Histology Lecture Introduction PDF

1st lecture in general histology Introduction By Dr.Maya AL-Joukhadar introduction-mayajoukhadar  Histology Histology: Latino term histo=tissue –logy =suffix denoting a subject of study or interest. DEFINE :branch of anatomy that study the tissues of the body and...

1st lecture in general histology Introduction By Dr.Maya AL-Joukhadar introduction-mayajoukhadar  Histology Histology: Latino term histo=tissue –logy =suffix denoting a subject of study or interest. DEFINE :branch of anatomy that study the tissues of the body and how these tissues are arranged to constitute organs This subject involves all aspects of tissue biology With the focus on how cells’ structure and arrangement optimize functions specific to each organ. introduction-mayajoukhadar  Cells Space between cells(ECM) But quantity of these components and the way of cell arrangement varies from tissue to another introduction-mayajoukhadar Tissues have two interacting component : – Cells the basic structural and functional units, the smallest living parts of the body (have membrane limited nuclei surrounded by cytoplasm which contains various membrane- limited organelles and the cytoskeleton, these cells differ in origin –shape- biology-function- movement..etc according to its tissue) – Extracellular matrix (ECM) which is productive and consists of: Ground substance(GS) Fibers ECM functions: supports the cells-the fluid transporting nutrients to the cells, and carrying introduction-mayajoukhadar away their wastes and secretory products  Cells produce the ECM locally and are in turn strongly influenced by matrix molecules. Many matrix components bind to specific cell surface receptors that span the cell membranes and connect to structural components inside the cells introduction-mayajoukhadar introduction-mayajoukhadar  MANY ORGANS MAKE SYSTEM : CELL FIRST UNIT IN THE BODY CELLS +ECM= TISSUE Four basic tissue types: MANY TISSUE MAKE ORGAN introduction-mayajoukhadar The small size of cells and matrix components in tissues makes histology dependent on the use of microscopes and molecular methods of study. But we need to study these tissue and cells in the state that they are in the body So we need PREPARATION OF TISSUES FOR STUDY introduction-mayajoukhadar  PREPARATION OF TISSUES FOR STUDY the preparation of tissue slices or “sections” that can be examined visually with transmitted light. The ideal microscopic preparation is preserved so that the tissue (which taken from biopsy) on the slide has the same structural features it had in the body. However, this is often not feasible because the preparation process can remove cellular lipid, with slight distortions of cell structure. introduction-mayajoukhadar  Biopsy: tissue which taken from the body to examine under microscope introduction-mayajoukhadar fixation introduction-mayajoukhadar  Fixation To preserve tissue structure and prevent degradation and alteration subsequent to death made by enzymes released from the cells or microorganisms, and maintain its normal architecture pieces of organs are placed as soon as possible after removal from the body in solutions of stabilizing or cross-linking compounds called fixatives: – formalin,a buffered isotonic solution of 10% formaldehyde ,react with the amine groups (NH2) of proteins. – for Electron microscopy glutaraldehyde treated tissue is then immersed in buffered osmium tetroxide, which preserves (and stains) cellular lipids as well as proteins. – Bouin’s fluid introduction-mayajoukhadar  Dehydration having its water extracted gradually by transfers through a series of increasing ethanol solutions, beginning 50% and ending in 100% ethanol. clearing The ethanol is then replaced by an organic solvent miscible with both alcohol and the embedding medium,xylene because infiltration with the reagents used here gives the tissue a translucent appearance introduction-mayajoukhadar embedding introduction-mayajoukhadar introduction-mayajoukhadar  infiltration and then embedding The fully cleared tissue is then placed in melted paraffin in an oven at 52°-60°C. Then it is placed into a small receptacle covered with melted paraffin and let it to harden at room temperature forming a paraffin block containing the tissue introduction-mayajoukhadar sectioning introduction-mayajoukhadar  Sectioning &mounting Sectioning is placing and trimming in an instrument called a microtome. Paraffin sections are typically cut at 3-10 μm thickness for light microscopy, but electron microscopy requires sections less than 1 μm thick. One micrometer (1 μm) equals 1/1000 of a millimeter (mm). The sections are placed on glass slides(mounting) and stained for light microscopy or on metal grids for electron microscopic staining and examination introduction-mayajoukhadar  Frozen section If results of such analyses are required before the medical procedure is completed, for example to know whether a growth is malignant before the patient is closed, a much more rapid