Basic and Advanced Laboratory Techniques in Histopathology and Cytology Pranab Dey

Basic and Advanced Laboratory Techniques in Histopathology and Cytology Pranab Dey Basic and Advanced Laboratory Techniques in Histopathology and Cytology Pranab Dey Department of Cytology and Gynecologic Pathology Post Graduate Institute of Medical Education and Research Chandigarh India ISBN 978-981-10-8251-1 ISBN 978-981-10-8252-8 (eBook) https://doi.org/10.1007/978-981-10-8252-8 Library of Congress Control Number: 2018941817 © Springer Nature Singapore Pte Ltd. 2018 This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. The publisher, the authors and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, express or implied, with respect to the material contained herein or for any errors or omissions that may have been made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Printed on acid-free paper This Springer imprint is published by the registered company Springer Nature Singapore Pte Ltd. part of Springer Nature. The registered company address is: 152 Beach Road, #21-01/04 Gateway East, Singapore 189721, Singapore Dedicated to Shree Shree Satyananda Giri, Rini and Madhumanti Preface Laboratory techniques in histopathology and cytology are the foundation of the diagnostic pathology. It is extremely essential to know all the basic and advanced techniques in laboratory. This book discusses the principles, steps, and troubleshooting areas of all the essential laboratory techniques in both histology and cytology laboratories. It contains multiple illustrations, microphotographs, tables, and boxes that explain the techniques. In addition to the various advanced techniques, microscopy and quality control in the laboratory have been discussed. I hope that the book will help all the postgraduate students in pathology, practising pathologists, and laboratory technologists. Chandigarh, India Pranab Dey 2017 September vii Acknowledgements I wish to express my thanks to Dr. Naren Aggarwal and Ms. Jagjeet Kaur Saini of Springer Nature who encouraged me in every stage in this work. I am thankful to Dr. Suvradeep Mitra who gave me his valuable sugges- tions at the time of the preparation of this manuscript. He also provided me many microphotographs. My sincere thanks to Dr. Charan Singh Rayat, Lecturer in Histopathology, who shared his vast experience in laboratory technology with me. I am grateful to my technical staffs who exchanged their views and opinions with me. My wife Rini and daughter Madhumanti constantly encouraged me during the writing of this book. They are my source of inspiration. Lastly I wish to express my gratitude to God because without His blessing nothing can be done. ix Contents Part I Basic Laboratory Techniques in Histopathology Laboratory 1 Fixation of Histology Samples: Principles, Methods and Types of Fixatives �� 3 1.1 Introduction �� 3 1.2 Aims of Fixation �� 3 1.3 Ideal Fixative �� 3 1.4 Tissue Changes in Fixation �� 3 1.5 Types of Fixation �� 4 1.5.1 Description of Nature of Fixation �� 5 1.6 Essential Precautions for Fixation in General �� 6 1.7 Mechanism of Fixation �� 6 1.8 Factors Affecting Fixation �� 8 1.9 Commonly Used Fixatives in the Laboratory �� 10 1.9.1 Formaldehyde �� 10 1.9.2 Preparation of Different Formalin Solution �� 10 1.9.3 Glutaraldehyde �� 11 1.9.4 Osmium Tetroxide �� 12 1.9.5 Methyl and Ethyl Alcohol �� 12 1.9.6 Acetone �� 12 1.9.7 Bouin’s Fixative �� 12 1.10 Mercury Salt-Containing Fixatives �� 13 1.10.1 Zenker’s Fluid �� 13 1.10.2 Helly’s Fluid �� 13 1.10.3 B5 Fixatives �� 13 1.10.4 Fixatives of Choice �� 13 1.11 Fixation Artefact �� 15 References �� 17 2 Processing of Tissue in Histopathology Laboratory �� 19 2.1 Factors that Influence Tissue Processing �� 19 2.2 Dehydration �� 20 2.3 Individual Dehydrating Agent �� 21 2.3.1 Alcohol �� 21 2.3.2 Dehydrating Agents Other than Alcohol �� 21 2.4 Clearing �� 22 2.4.1 Individual Clearing Agent �� 22 2.4.2 Other Clear Agents �� 23 xi xii Contents 2.5 Infiltration and Embedding �� 24 2.5.1 Different Impregnating Medium �� 24 2.6 Tissue Processing Methods �� 25 2.7 Overall Precautions of Tissue Processing �� 26 2.7.1 Time Schedule for Overnight Processing �� 26 2.7.2 Manual Tissue Processor �� 26 2.7.3 Microwave Processing �� 27 References �� 27 3 Embedding of Tissue in Histopathology �� 29 3.1 Embedding Medium �� 29 3.2 Different Types of Mould Used for Block �� 30 3.3 Tissue Embedding Method �� 30 3.4 Tissue Orientation and Embedding �� 32 3.5 Tissue Marking �� 32 References �� 33 4 Decalcification of Bony and Hard Tissue for Histopathology Processing �� 35 4.1 Introduction �� 35 4.1.1 Factors Controlling the Rate of Decalcification �� 36 4.2 The Methods of Decalcification �� 36 4.3 Chelating Agents �� 37 4.3.1 Other Procedures of Decalcification �� 38 4.4 Surface Decalcification �� 38 4.5 End Point Determination of Decalcification �� 39 Reference �� 39 5 Tissue Microtomy: Principle and Procedure �� 41 5.1 Introduction �� 41 5.2 Microtomes �� 41 5.2.1 Microtome Knife �� 43 5.2.2 Disposable Knife �� 44 5.2.3 Materials Used in Knife �� 44 5.2.4 Angles of Knife �� 44 5.3 Microtome Knife Sharpening �� 44 5.3.1 Manual Method �� 44 5.3.2 Factors Involved in Cutting ��

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