Blood platelets play a central role in the arrest of bleeding and the development of thrombosis. Unraveling the complex processes of platelet biogenesis from megakaryocytes, platelet adhesion, aggregation, and secretory responses are important topics in the field of hemostasis and thrombosis. Analysis of the ultrastructural changes that occur during these processes is essential for understanding the rapid membrane dynamics and has contributed substantially to our present knowledge of platelet formation and functioning. Recent developments in real-time imaging, correlative light and electron microscopy imaging (CLEM), and 3D (cryo) electron microscopy and tomography offer exciting opportunities to improve studies of the platelet adhesive responses and secretion at the ultrastructural level in a close to native environment. In this chapter we discuss and illustrate cryo preparation techniques (high-pressure freezing, vitrification), correlative LM and EM workflows, and 3D cryo-electron tomography that we apply in our current research projects.
Keywords: Correlative light and electron microscopy (CLEM); Cryo-EM; Fixation and cryo-immobilization; High pressure freezing EM tomography; Immuno-EM; TEM.