Practical Forensic Microscopy

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1 Practical Forensic Microscopy A Laboratory Manual Barbara P. Wheeler and Lori J. Wilson Department of Chemistry, Eastern Kentucky University Richmond, KY, USA

3 Practical Forensic Microscopy A Laboratory Manual

5 Practical Forensic Microscopy A Laboratory Manual Barbara P. Wheeler and Lori J. Wilson Department of Chemistry, Eastern Kentucky University Richmond, KY, USA

6 Copyright 2008 John Wiley & Sons Ltd, The Atrium, Southern Gate, Chichester, West Sussex PO19 8SQ, England Telephone (+44) (for orders and customer service enquiries): Visit our Home Page on or All Rights Reserved. No part of this publication may be reproduced, stored in a retrieval system or transmitted in any form or by any means, electronic, mechanical, photocopying, recording, scanning or otherwise, except under the terms of the Copyright, Designs and Patents Act 1988 or under the terms of a licence issued by the Copyright Licensing Agency Ltd, 90 Tottenham Court Road, London W1T 4LP, UK, without the permission in writing of the Publisher. Requests to the Publisher should be addressed to the Permissions Department, John Wiley & Sons Ltd, The Atrium, Southern Gate, Chichester, West Sussex PO19 8SQ, England, or ed to permreq@wiley.co.uk, or faxed to (+44) Designations used by companies to distinguish their products are often claimed as trademarks. All brand names and product names used in this book are trade names, service marks, trademarks or registered trademarks of their respective owners. The Publisher is not associated with any product or vendor mentioned in this book. This publication is designed to provide accurate and authoritative information in regard to the subject matter covered. It is sold on the understanding that the Publisher is not engaged in rendering professional services. If professional advice or other expert assistance is required, the services of a competent professional should be sought. Other Wiley Editorial Offices John Wiley & Sons Inc., 111 River Street, Hoboken, NJ 07030, USA Jossey-Bass, 989 Market Street, San Francisco, CA , USA Wiley-VCH Verlag GmbH, Boschstr. 12, D Weinheim, Germany John Wiley & Sons Australia Ltd, 42 McDougall Street, Milton, Queensland 4064, Australia John Wiley & Sons (Asia) Pte Ltd, 2 Clementi Loop #02-01, Jin Xing Distripark, Singapore John Wiley & Sons Canada Ltd, 6045 Freemont Blvd, Mississauga, Ontario, L5R 4J3, Canada Wiley also publishes its books in a variety of electronic formats. Some content that appears in print may not be available in electronic books. Library of Congress Cataloging-in-Publication Data Wheeler, Barbara P. Practical forensic microscopy : a laboratory manual / by Barbara P. Wheeler and Lori J. Wilson. p. cm. Includes index. ISBN Medical jurisprudence Laboratory manuals. 2. Microscopy Laboratory manuals. I. Wilson, Lori. II. Title. RA1034.W dc British Library Cataloguing in Publication Data A catalogue record for this book is available from the British Library ISBN: Typeset in 10/12 Times by Laserwords Private Limited, Chennai, India Printed and bound in Singapore by Markono Ltd This book is printed on acid-free paper.

7 Contents Preface Acknowledgements Laboratory Safety Microscope Maintenance The Micro Kit ix xi xiii xv xvii Experiments 1 Chapter 1 The Stereomicroscope 3 Experiment 1A: Familiarization with the Stereomicroscope 4 Chapter 2 The Compound Light Microscope 13 Experiment 2A: Familiarization with the Compound Light Microscope 14 Experiment 2B: Measurements Using the Ocular Micrometer 20 Experiment 2C: Microscopic Mounting Techniques 25 Experiment 2D: Determining Refractive Index 29 Chapter 3 The Polarized Light Microscope 39 Experiment 3A: Familiarization with the Polarized Light Microscope 41 Experiment 3B: Determining Refractive Index of Anisotropic Materials 48 Experiment 3C: Determining Birefringence and Sign of Elongation 53 Chapter 4 The Fluorescence Microscope 59 Experiment 4A: Familiarization with the Fluorescence Microscope 61 Chapter 5 The Phase Contrast Microscope 67 Experiment 5A: Familiarization with the Phase Contrast Microscope 69 Application Experiments 75 Chapter 6 Experiment 6: Physical Match Examinations 77 Chapter 7 Experiment 7: Construction Examinations of Evidence 83 Chapter 8 Experiment 8: Lamp Filament Examinations 93 Chapter 9 Experiment 9: Fingerprint Examinations and Comparison 103 Chapter 10 Experiment 10: Tool Mark Examinations 111

