Medical University of Graz

Here we describe and compare two positions for obtaining the apical four-chamber view in mice. These positions enable the quantification of the right ventricular function, provide comparable results, and can be used interchangeably.

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Neuroscience

An Unbiased Approach of Sampling TEM Sections in Neuroscience
Stefan Wernitznig 1, Florian Reichmann 2, Mariella Sele 1, Christoph Birkl 3, Johannes Haybäck 4,5, Florian Kleinegger 4, Anna Birkl-Töglhofer 4, Stefanie Krassnig 4, Christina Wodlej 4, Peter Holzer 2, Daniel Kummer 1, Elisabeth Bock 1, Gerd Leitinger 1
1Department of Cell Biology, Histology and Embryology, Gottfried Schatz Research Center, Medical University of Graz, 2Department of Pharmacology, Otto Loewi Research Center, Medical University of Graz, 3Division of General Neurology, Department of Neurology, Medical University of Graz, 4Institute of Pathology, Medical University of Graz, 5Department of Pathology, Medical Faculty, Otto von Guericke University Magdeburg

We introduce a novel workflow for electron microscopy investigations of brain tissue. The method allows the user to examine neuronal features in an unbiased fashion. For elemental analysis, we also present a script that automatizes most of the workflow for randomized sampling.

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Medicine

Detection of Vascular Pathways of Oral Mucosa Influencing Soft- and Hard Tissue Surgeries by Latex Milk Injection
Arvin Shahbazi 1,2, Ulrike Pilsl 3, Bálint Molnár 2, Georg Feigl 3
1Department of Anatomy, Histology and Embryology, Semmelweis University, 2Department of Periodontology, Semmelweis University, 3Department of Macroscopical and Clinical Anatomy, Medical University of Graz

Here, we describe the staining of blood vessels using latex milk injections. This procedure provides anatomical knowledge of the oral blood supply for detailed macroscopic vascular mapping of the oral mucosa, as well as understanding of proper flap design to prevent complications and promote postoperative wound healing.

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Cancer Research

Exploring the Arginine Methylome by Nuclear Magnetic Resonance Spectroscopy
Hansjörg Habisch *1, Fangrong Zhang *1,2, Qishun Zhou 1, Tobias Madl 1
1Gottfried Schatz Research Center for Cell Signaling, Metabolism and Aging, Molecular Biology and Biochemistry, Medical University of Graz, 2Ministry of Education, School of Basic Medical Sciences, Key Laboratory of Gastrointestinal Cancer (Fujian Medical University)

The present protocol describes the preparation and quantitative measurement of free and protein-bound arginine and methyl-arginines by 1H-NMR spectroscopy.

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Cancer Research

Engineering Oncogenic Heterozygous Gain-of-Function Mutations in Human Hematopoietic Stem and Progenitor Cells
Tommaso Sconocchia 1, Johannes Foßelteder 1, Thomas Köhnke 2, Ravindra Majeti 2, Andreas Reinisch 1,3
1Division of Hematology, Department of Internal Medicine, Medical University of Graz, 2Division of Hematology, Stanford Cancer Institute, Stanford Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, 3Department of Blood Group Serology and Transfusion Medicine, Medical University of Graz

Novel strategies to faithfully model somatic mutations in hematopoietic stem and progenitor cells (HSPCs) are necessary to better study hematopoietic stem cell biology and hematological malignancies. Here, a protocol to model heterozygous gain-of-function mutations in HSPCs by combining the use of CRISPR/Cas9 and dual rAAV donor transduction is described.

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Developmental Biology

Ex Vivo Placental Explant Flow Culture - Mimicking the Dynamic Conditions In Utero
Nadja Kupper 1, Elisabeth Pritz 1, Monika Siwetz 1, Jacqueline Guettler 1, Berthold Huppertz 1
1Division of Cell Biology, Histology and Embryology, Gottfried Schatz Research Center, Medical University of Graz

Here is a protocol for culturing placental explants under constant flow conditions. This approach enhances traditional static villous culture systems by enabling the replication of dynamic physiological environments.

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