Introduction To Histology And Microscopy PDF

BMS BSc Year 1: Fundamentals of Life Science Introduction to Histology and Microscopy Dr Genevieve Melling Join the Vevox session Go to vevox.app Enter the session ID: 183-136-906 Or scan the QR code  Learning Outcomes At the end of the se...

BMS BSc Year 1: Fundamentals of Life Science Introduction to Histology and Microscopy Dr Genevieve Melling Join the Vevox session Go to vevox.app Enter the session ID: 183-136-906 Or scan the QR code  Learning Outcomes At the end of the session, you should be able to:  List the organelles within a typical cell and consider their histological appearance.  Discuss the concept of the four fundamental categories of tissues and summarise the basic functions of each.  Discuss how tissue function is related to structure, particularly at a histological level.  Appreciate the relevance of histology to biological sciences, research and also to medicine.  Appreciate the techniques used in preparing specimens for microscopic examination.  Appreciate the magnification of light and electron microscopes and predict the tissues and cellular structures revealed.  Name some common histological stains and describe the tissues or cellular structures which they stain.  Recommended reading/ resources For most of your histology sessions: Wheater’s Functional Histology (Sixth edition) Another textbook for description? Junqueira’s (Find it @ bham) Clinical key For today’s session: Wheater’s Functional Histology (Sixth edition) Part 1 Chapter 1; Pages: 2 – 31, selected sections in Part 2 and Appendices 1 – 3, pages 428 – 434 Histology Teaching in Biomedical Science Year 1 In FLS: Workshop 1) Introduction to Histology and Microscopy Workshop 2) Histology of Epithelial tissues-FLIPPED! Workshop 3) Histology of Muscles- INTERACTIVE LECTURE Workshop 4) Histology of Connective Tissue-FLIPPED Workshop 5) Histology of Skin- FLIPPED Workshop 6) Histology of Cartilage and bone-INTERACTIVE LECTURE Histology workshops- Flipped sessions Flipped sessions Before the timetabled workshop: Access pre-session material on Labtutor (link on Canvas) Watch podcasts Attempt to answer questions on Lab tutor Ask any questions on a padlet (link on Canvas)/ OR think of questions you would like to ask during the workshop During the interactive workshop: Lab tutor questions will be discussed Knowledge from the podcasts will be applied Knowledge will be tested (polling) Your questions will be answered  Histology workshops- Flipped sessions Why use Flipped teaching? Hew et al., (2018) – performed meta analysis showing improved student learning, especially when quizzes were used Zhong et al., (2022)- found online flipped histology sessions were more enjoyable for students, increased interactivity and improved quiz scores Guo, et al. (2024)- found flipped sessions for histology led to better exam marks So Flipped sessions: Improve understanding Improve learning Improves engagement Improves interactivity Enable the teaching to student-centred Hew et al., BMC Med Educ 18, 38 (2018). Zhong, J., Li, Z., Hu, X. et al. BMC Med Educ 22, 795 (2022). Guo, et al. BMC Med Educ 24, 388 (2024). Intro: What is Histology? Microanatomy vs gross anatomy Form dictates function Important: for understanding for cells are organised into specific functional tissues> organs…. Intro: What is Histology? Microanatomy vs gross anatomy Form dictates function Important: for understanding for cells are organised into specific functional tissues> organs…. For diagnostics Researching human disease Researching drug treatments Researching new biomarkers Q) Any examples of diseases can be diagnosed by histology? How do we study histology? Created with BioRender.com  Four basic classifications of tissues Wheater’s Functional Histology Fig 8.3a (1), Fig 5.03b (2), Fig 10.01a (3), Fig 07.13 (4)  Four basic classifications of tissues Wheater’s Functional Histology Fig 8.3a (1), Fig 5.03b (2), Fig 10.01a (3), Fig 07.13 (4)  Four basic classifications of tissues Wheater’s Functional Histology Fig 8.3a (1), Fig 5.03b (2), Fig 10.01a (3), Fig 07.13 (4)  Four basic classifications of tissues Wheater’s Functional Histology Fig 8.3a (1), Fig 5.03b (2), Fig 10.01a (3), Fig 07.13 (4)  Four basic classifications of tissues 1) Muscle Epithelia 2) Connective tissue Nervous tissue 3) 4) Wheater’s Functional Histology Fig 8.3a (1), Fig 5.03b (2), Fig 10.01a (3), Fig 07.13 (4)  Histology in