Abstract TP214: 3D-Electron Microscopy for the Analysis of the Microstructure of Acute Ischemic Stroke Clots

Abstract

Introduction: Characterization of acute ischemic stroke thrombi has typically focused on the two dimensional (2D) histological characterization of the thrombus. Little is known about the three dimensional (3D) structure of these occlusive thrombi. Electron Microscopy (EM) can be used to obtain 3D information about the organization of clot components. Individual structures can be segmented and qualitative and quantitative information obtained from the clots. Methods: Following IRB approval from Mayo Clinic, acute ischemic stroke clot samples were collected from 10 patients treated at Mayo Clinic. Samples were immediately fixed in fresh modified Trumps fixative solution. A representative fragment of clot was taken and split longitudinally into two pieces; one piece was sent for 3D-EM analysis and the other for histological processing and analysis. Hematoxylin and eosin stain (H&E) and Martius Scarlett Blue (MSB) histological stains were used for conventional histopathology evaluation and components analysis. Immunohistochemical (IHC) staining using anti-CD42b (Platelets) was applied to confirm the presence of platelets in each clot. Serial block-face scanning electron microscopy was used to generate high-resolution three-dimensional images. Results: The 2D morphological features of the clot were identified using H&E, MSB and anti-CD42b staining. The 3D organization of thrombi varies greatly, both within and between clots depending on the constituents. Red Blood Cell-rich areas are mainly composed of Red Blood Cells packaged tightly together, with a small number of Fibrin strands interwoven between the cells. Fibrin-rich areas are characterized by dense networks of fibrin strands along with platelets and White Blood Cells with some sparse Red Blood Cells present. Platelet-rich areas are composed of large volumes of de-granulated platelets, some fibrin stands and white blood cells and an absence of Red Blood Cells. Conclusions: 3D electron microscopy can provide important information on the organization of components within thrombi. This could provide key information on the age and etiology of the thrombus. In addition, it may help to inform thrombolytic treatment and medical device design.