Histology_Lecture1_introduction_to_Histology_and_Histological_techniques.pdf

University of Alkafeel, College of Medicine, Year 2, Course 1...

University of Alkafeel, College of Medicine, Year 2, Course 1 Histology - Lecture 1 Introduction to Histology and Histological techniques By: Assistant Professor Dr. Rasha Dosh 1 alkafeel.edu.iq [email protected] 18/9/2024  Learning objectives At the end of the lecture you should be able to: 1- Define human histology. 2- Know the organization and components of the human body. 3- Know the histological techniques for preparation and staining of tissue sections and different types of microscopes. 2 alkafeel.edu.iq [email protected] 18/9/2024  What is Histology? Histology: is the study of tissues and organs under the microscope. It is the microscopic anatomy, to study the detailed structure with the aid of light microscope or electron microscope (EM) (Transmission Electron Microscope (TEM) and Scanning Electron Microscope (SEM). histos is Greek for tissue, logia is Greek for science Cytology: is the study of the structure and function of the cell. 3 alkafeel.edu.iq [email protected] 18/9/2024  Why Study Cells and Tissues? Levels of Organizations: Cells are the smallest living units of the levels of organization. Tissues are many cells + extracellular ground substance  Four basic tissues:  Epithelial tissue  Connective tissue  Muscular tissue  Nervous tissue Organs: made up of tissue, have special shape, structure and function System: organs which have related function get together. 4 alkafeel.edu.iq [email protected] 18/9/2024  Tissue Processing Haematoxylin and Eosin (H&E)-stained sections of formalin-fixed tissue are the specimens most commonly examined for histologic studies with the light microscope. Steps of Tissue Processing: 1. Receipt & Identification 2. Labelling of the specimen with numbering 3. Fixation 4. Dehydration 5. Clearing 6. Infiltration 7. Embedding 8. Sectioning 9. Staining 10. Mounting 5 alkafeel.edu.iq [email protected] 18/9/2024  6 alkafeel.edu.iq [email protected] 18/9/2024  Automated tissue processor Consist of 12 stations 1 jar – formalin 7 jars – grades of alcohol 2 jars – xylene 2 jars – molten paraffin wax 7 alkafeel.edu.iq [email protected] 18/9/2024  Fixation Any tissue once taken out of the body will decompose due to:- 1- Loss of bloody supply and oxygen 2- Accumulation of products of metabolism 3- Action of autolytic enzymes 4- Putrefaction by bacteria 5- All the above changes PREVENTED BY FIXATION! Fixation: which helps in the following: 1- Terminate cell metabolism. 2- Prevent enzymatic degradation of cells and tissues by autolysis (self-digestion). 3- Kill pathogenic microorganisms such as bacteria, fungi, and viruses. 4- Harden the tissue as a result of either cross-linking or denaturing protein molecules. he most common fixative is Formalin, a 37% aqueous solution of formaldehyde. 8 alkafeel.edu.iq [email protected] 18/9/2024  Dehydration It is the process in which the water content in the tissue to be processed is completely reduced by passing the tissue through increasing concentrations of dehydrating agents (alcohol). Water removed from tissues and cells – this space is occupied by wax. The duration of the procedure can be noted down as 1. 50 % alcohol – 1 hour 2. 60 % alcohol – 1 hour 3. 70 % alcohol – 1 hour 4. 80 % alcohol – 1 hour 5. 90 % alcohol – 1 hour 6. Absolute alcohol – 1 hour 7. Absolute alcohol – 1 hour Dehydration is done so that the wax i.e Paraffin wax, which is used for infiltration, can be easily mixed as it is unmixed with water. 9 alkafeel.edu.iq [email protected] 18/9/2024  Clearing ( dealcoholization) 1- This is the step between dehydration and paraffin infiltration. 2- The purpose of this step is removal of the alcohol and replacement with a fluid that will mix with paraffin. 3- This step is called clearing because they penetrate, make the tissue transparent and also harden it. 4- Xylene is the most commonly used clearing agent. 10 alkafeel.edu.iq [email protected] 18/9/2024  Infiltration and embedding Empty spaces in tissues and cells , after removal of clearing agent, are taken by molten paraffin wax for 6-8 hours. Melting point of wax (52- 60 ºC) Hardens the tissue and helps in easy sectioning of the tissue. Embedding is done by transferring the tissue which has been cleared of the alcohol to a mould filled with molten wax & is allowed to cool & solidify. After solidification, a wax block is obtained which is then sectioned to obtain ribbons. 11 alkafeel.edu.iq [email protected] 18/9/2024  Sectioning It is the procedure in which the blocks which have been prepared are cut or sectioned and thin strips of varying thickness are prepared. The instrument by which this is done is called a Microtome. 12 alkafeel.edu.iq [email protected] 18/9/2024  Routine staining (H&E) 1- Staining of the section is done to bring out the particular details in the tissue under study. 2- The most commonly used stain in routine practice is Haematoxylin & Eosin stain. Haematoxylin – nuclear stain (blue) Eosin – cytoplasmic stain (pink) 3- Haematoxylin can be considered as a basic dye. It is used to stain acidic (or basophilic) structures a purplish blue, thus the nucleus is stained purple. 4- Eosin is an acidic dye: It stains basic (or acidophilic) structures red Brush Border or pink. This is also sometimes termed eosinophilic, thus the cytoplasm is stained pink. 13 alkafeel.edu.iq [email protected] 18/9/2024  14 alkafeel.edu.iq [email protected] 18/9/2024  Types of Microscopes light microscope TEM SEM 15 alkafeel.edu.iq [email protected] 18/9/2024  References 1- Junqueira’s Basic Histology: Text and Atlas, 16th Edition, by Anthony L. Mescher. 2- A Text and Atlas: With Correlated Cell and Molecular Biology, 8th edition, by Wojciech Pawlina, Michael H. Ross. 16 alkafeel.edu.iq [email protected] 18/9/2024  ANY QUESTIONS? 17 alkafeel.edu.iq [email protected] 18/9/2024  Thank you for Listing 18 alkafeel.edu.iq [email protected] 18/9/2024