the body is complex Most structures in the body consist of several of the different types of tissue for example: Trachea (windpipe) https://www.histology.leeds.ac.uk/r H&E/Alcian blue (LP) espiratory/basic_structure.php Wheater’s Functional Histology  A typical mammalian cell structure and function Wheater’s Functional Histology Fig 1.1 and Fig 5.03b  Electron microscopy Transmission vs Scanning x16 500 x2000 Wheater’s Functional Histology Fig 1.1 and Fig 12.09  Which of the following was taken on a ##/## Join at: vevox.app ID: XXX-XXX-XXX Question slide Scanning Electron micrograph: FIG. 3.7 Erythrocytes. Wheater’s Functtional Histology  Which of the following was taken on a ##/## Join at: vevox.app ID: XXX-XXX-XXX Results slide Scanning Electron micrograph: RESULTS SLIDE FIG. 3.7 Erythrocytes. Wheater’s Functtional Histology  Microscopy https://ib.bioninja.com.au/standard-level/topic-1-cell-biology/12-ultrastructure-of- Electron microscopy- SEM vs TEM cells/electron-microscopy.html Wheater’s Functional Histology Fig 1.1 and Fig 5.03b  ##/## Join at: vevox.app ID: XXX-XXX-XXX Question slide What is this organelle? Mitochondrion ##.##% Smooth endoplastic reticulum ##.##% nucleus ##.##% rough endoplastic reticulum ##.##% Stevens & Lowe's Human Histology, Fifth Edition Fig 2. 15  ##/## Join at: vevox.app ID: XXX-XXX-XXX Results slide What is this organelle? Mitochondrion ##.##% Smooth endoplastic reticulum ##.##% nucleus ##.##% rough endoplastic reticulum ##.##% RESULTS SLIDE Stevens & Lowe's Human Histology, Fifth Edition Fig 2. 15 ##/## Join at: vevox.app ID: XXX-XXX-XXX Question slide What is the function of this organelle? ##/## Join at: vevox.app ID: XXX-XXX-XXX Results slide What is the function of this organelle? RESULTS SLIDE  Zooming in on organelles ? Free vs ER attached ribosomes Stevens & Lowe's Human Histology, Fifth Edition Fig 2.19 & 2.8  Zooming in on organelles ? What is this organelle? Stevens & Lowe's Human Histology, Fifth Edition Fig 2.21  ##/## Join at: vevox.app ID: XXX-XXX-XXX Question slide What is the function(s) of the golgi? protein translation ##.##% protein transcription ##.##% protein modification ##.##% protein sorting ##.##% protein transport ##.##% Stevens & Lowe's Human Histology, Fifth Edition Fig 2.21  ##/## Join at: vevox.app ID: XXX-XXX-XXX Results slide What is the function(s) of the golgi? protein translation ##.##% protein transcription ##.##% protein modification ##.##% protein sorting ##.##% protein transport ##.##% RESULTS SLIDE Stevens & Lowe's Human Histology, Fifth Edition Fig 2.21  Zooming in on organelles ? Euchromatin vs Heterochromatin Nucleus vs nucleolus Stevens & Lowe's Human Histology, Fifth Edition Fig 2.9  Zooming in on organelles ? Euchromatin vs Heterochromatin Nucleus vs nucleolus Euchromatin= light areas, lightly bound chromatin, actively transcribed DNA Heterochromatin= dense areas, transcriptionally inactive Stevens & Lowe's Human Histology, Fifth Edition Fig 2.9  How can we study cells? Many ways! One way is to culture cells In vitro What is the difference between primary and cell line?  How can we study cells? Many ways! One way is to culture cells In vitro What is the difference between primary and cell line? Dr Manu Carollo (Oxford Brookes University) growing many millions of cells from cervical cancer cell line; HeLa.  How can we study cells? Many ways! One way is to culture cells In vitro What is the difference between primary and cell line? Advantages and disadvantages? Dr Manu Carollo (Oxford Brookes University) Ethics? growing many millions of cells from cervical cancer cell line; HeLa. Picture from Amazon.co.uk How do we study histology? Fixation Dissection Embedding Sectioning Staining Visualisation Created with BioRender.com  Histology techniques- fixing Fixation Fixing tissue prevents degradation and helps Dissection maintain architecture Embedding Two methods: Chemical vs physical Sectioning Chemical= formalin based Staining Physical= ultra low freezing- Liquid Nitrogen Visualisation Different fixation for different microscopy e.g. electron microscopy-uses osmium tetroxide Fixation First, then dissection Wheater’s Pathology Fig 1.1  Histology techniques- embedding and sectioning Embedding the tissue in a wax or plastic resin. This enables us to cut very thin sections of tissue Using a microtome cryostat for frozen tissues Paraffin wax tissue block https://makeagif.com/gif/rotary-microtome-section-JBUPqN Wheater’s Pathology Fig 1.1 Histology techniques- staining Fixation Why do we stain? Dissection Embedding Sectioning Staining Visualisation  Histology techniques- staining Fixation Why do we stain? Dissection Embedding Sectioning Staining Can be specific stain or more general dye. Visualisation Chemistry of most dyes: acid/basic dyes Bind to different areas of the cell 1. Kamemoto LE et al 2010. Near-Infrared Micro-Raman Spectroscopy for in Vitro Detection of Cervical Cancer. Applied Spectroscopy. 