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Medical University Vienna

5 ARTICLES PUBLISHED IN JoVE

Bioengineering

Measuring TCR-pMHC Binding In Situ using a FRET-based Microscopy Assay

Markus Axmann¹, Gerhard J. Schütz¹, Johannes B. Huppa²

¹Institute for Applied Physics - Biophysics, Vienna University of Technology, ²Institute for Hygiene and Applied Immunology, Medical University of Vienna

This manuscript describes how to conduct (single molecule) Förster Resonance Energy Transfer (FRET)- based assays to measure the binding dynamics between T-cell antigen receptor (TCR) and antigenic peptide-loaded MHC molecules as they occur within the immunological synapse of a T-cell in contact with a functionalized planar supported lipid bilayer.

Bioengineering

Single Molecule Fluorescence Microscopy on Planar Supported Bilayers

Markus Axmann¹, Gerhard J. Schütz¹, Johannes B. Huppa²

¹Institute of Applied Physics - Biophysics, Vienna University of Technology, ²Institute for Hygiene and Applied Immunology, Immune Recognition Unit, Medical University of Vienna

Preparation of protein-functionalized planar glass-supported lipid bilayers, determination of protein mobility within and measurement of protein densities is shown here. A roadmap to building a noise-reduced Total Internal Reflection microscope is outlined, which allows visualizing single bilayer-resident fluorochromes with high spatiotemporal resolution.

Biology

A Rapid Automated Protocol for Muscle Fiber Population Analysis in Rat Muscle Cross Sections Using Myosin Heavy Chain Immunohistochemistry

Konstantin D. Bergmeister^1,3, Marion Gröger², Martin Aman¹, Anna Willensdorfer¹, Krisztina Manzano-Szalai¹, Stefan Salminger¹, Oskar C. Aszmann¹

¹CD Laboratory for the Restoration of Extremity Function, Division of Plastic and Reconstructive Surgery, Department of Surgery, Medical University of Vienna, ²Core Facility Imaging, Core Facilities, Medical University Vienna, ³Department of Hand, Plastic and Reconstructive Surgery, Burn Center, BG Trauma Center Ludwigshafen, Plastic and Hand Surgery, University of Heidelberg

Here, we present a protocol for rapid muscle fiber analyses, which allows improved staining quality, and thereby automatic acquisition and quantification of fiber populations using the freely available software ImageJ.

Biochemistry

Enrichment of Native Lipoprotein Particles with microRNA and Subsequent Determination of Their Absolute/Relative microRNA Content and Their Cellular Transfer Rate

Markus Axmann¹, Andreas Karner², Sabine M. Meier¹, Herbert Stangl¹, Birgit Plochberger²

¹Center for Pathobiochemistry and Genetics, Institute of Medical Chemistry and Pathobiochemistry, Medical University Vienna, ²School of Medical Engineering and Applied Social Sciences, University of Applied Sciences Upper Austria

Here, a quantitative real-time polymerase chain reaction-based protocol is presented for the determination of the native micro-RNA content (absolute/relative) of lipoprotein particles. In addition, a method for increasing the micro-RNA level, as well as a method for determining the cellular uptake rate of lipoprotein particles, is demonstrated.

Medicine

Targeted Muscle Reinnervation: Surgical Protocol for a Randomized Controlled Trial in Postamputation Pain

Emily Pettersen^1,2,3,4, Paolo Sassu⁵, Francesca Alice Pedrini^1,5, Hannes Granberg^1,2, Carina Reinholdt^2,6, Juan Manuel Breyer⁷, Aidan Roche⁸, Andrew Hart^9,10, Adil Ladak¹¹, Hollie A. Power¹¹, Michael Leung¹², Michael Lo¹², Ian Valerio¹³, Kyle R. Eberlin¹³, Theodore A. Kung¹⁴, Paul Cederna¹⁴, Jason M. Souza^15,16, Oskar Aszmann¹⁷, Jason Ko¹⁸, Gregory A. Dumanian¹⁸, Max Ortiz-Catalan^1,4,19,20

¹Center for Bionics and Pain Research, ²Center for Advanced Reconstruction of Extremities, Sahlgrenska University Hospital, ³Department of Electrical Engineering, Chalmers University of Technology, ⁴Bionics Institute, ⁵IV Clinica Ortoplastica, IRCCS Istituto Ortopedico Rizzoli, ⁶Department of Hand Surgery, Institute of Clinical Sciences, Sahlgrenska Academy, University of Gothenburg, Sahlgrenska University Hospital, ⁷Department of Orthopedic, Hand Unit, Worker Hospital, ⁸College of Medicine and Veterinary Medicine, The Queen's Medical Research Institute, The University of Edinburgh, ⁹Canniesburn Plastic Surgery Unit, Glasgow Royal Infirmary, ¹⁰College of Medicine, Veterinary & Life Sciences, The University of Glasgow, ¹¹Division of Plastic Surgery, Department of Surgery, Faculty of Medicine and Dentistry, University of Alberta, ¹²Plastics and Reconstructive Surgery, Dandenong Hospital, Monash Health, ¹³Division of Plastic and Reconstructive Surgery, Massachusetts General Hospital &, Harvard Medical School, ¹⁴Section of Plastic Surgery, Department of Surgery, Michigan Medicine, ¹⁵Department of Surgery, Uniformed Services University of the Health Sciences and Walter Reed National Military Medical Center, ¹⁶Department of Plastic and Reconstructive Surgery, Ohio State University, ¹⁷Clinical Laboratory for Bionic Extremity Reconstruction, Department of Plastic, Reconstructive and Aesthetic Surgery, Medical University Vienna, ¹⁸Division of Plastic Surgery, Department of Surgery, Northwestern Feinberg School of Medicine, ¹⁹Medical Bionics Department, University of Melbourne, ²⁰Prometei Pain Rehabilitation Center

The protocol outlines the surgical procedure for the treatment of postamputation pain using Targeted Muscle Reinnervation (TMR). TMR will be compared with two other surgical techniques, specifically Regenerative Peripheral Nerve Interface (RPNI) and neuroma excision, followed by immediate burying within muscle under the context of an international, randomized controlled trial.