12 The average human body has on the order of 10 circulating platelets. They are crucial for hemostasis, and yet excessive platelet activation is a major cause of m- bidity and mortality in Western societies. It is therefore not surprising that platelets have become one of the most extensively investigated biological cell types. We are, however, far from understanding precisely how platelets become activated under physiological and pathophysiological conditions. In addition, there are large gaps in our knowledge of platelet production from their giant precursor cell, the megakar- cyte. Understanding megakaryocyte biology will be crucial for the development of platelet gene targeting. The aim of Platelets and Megakaryocytes is therefore to bring together established and recently developed techniques to provide a comprehensive guide to the study of both the platelet and the megakaryocyte. It consists of five s- tions split between two volumes. The more functional assays appear in Volume 1, whereas Volume 2 includes signaling techniques, postgenomic methods, and a n- ber of key perspectives chapters. Part I of Volume 1, Platelets and Megakaryocytes: Functional Assays, describes many well-established approaches to the study of platelet function, including aggregometry, secretion, arachidonic acid metabolism, procoagulant responses, pla- let adhesion under static or flow conditions, flow cytometry, and production of microparticles. Although one would ideally wish to perform experiments with human platelets, studies within the circulation using intravital microscopy require the use of animal models, which are described in Chapter 16, vol. 1.

Part I. Perspectives Signaling Receptors on Platelets and Megakaryocytes Donna Woulfe, Jing Yang, Nicolas Prevost, Peter O''Brien, Ryan Fortna, Massimiliano Tognolini, Hong Jiang, Jie Wu, and Lawrence F. Brass Manipulation of Gene Expression in Megakaryocytes Nikla R. Emambokus, George J. Murphy, and Jonathan Frampton In Vitro Changes of Platelet Parameters: Lessons From Blood Banking Jose Rivera, María L. Lozano, and Vicente Vicente Pharmacological Approaches to Studying Platelet Function: An Overview Susanna M. O. Hourani Inhibitors of Cellular Signaling Targets: Designs and Limitations Chris J. Vlahos and Louis F. Stancato Part II Signaling Techniques. Ligand-Binding Assays for Collagen Stephanie M. Jung and Masaaki Moroi Use of Radiolabeled 2-Methylthio-ADP to Study P2Y Receptors on Platelets and Cell Lines Pierre Savi and Jean-Marc Herbert Ligand-Binding Assays: Fibrinogen Dermot Cox Techniques for the Analysis of Proteins by SDS-Polyacrylamide Gel Electrophoresis and Western Blotting Jonathan M. Gibbins Study of Tyrosine Kinases and Protein Tyrosine Phosphorylation Jonathan M. Gibbins Protein Tyrosine Phosphatases Matthew L. Jones and Alastair W. Poole The Study of Serine-Threonine Kinases Gertie Gorter and Jan Willem Akkerman Phosphoinositides: Lipid Kinases and Phosphatases Bernard Payrastre Isolation and Analysis of Platelet Lipid Rafts Corie N. Shrimpton, Karine Gousset, Fern Tablin, and Jose A. Lopez Measurement and Manipulation of [Ca2+]i in Suspensions of Platelets and Cell Cultures Philippe Ohlmann, Beatrice Hechler, Jean-Pierre Cazenave, and Christian Gachet Measurement and Manipulation of Intracellular Ca2+ in Single Platelets and Megakaryocytes Michael J. Mason and Martyn P. Mahaut-Smith Patch-Clamp Recordings of Electrophysiological Events in the Platelet and Megakaryocyte Martyn P. Mahaut-Smith Evaluation of Nitrotyrosine-Containing Proteins in Blood Platelets Khalid M. Naseem, Rocio Riba, and Max Troxler Nitric Oxide Signaling in Platelets Sylvia Y. Low and K. Richard Bruckdorfer Snake Venom Toxins Affecting Platelet Function Robert K. Andrews, Elizabeth E. Gardiner, and Michael C. Berndt Peptide Synthesis in the Study of Collagen-Platelet Interactions C. Graham Knight, Catherine M. Onley, and Richard W. Farndale Platelet Permeabilization Robert Flaumenhaft Part III. Molecular and Post-Genomic Techniques Using Retroviruses to Express Genes in Primary Megakaryocyte Lineage Cells Meenakshi Gaur, George J. Murphy, Jonathan Frampton, and Andrew D. Leavitt Use of Antisense Oligonucleotide Technology to Investigate Signaling Pathways in Megakaryocytes Hava Avraham, Shalom Avraham, and Radoslaw Zagozdzon GFP Fusion Proteins to Study Signaling in Live Cells Simon A. Walker, Gyles E. Cozier, and Peter J. Cullen Two-Dimensional Polyacrylamide Gel Electrophoresis for Platelet Proteomics Katrin Marcus and Helmut E. Meyer Preparation of mRNA and cDNA Libraries From Platelets and Megakaryocytes Benjamin Z. S. Paul, Jiango Jin, and Satya P. Kunapuli Platelet Receptor Structures and Polymorphisms Thomas J. Kunicki, Steven Head, and Daniel R. Salomon Gene Array Technology and the Study of Platelets and Megakaryocytes Lloyd T. Lam and Emery H. Bresnick Index