64(3):255-261.  Histology techniques- staining Haematoxylin and Eosin Fixation Dissection Basic dyes (Haematoxylin) bind to _____ parts of the cells Embedding Sectioning Therefore: Acid dyes (eosin) bind to _____ Staining parts of the cells Visualisation Basophilic vs Acidophilic (eosinophilic)? Wheater’s Functional Histology Fig 5.03b  Stains.. Other stains: Elastic Van Gieson Masson’s trichrome  General stains Wheater’s Functional Histology Fig 5.03b Other stains… 4 5 6  General stains What stain has been used here? H&E Clue.. look at the connective tissue Elastic van Gieson Masson trichrome Wheater’s Functional Histology Fig 19.26 ##/## Join at: vevox.app ID: XXX-XXX-XXX Question slide What stain has been used here? H&E ##.##% Elastic van Gieson ##.##% Masson 's Trichrome ##.##% ##/## Join at: vevox.app ID: XXX-XXX-XXX Results slide What stain has been used here? H&E ##.##% Elastic van Gieson ##.##% Masson 's Trichrome ##.##% RESULTS SLIDE  General stains What stain has been used here? H&E Clue.. look at the connective tissue Elastic van Gieson Masson trichrome Human Histology Steven and Lowe Fig 18.14c ##/## Join at: vevox.app ID: XXX-XXX-XXX Question slide What stain has been used here? normal H & E ##.##% Elastic van Gieson ##.##% Mosson Trichrome ##.##% ##/## Join at: vevox.app ID: XXX-XXX-XXX Results slide What stain has been used here? normal H & E ##.##% Elastic van Gieson ##.##% Mosson Trichrome ##.##% RESULTS SLIDE  Specific stains Immunohistochemistry Immunocytochemistry Can be used to assess the expression of particular proteins, depending on antibody used. Figure 1.3 Wheater’s Pathology  Specific stains- immunohistochemistry cytokeratin= intermediate filament found in epithelial tissues Can be used to assess the expression of particular proteins, depending on antibody used. Figure 1.3 Wheater’s Pathology Figure 1.14 Wheater’s Functional Histology ##/## Join at: vevox.app ID: XXX-XXX-XXX Question slide What type of protein has been stained for here? nuclear protein ##.##% membrane receptor protein ##.##% cytoskeletal protein ##.##% mitochondrial protein ##.##% ##/## Join at: vevox.app ID: XXX-XXX-XXX Results slide What type of protein has been stained for here? nuclear protein ##.##% membrane receptor protein ##.##% cytoskeletal protein ##.##% mitochondrial protein ##.##% RESULTS SLIDE  Question Imagine: It is 2031, you have trained to become an NHS Biomedical scientist specialising in histopathology (obviously, because you love histology!). The surgical assistant has handed you a tissue biopsy from the tissue margins of their mastectomy to check whether they have fully removed all the cancerous tissue from a person’s breast, before the surgical team close up the skin. What kind of tissue preparation would you need to perform to ensure quick analysis of the tissue? https://www.urmc.rochester.edu/pathology-labs/blog/march- 2016/cut-and-dry-a-day-in-the-life-of-a-histologist.aspx  Group discussion Imagine: It is 2031, you have trained to become an NHS Biomedical scientist specialising in histopathology (obviously, because you love histology!). The surgical assistant has handed you a tissue biopsy from the tissue margins of their mastectomy to check whether they have fully removed all the cancerous tissue from a person’s breast, before the surgical team close up the skin. What kind of tissue preparation would you need to perform to ensure quick analysis of the tissue? 1) How would you fix the tissue? Chemical e.g. formalin, physical e.g. freezing, none 2) What kind of embedding would you choose to perform? Paraffin, plastic resin, none 3) What kind of stain would be most appropriate? Van Gieson, H&E, Masson’s trichrome, none 4) What kind of microscopy would you choose to perform? Light (bright field), fluorescence, electron microscopy https://www.urmc.rochester.edu/pathology-labs/blog/march- 2016/cut-and-dry-a-day-in-the-life-of-a-histologist.aspx