8 vi CONTENTS Chapter 11 Firearms Examinations 115 Experiment 11: Firearms Examinations 115 Experiment 11A: Gunshot Residue Examinations 122 Chapter 12 Experiment 12: Shoe and Tire Print/Impression Examinations 131 Chapter 13 Experiment 13: Botanical Examinations 137 Chapter 14 Experiment 14: Paint Examinations 143 Chapter 15 Hair Examinations 149 Experiment 15: Hair Examinations 149 Experiment 15A: Animal Hair Examinations 160 Experiment 15B: Determination of Racial and Somatic Origin Characteristics of Human Hair 169 Experiment 15C: Human Hair Examinations and Comparisons 178 Experiment 15D: Evaluation of Human Hair for DNA 186 Chapter 16 Glass Examinations 193 Experiment 16: Glass Examinations 193 Experiment 16A: Glass Breakage Determinations 201 Chapter 17 Fiber Examinations 209 Experiment 17: Textile Fibers Examinations 209 Experiment 17A: Natural Fiber Examinations 215 Experiment 17B: Man-made Fiber Examinations 227 Experiment 17C: Fiber Comparisons 237 Chapter 18 Soil Examination 243 Experiment 18: Soil Examinations 243 Experiment 18A: Identification of Minerals in Soil 251 Chapter 19 Experiment 19: Microchemical Testing Inorganic Ions 265 Chapter 20 Experiment 20: Microscopic Analysis of Controlled Substances 277 Chapter 21 Experiment 21: Semen Examinations 289 Instrumental Microscopy 295 Chapter 22 Experiment 22: Fourier Transform Infrared Microspectrometry 297 Chapter 23 Experiment 23: UV-Visible-NIR Microspectrophotometry 307 Chapter 24 Experiment 24: Thermal Microscopy 315 Chapter 25 Experiment 25: Scanning Electron Microscopy 323 Appendices 331 A Optical Properties of Natural Fibers 331 B Optical Properties of Man-Made Fibers 333

9 CONTENTS vii C Michel-Lévy Chart 337 D Dispersion-Staining Graph 339 E Mock Case Scenario 341 F Circle Template 345 Glossary of Microscopy Terms 347 Index 357

11 Preface Forensic science is a discipline that has evolved from the application of science to questions arising from crime or litigation. Since the popularity of forensic science as a career choice has emerged, many colleges and universities have developed criminalistics and forensic science programs. Swelling enrollments have created a market for texts within this field. Because of this heightened interest, there are many texts that concentrate on the general aspects of the field, providing an introduction to forensic science. Due to rising public interest, there are even a few texts that bring attention to separate disciplines within forensic science, for instance, firearms, drugs, and DNA. However, in the past, forensic microscopy has had little distinction, being overshadowed by more visible disciplines. In recent years, some highly publicized criminal cases have brought forensic microscopy into the spotlight. Practical Forensic Microscopy is a comprehensive lab manual that adapts microscopic procedures used in the forensic laboratory to practical experiments that can be taught in college laboratories. The manual is written by a practitioner and an academician, and so a balanced approach to the topic was able to be reached. This laboratory manual provides a general overview and understanding of the numerous microscopes and microscopic techniques used within the field of forensic science. Each topic covered begins with a list of simple objectives for the experiment. To assist the student in obtaining the objective, an explanation of the topic, selected reading references, and an experiment are used. Worksheets and drawing templates have also been included to compile analytical results. Instructors may find it useful to download the worksheets and templates from To test the student s knowledge, report requirements and questions are included. This manual is unique among other laboratory manuals in the fact that the microscopic techniques commonly used by scientists have been applied to forensic disciplines. However, in some cases it is impractical to use forensic laboratory procedures in an educational setting due to the large number of students or when equipment and supplies such as controlled substances are not available. Every attempt has been made to adapt forensic laboratory procedures to best address these concerns. When significant modifications are made for the educational setting, the scientifically accepted theory and principles of the forensic lab procedures are still covered thoroughly in the introduction. To address these concerns, it was at times necessary to make concessions for accuracy or precision. For example, the absolute measurement of density of glass evidence in a forensic laboratory would use a temperature controlled system, whereas in our procedure students use standard laboratory glassware and balances. However, we felt that the need to cover the topic of measurement of the density of glass far outweighed the concessions made in accuracy and precision. This manual is an attempt to bring forensic microscopy to the student so that the future of this discipline within forensic science will continue to flourish. Forty laboratory experiments have been developed to cover the variety of evidence disciplines within the forensic science field. The manual starts with the use of simple stereomicroscopes and gradually introduces more complex microscope systems used in a forensic laboratory. Each forensic science discipline, which uses microscopes, is

12 x PREFACE covered so that the student will obtain a general understanding of the microscopes and microscopic techniques used in examinations. For example, impression evidence such as fingerprints, shoe print patterns, tool marks, and firearms are analyzed using simple stereomicroscopic techniques. Biological, drug, and trace evidence (i.e., paint, glass, hair fiber) are covered by a variety of microscopes and specialized microscopic techniques. The authors have successfully used a mock case scenario at the end of each semester where students are placed into groups of three and provided with items of evidence. This has been an effective way to solidify the topics covered throughout the semester and in some cases extend the procedures covered. The group divides the tasks and can ask for additional exhibits if available. The group fills out a laboratory report form (Appendix E) and provides individual notes as their lab report. The ability to reach a conclusion and communicate the necessary information in a concise manner is one of the goals of the mock case scenario. Obtaining the knowledge and developing the skills will allow students to bring forensic microscopy once again to the forefront.

