Main description:

This detailed volume covers diverse aspects of P2X7 receptor analysis, ranging from its molecular structure to related pharmacological and immunological tools, via its analysis in heterologous expression systems as well as assays using primary cells and whole animal models. After three introductory chapters that focus on its structure, ligands, and physiological functions, the book details the generation of antibody and nanobody tools for P2X7 receptors, provides protocols for the analysis of expressed P2X7 receptors with a focus on their electrophysiological analysis, as well as protocols for the investigation of P2X7 down-stream signaling in immune cells by flow cytometry. Mouse models and procedures suited to investigate P2X7-mediated effects in other primary cells and in vivo are also explained. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls.
Authoritative and practical, The P2X7 Receptor: Methods and Protocols is a valuable reference not only for the growing community fascinated by this unusual ion channel but also for a broad readership interested in ion channels or purinergic receptors.

Contents:

1. How Structural Biology Has Directly Impacted Our Understanding of P2X Receptor Function and Gating

Steven E. Mansoor

2. Agonists, Antagonists, and Modulators of P2X7 Receptors

Christa E. Muller and Vigneshwaran Namasivayam

3. Modulation of Cell Energy Metabolism by the P2X7 Receptor

Francesco Di Virgilio, Simonetta Falzoni, Alba Clara Sarti, Paola Chiozzi, Valentina Vultaggio-Poma, and Anna Lisa Giuliani

4. Fully Flexible Ligand Docking for the P2X7 Receptor Using ROSIE

Sudad Dayl and Ralf Schmid

5. Preparation of the Murine Anti-Human P2X7 Receptor Monoclonal Antibody (Clone L4)

Amal Elhage, Ross J. Turner, Peter Cuthbertson, Debbie Watson, and Ronald Sluyter

6. Development of Antibody and Nanobody Tools for P2X7

Tobias Stahler, Welbeck Danquah, Melanie Demeules, Henri Gonde, Romain Hardet, Friedrich Haag, Sahil Adriouch, Friedrich Koch-Nolte, and Stephan Menzel

7. Enhanced Transduction of P2X7-Expressing Cells with Recombinant rAAV Vectors

Anna Marei Mann, Waldemar Schafer, Sahil Adriouch, Kathleen Boerner, Dirk Grimm, Ingke Braren, and Friedrich Koch-Nolte

8. Immunofluorescence Staining of P2X7 Receptors in Whole-Mount Myenteric Plexus Preparations

Tina Jooss, Birgit Kuch, Jiong Zhang, Michael Schemann, and Annette Nicke

9. Established Protocols for cRNA Expression and Voltage-Clamp Characterization of the P2X7 Receptor in Xenopus laevis Oocytes

Gunther Schmalzing and Fritz Markwardt

10. A Simplified Protocol to Incorporate the Fluorescent Unnatural Amino Acid ANAP into Xenopus laevis Oocyte-Expressed P2X7 Receptors

Anna Durner and Annette Nicke

11. Using Whole-Cell Electrophysiology and Patch Clamp Photometry to Characterize P2X7 Receptor Currents

Xin Liang, Laura Janks, and Terrance M. Egan

12. Using Symmetrical Organic Cation Solutions to Study P2X7 Ion Permeation

Kate Dunning, Laurie Peverini, and Thomas Grutter

13. Methods for Studying Cholesterol-Dependent Regulation of P2X7 Receptors

Ruth D. Murrell-Lagnado and Toshimitsu Kawate

14. Visualizing P2X7-Dependent Inflammasome Formation in Human Monocytes by Fluorescence Microscopy and Flow Cytometry

Samantha Wittke, Leif Ancker, Sana Javed, and Friedrich Haag

15. Characterization of P2X7 Receptors in Human Blood Cells

Laura Hurtado-Navarro, Alberto Baroja-Mazo, and Pablo Pelegrin

16. Real-Time Flow Cytometry as a Tool to Monitor Cellular Consequences of P2X7 Activation in Multiple Cell Populations Mixed in a Single FACS Tube

Alexander W. Veltkamp, Tim Magnus, and Bjoern Rissiek

17. Administration of P2X7 Receptor Blockers in Oncological Experimental Models

Elena De Marchi, Anna Pegoraro, and Elena Adinolfi

18. Humanized Mouse Model to Study the P2X7 Receptor in Graft-Versus-Host Disease

Debbie Watson, Sam R. Adhikary, Peter Cuthbertson, Nicholas J. Geraghty, Katrina M. Bird, Amal Elhage, Chloe Sligar, and Ronald Sluyter

19. Analysis of P2X7-Induced Neuronal Branching

Paula Mut-Arbona and Beata Sperlagh

20. Studying the Role of P2X7 Receptor in Axonal Growth Using In Utero Electroporation Technique

Alvaro Sebastian-Serrano, Carolina Bianchi, Caterina Di Lauro, Lucia Soria-Tobar, Beatriz Alvarez-Castelao, and Miguel Diaz-Hernandez

21. Analyzing the Role of the P2X7 Receptor in Epilepsy

Mariana Alves, Laura de Diego-Garcia, and Tobias Engel