Cytology and Histology L1 Introduction PDF

Introduction Prepared by Dr. Hagar Ali Abdul Raheem Abu Quora Lecturer of Cytology, Histology, and Histochemistry – Zoology Department – Faculty of Science – Menoufia University - Egypt...

Introduction Prepared by Dr. Hagar Ali Abdul Raheem Abu Quora Lecturer of Cytology, Histology, and Histochemistry – Zoology Department – Faculty of Science – Menoufia University - Egypt Cytology and Histology – First semester 2023/2024  Cytology and Histology Cytology is the science that deals with the study of cell structure and function. (Cyto. Means cell, and logia, means branch of science). Histology is the science that deals with the study of normal tissues of the body and how these tissues are arranged to constitute organs. (Gr. Histo means tissue, and Logia, means branch of science). 18/10/2023 Introduction Cytology & Histology L1 2 Introduction The living organisms are made up of many tiny living compartments called cells. During development, cells and extracellular matrix are associated together to give raise different types of tissues with characteristic structural features. Organs are formed by an arranged combination of these tissues and their precise arrangement allows the functioning of each organ and of the organism as a whole (Fig. 1). 18/10/2023 Introduction Cytology & Histology L1 3  Fig. 1 Relationship between cells to form a whole living organism. 18/10/2023 Introduction Cytology & Histology L1 4 Preparation of Electron microscope samples  A very small piece of tissue is obtained from the living organism (diagnostic or surgical biopsy) or after complete dissection (samples) are immediately placed in fixatives like glutaraldehyde followed by osmium. The aim of fixatives is to prevent post-mortem degeneration, preserve tissues as it is in live and protect the misleading changes in cells and tissues. After dehydration and clearing , embedding by plastic Araldite. Ultra sections (1/40µm) by ultramicrotome used diamond or glass knives. Sections are mounted on copper grids and stained with uranyl acetate and lead citrate. 18/10/2023 Introduction Cytology & Histology L1 5 Transmission and Scanning Electron Microscope Fig. 2 TEM and SEM 18/10/2023 Introduction Cytology & Histology L1 6  Fig. 3 TEM and SEM pictures. 18/10/2023 Introduction Cytology & Histology L1 7  Albino Rat Albino Mouse Dissected Rat Kidney Section of kidney H&E 40x Fig. 4 From animal models (rat & mouse) to tissue sections. 18/10/2023 Introduction Cytology & Histology L1 8 Preparation of microscopically sections by paraffin sections Fixation: A small piece of tissue is obtained from the living organism (diagnostic or surgical biopsy) or after complete dissection (samples) are immediately placed in fixatives. There are many types of fixatives according to the type of tissue and stain like formaldehyde, acetone, picric acid, bouin, and so on …. The aim of fixatives is to prevent post-mortem degeneration, preserve tissues as it is in live and protect the misleading changes in cells and tissues. Post fixation treatment by running tap water overnight. 18/10/2023 Introduction Cytology & Histology L1 9 Preparation of microscopically sections by paraffin sections Dehydration:  gradually remove the water by ascending series of alcohol to prevent the shrinkage of tissues (70%, 80%, 90%, 100%I, and 100%II). 18/10/2023 Introduction Cytology & Histology L1 10 Preparation of microscopically sections by paraffin sections Clearing: Tissues are placed in paraffin solvent (xylene or xylol) to completely replaced alcohol by xylene in preparing to paraffin embedding. Infiltration: The tissues are then placed in melted paraffin in an oven (52-60oC) until completely infiltrated by this substance. Embedding: The paraffin infiltrated tissue is placed in a small hardened block to facilitate sectioning (Fig. 3). 18/10/2023 Introduction Cytology & Histology L1 11  Fig. 5 steps from fixation to embedding. 18/10/2023 Introduction Cytology & Histology L1 12 Mc Graw-Hill Companies Preparation of microscopically sections by paraffin sections Trimming: Tissue blocks are trim to be suitable for sectioning by microtome. Sectioning: The paraffin blocks are fixed on the holder of the microtome and sections are cut. The thickness of sections is ranged from 5-7µm (Fig. 4). Fig. 6 Microtome instrument. 18/10/2023 Introduction Cytology & Histology L1 13 Mc Graw-Hill Companies 18/10/2023 Introduction Cytology & Histology L1 Video 1 BioGenex 1996 14 Preparation of microscopically sections by paraffin sections Affixation: Paraffin sections are attached to the glass slides by sticky material. Staining (H&E) and mounting: Cell components such as DNA with a negative charge are stained by basophilic stain (hematoxylin), while other cytoplasmic components are stained by acidophilic stain (eosin). After that, the stained sections mount with Canada balsam or D.PX. 18/10/2023 Introduction Cytology & Histology L1 15 H & E Stain Dewaxing Hydration Xylene I – Xylene II – X: A – 100% Alcohol I – 100% A II - 90% A - 80% A- 70% A- Distilled Cytoplasm Nuclei stain Differentiation stain water – Haematoxylin - Dist. Water – 70% acidic A- running tap water until blueing - Eosin – Dehydration Clearing Mounting 95% A I – 95% AII - 100% A III – 100% A IV – Xylene III – Xylene IV – Canada balsam / D.P.X. 18/10/2023 Introduction Cytology & Histology L1 16 18/10/2023 Introduction Cytology & Histology L1 Video 2 LeicaBiosystems 2018 17 Bright field Light Microscope The paraffin sections are stained with H&E and examined with ordinary light passing through the optical system of a bright microscope. The optical components are the condenser, objective lens, and eyepiece. The condenser focused the light on the objective to be easily studied, the objective lens enlarging and projecting the image of the object and the eyepiece enlarging this image onto the viewer’s retina (Fig. 5). 18/10/2023 Introduction Cytology & Histology L1 18 Light Microscope Fig. 7 Bright light microscope. 18/10/2023 Introduction Cytology & Histology L1 19 Mc Graw-Hill Companies  4X 10X Fig. 8 Sections in lymph nodes in breast cancer Abu Quora et al., 2021 18/10/2023 Introduction Cytology & Histology L1 20  Cytology Cell 1. Cytoplasmic organelles A. Membranous organelles ( cell membrane, Mitochondria, Golgi apparatus, rough Endoplasmic Reticulum, smooth Endoplasmic reticulum, lysosomes) B. Non-membranous organelles ribosomes, cytoskeleton flagella, cilia, microtubules, ) 2. Nucleus 18/10/2023 Introduction Cytology & Histology L1 21 18/10/2023 Introduction Cytology & Histology L1 22