University of Debrecen

6 ARTICLES PUBLISHED IN JoVE

Developmental Biology

Prediction and Validation of Gene Regulatory Elements Activated During Retinoic Acid Induced Embryonic Stem Cell Differentiation

Zoltan Simandi^*1, Attila Horvath^*2, Peter Nagy¹, Laszlo Nagy^1,2,3

¹Sanford-Burnham-Prebys Medical Discovery Institute at Lake Nona, ²Department of Biochemistry and Molecular Biology, Research Center for Molecular Medicine, Medical and Health Science Center, University of Debrecen, ³MTA-DE “Lendulet” Immunogenomics Research Group, University of Debrecen

In this work we provide an experimental workflow of how active enhancers can be identified and experimentally validated.

Biochemistry

Use of Recombinant Fusion Proteins in a Fluorescent Protease Assay Platform and Their In-gel Renaturation

Beáta Bozóki^*1,2, János András Mótyán^*1, Márió Miczi¹, Lívia Diána Gazda¹, József Tőzsér¹

¹Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, ²Biotechnology Research Department, Gedeon Richter Plc

Here, we present the detailed procedure of a recently developed protease assay platform utilizing N-terminal hexahistidine/maltose-binding protein and fluorescent protein-fused recombinant substrates attached to the surface of nickel-nitrilotriacetic acid magnetic agarose beads. A subsequent in-gel analysis of the assay samples separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis is also presented.

Bioengineering

Three-Dimensional Echocardiographic Method for the Visualization and Assessment of Specific Parameters of the Pulmonary Veins

Csaba Jenei¹, Laszlo Nagy¹, Reka Urbancsek¹, Daniel Czuriga¹, Zoltan Csanadi¹

¹Division of Cardiology, Department of Cardiology, Faculty of Medicine, University of Debrecen

The dimensions of the pulmonary veins (PV) are important parameters when planning pulmonary vein isolation. 2D transoesophageal echocardiography can only provide limited data about the PVs; however, 3D echocardiography can evaluate relevant diameters and areas of the PVs, as well as their spatial relationship to surrounding structures.

Biology

Assessing Protein Interactions in Live-Cells with FRET-Sensitized Emission

György Vámosi¹, Sarah Miller², Molika Sinha², Maria Kristha Fernandez², Gabor Mocsár¹, Malte Renz²

¹Department of Biophysics and Cell Biology, Faculty of Medicine, University of Debrecen, ²Gynecologic Oncology Division, Stanford University School of Medicine

Förster Resonance Energy Transfer (FRET) between two fluorophore molecules can be used for studying protein interactions in the living cell. Here, a protocol is provided as to how to measure FRET in live cells by detecting sensitized emission of the acceptor and quenching of the donor molecule using confocal laser scanning microscopy.

Cancer Research

High-Content Screening Assay for the Identification of Antibody-Dependent Cellular Cytotoxicity Modifying Compounds

Eliza Guti^*1,2, Ákos Máté Bede^*1,3, Csongor Váróczy^*1,3, Csaba Hegedűs¹, Máté Á. Demény^1,4, László Virág^1,4

¹Department of Medical Chemistry, Faculty of Medicine, University of Debrecen, ²Doctoral School of Molecular Medicine, University of Debrecen, ³National Academy of Scientist Education, University of Debrecen, ⁴ELKH-DE Cell Biology and Signaling Research Group

This protocol presents an automated, image-based high-throughput technique to identify compounds modulating natural killer cell-mediated breast cancer cell killing in the presence of a therapeutic anti-HER-2 antibody.

Cancer Research

Quantifying Antibody-Dependent Cellular Cytotoxicity in a Tumor Spheroid Model: Application for Drug Discovery

Isotta Sturniolo^1,2, Csongor Váróczy^1,3, Ákos Máté Bede^1,3, Csaba Hegedűs¹, Máté Á. Demény^1,4, László Virág^1,4

¹Department of Medical Chemistry, Faculty of Medicine, University of Debrecen, ²Doctoral School of Molecular Medicine, University of Debrecen, ³National Academy of Scientist Education, ⁴HUN-REN-DE Cell Biology and Signaling Research Group

Here, we present a method to identify compounds that modulate the ADCC mechanism, an important cancer cell-killing mechanism of antitumor antibodies. The cytotoxic effect of NK cells is measured in breast cancer cell spheroids in the presence of Trastuzumab. Image analysis identifies live and dead killer and target cells in spheroids.