13 Acknowledgements When we agreed to write this laboratory manual, we couldn t begin to comprehend the time and effort that would be required to complete the manuscript. Thanks to the work and encouragement of many individuals, our thoughts and ideas have been realized. To begin, we wish to thank Dr Vernon Stubblefield who instilled in us the beginnings of forensic microscopy. Without his initial insight and commitment to the fundamentals of polarized light microscopy, we wouldn t be the microscopists we are today. Many others have also contributed a significant amount of effort toward the manual. We are most grateful to Melanie Bentley of the College of Arts and Sciences at EKU who spent an extraordinary amount of time and effort assisting with sketches, photographs, and the final layouts of our figures. Because of her skills, many of the experiments are enhanced by visual aids. Her perfectionist quality and willingness to assist in this project greatly improved the lab manual. We also appreciate the work of Forensic Science majors Jesse Meiers, for his assistance with photographs, and Ethan Harlacher, for his assistance with references and glossary items. We are grateful to Marci Adkins, Lara Mosenthin, and Patrick McLaughlin of the Kentucky State Police Central Forensic Laboratory for providing technical assistance in each of their areas of the forensic science discipline. We also appreciate the laboratory experiments contributed by Dr Larry Kaplan, Williams College, Dr Larry Quarino, Cedar Crest College, and Mr Joe Wallace, Kentucky Department of Criminal Justice Training/EKU Forensic Science Program. We feel it is also important to acknowledge the support of Dr Diane Vance and the other faculty members of the Department of Chemistry at Eastern Kentucky University for their support and encouragement during the project. The support of the Department of Chemistry and Eastern Kentucky University also helped make this project possible. We wish to acknowledge the government agencies, instrument manufacturers, and private companies cited in the manual for contributing their photographs and illustrations. Finally we wish to thank Fiona Woods and the staff at John Wiley & Sons, Ltd. And most of all, special thanks is given to our families for their endless patience, encouragement, and support during the project. We promise not to burn any more dinners until our next project.

14 xii ACKNOWLEDGEMENTS Let s take a closer view at the amazing field of forensic microscopy. (Photograph by Chris Radcliffe, reproduced with permission of Eastern Kentucky University)

15 Laboratory Safety Laboratory work can be very interesting and exciting, however certain safety concerns should always be taken into consideration. General laboratory safety rules follow. Each laboratory will have its own set of rules, so make sure that you have read those for your specific laboratory, understand them, and comply with them. When there is a question concerning laboratory safety, please ask the instructor. 1. Many materials in a laboratory may cause eye injury. Wear approved safety glasses to protect against chemical splashes and stray impacts. 2. Wear a protective laboratory apron or coat and close-toed shoes. 3. No eating, drinking, smoking, or applying makeup in the lab. 4. Keep your work area clean and free of clutter. 5. Be prepared to work while you are in the laboratory. No horseplay is permitted in the lab. 6. Do not perform any unauthorized experiments. 7. Handle scalpels and razor blades with extreme caution. Never cut materials toward you. 8. Use a fume hood for all substances that produce strong odors or fumes. 9. Do not remove any materials from the laboratory. 10. Dispose of all waste properly. Ask your instructor for directions if you are not sure what to do. To avoid contamination, never return chemicals to their original containers. 11. Do not work alone in the laboratory. 12. Check equipment to be sure that it is in good condition. Don t use chipped or cracked glassware. 13. Never pipette by mouth. Always use suction bulbs or disposable pipettes. 14. Never touch, taste, or smell a chemical. If you are instructed to note the smell, gently wave your hand over the opening to direct fumes toward your nose. Do not inhale the fumes directly from the container. 15. Rinse off any acid or base spills on your skin/clothing. Clean up all spills immediately and notify your instructor. Ask your instructor for assistance if you are not sure what to do. 16. Read labels carefully to be sure that you are using the correct reagent for an experiment. 17. Know the location and operation of the eyewash, safety shower, spill materials, and fire extinguisher in the lab. 18. Know the location of the safety kit. 19. All accidents and injuries should be reported to the instructor immediately. 20. Follow other safety rules as set by your laboratory.

Practical Forensic Microscopy

Practical Forensic Microscopy A Laboratory Manual Barbara P. Wheeler and Lori J. Wilson Department of Chemistry, Eastern Kentucky University Richmond, KY, USA Practical Forensic Microscopy A Laboratory