processing method is used. The biopsy is rapidly frozen in liquid nitrogen, preserving cell structures and making the tissue hard and ready for sectioning. A microtome called a cryostat in a cabinet at subfreezing temperature is used to section the block with tissue, and the frozen sections are placed on slides for rapid staining and microscopic examination by a pathologist also effective in histochemical studies of very sensitive enzymes also useful when structures containing lipids introduction-mayajoukhadar staining introduction-mayajoukhadar  staining Most cells and extracellular material are completely colorless, and to be studied microscopically tissue sections must be stained (dyed). The purpose is to make tissues distinguishable from one another Dyes stain material more or less selectively, often behaving like acidic or basic compounds and forming electrostatic (salt) linkages with ionizable radicals of macromolecules in tissues Cell components such as nucleic acids with a net negative charge have an affinity for basic dyes and are termed basophilic proteins with many ionized amino groups, stain more readily with acidic dyes and are termed acidophilic Examples of basic dyes : Hematoxylin , toluidine blue, alcian blue, methylene blue. Acid dyes (ex , eosin, orange G, and acid fuchsin) introduction-mayajoukhadar introduction-mayajoukhadar staining methods: H&E(purple neucleus&pink cytoplasm ) trichrome stains :Masson trichrome (mallory &Masson) (dark blue neucleus&red cytoplasm&blue ECM ) Van gieson :(dark blue neucleus&yellow cytoplasm &red collagen ) PAS (periodic acid-Schiff) pink carbohydrate-rich tissue (mucin/ basement membrane) Sudan black (lipid/myelin ) Silver/gold methods(metallic salts) neurones orcein for elastic tissue introduction-mayajoukhadar H&E PAS Masson trichrome orcein for elastic tissue introduction-mayajoukhadar Slide preparation, from tissue fixation to observation with a light microscope, may take from 12 hours to 2½ days, depending on : the size of the tissue, the embedding medium, and the method of staining. The final step before microscopic observation is mounting a protective glass coverslip on the slide with clear adhesive.)‫(بلسم كندا‬ introduction-mayajoukhadar  Types of microscopes Light microscopes ELECTRON microscopes introduction-mayajoukhadar  Light MICROSCOPY based on the interaction of light with tissue components and are used to reveal and study tissue features Bright-field microscopy, ‫*مجاهرنا‬ Fluorescence‫المتألق‬, ultraviolet (UV) light and filters, different wavelengths emitted by the substances to be visualized. dark background, blue green yellow fluorescence Phase-contrast‫متباين الطور‬, principle that light changes its speed when passing through cellular and extracellular structures with different refractive indices,colorless but lighter or darker Confocal‫متحد البؤر‬, increase contrast ,achieves high sharpness, Digital images can be analyzed quantitatively using appropriate software gives 3D images Focuspolarizing microscopy‫المستقطب‬. recognition of stained or unstained structures made of highly organized subunits, normal light passes through a polarizing filter introduction-mayajoukhadar fluorescence phase-contrast bright-field microscopy, introduction-mayajoukhadar focuspolarizing microscopy  ELECTRON MICROSCOPY based on the interaction of tissue components with beams of electrons The wavelength in an electron beam is much shorter than that of light, allowing a 1000-fold increase in resolution. Transmission Electron Microscopy(TEM)‫نافذ‬ Scanning electron microscopy (SEM) ‫ماسح‬ present a three-dimensional view introduction-mayajoukhadar introduction-mayajoukhadar  Bright-field microscopy stained tissue is examined with ordinary light passing through the preparation this microscope includes an optical system and mechanisms to move and focus the specimen The optical components are: – the condenser )‫ (المكثف‬focusing light on the object to be studied – the objective )‫ (العدسة الجسمية‬lens enlarging and projecting the image of the object toward the observer; – and the eyepiece (or ocular lens) )‫ (العدسة العينية‬further magnifying this image and projecting it onto: a- the viewer’s retina or. B- charge coupled device (CCD) highly sensitive to low light levels with a camera and monitor. resolving power ) ‫ (القوة التمييزية‬The critical factor in obtaining a crisp, detailed image defined as the smallest distance between two structures at which they can ben seen as separate objects. The maximal resolving power of the light microscope is approximately 0.2 μm, which can permit clear images magnified 1000-1500 times. microscope’s resolving power determines the quality of the image which depends mainly onintroduction-mayajoukhadar the quality of its objective lens.  ‫ضابط المسافة بين الحدقتين‬ ‫عدسة عينية‬ ‫عدسة جسمية‬ ‫مالقط تثبيت العينة‬ ‫لوحة التشغيل‬ ‫مكثفة‬ ‫لوحة ضبط كثافة‬ ‫االضاءة‬ ‫‪introduction-mayajoukhadar‬‬ ‫لولب تحريك أمام خلف‬ ‫– يمين يسار‬ introduction-mayajoukhadar