Difference between revisions of "Histologic"

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'''Acknowledgements'''
  
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== 1. Histology ==
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This Virtual Microscopy Histology Laboratory Manual is a derivative work from the laboratory teaching materials produced over many years by anatomists from the University of Alabama at Birmingham School of Medicine.  The instructors who designed the curriculum, acquired the teaching slide sets and developed this laboratory manual were:  George Hand, Ph.D., Jim Sheetz, Ph.D. and Laura Cotlin, Ph.D.
  
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Histology is the study of the microscopic structure of biological material and the ways in which individual components are structurally and functionally related. It is central to medical science since it stands at the crossroads between biochemistry, molecular biology and physiology on the one side, and pathologic processes that cause disease on the other.  Although often thought of as an archaic discipline, practical knowledge of histology is in actuality an integral part of modern investigative techniques and current medical practice
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The virtual microscopy slides described in this manual are primarily scans of original glass slides used in the University of Alabama at Birmingham School of Medicine Cell Biology and Histology teaching program.  Additional virtual microscopy slides were kindly contributed by:  James L. Fishback, MD, University of Kansas School of Medicine; Mary Ann Sens, MD, PhD, University of North Dakota School of Medicine; and Richard M. Conran, M.D., Ph.D., Uniformed Services University of the Health Sciences.
  
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In this laboratory manual we will focus on the basic structure of human tissues.  We will concentrate on structure-function correlations that are important in the understanding of disease processes.  Thus, we will not attempt to provide a comprehensive review of all structures in the body; instead we will focus just on the structural relationships that are integral to disease.
 
  
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Almost all of the tissues we will review are human tissues obtained at autopsy or from surgical biopsies.  As a general rule all fresh tissues are fixed in 10% neutral buffered formalin and are embedded in paraffin wax before cutting microscopic tissue sections.  The embedding process requires dehydration of the tissues using organic solvents, permeation of the tissues with paraffin wax, and hardening of the wax for cutting.  Tissue sections are then cut at 5 to 7 m in thickness and placed on glass slides.  The tissues are then rehydrated and stained.  This dehydration-wax embedding - rehydration cycle results in dissolution of any lipid materials within the tissues.  This may lead to alterations in the morphology of tissues.  However, if you understand the process you can overlook these artifacts and still make accurate assessments of the tissue.  One classic “artifact” is the loss of fat from liver tissue obtained from a patient with fatty liver.  This leaves holes in the tissue where the fat globules had been situated before they were dissolved away.  These and other classic artifacts will become second nature to you as you review tissue sections. 
 
  
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== Overview of Tissue Preparation and Staining for Microscopy ==
 
  
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1.  '''Obtaining tissues''' - Human material is obtained at autopsy or from surgical biopsies.
 
  
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2.  '''Fixation''' - To preserve the tissue, it is placed immediately in a fixative which acts to preserve the cell and tissue constituents in as lifelike a manner as possible after death.  In postmortem tissue, considerable autolysis may have occurred prior to fixation.  Formalin (10%) is the fixative most often used by pathologists.
 
  
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3.  '''Dehydration''' - The fixed tissues must be dehydrated in order to embed them in paraffin for sectioning.  Water is removed from the tissues by passing them through a series of increasingly concentrated solutions of alcohol.
 
  
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4. '''Clearing''' - Absolute alcohol is not miscible with paraffin. Thus, the alcohol must be removed from the tissue and replaced with an agent that mixes with molten paraffin. The most commonly used clearing agent is xylene.  The xylene makes the tissues translucent or “clears” them.
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Peter G. Anderson, D.V.M., Ph.D.
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Professor & Director of Pathology Undergraduate Education
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Department of Pathology
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University of Alabama at Birmingham
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Volker Hall, 213
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1670 University Boulevard
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Birmingham, Alabama 35294-0019
  
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5.  '''Embedding''' - Following clearing, the tissue is placed in the embedding agent, molten paraffin, and allowed to steep until the tissue is thoroughly infiltrated by the embedding medium.  The preparation is then cooled, the paraffin solidifies, and the block of tissue can now be cut with a minimum of distortion.  The paraffin infiltrates the interstices of the tissue and thus provides internal support as well as external support for sectioning.
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Phone: 205-934-2414
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Email Address: pga@uab.edu
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6.  '''Sectioning''' - The tissue is now cut into very thin slices, usually 5 to 7 m, with a microtome.  The sections are then mounted on glass slides and stained.
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Web page: http://peir.net
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7.  '''Staining''' - For morphologic study, it is necessary to create color contrasts in the tissues by staining.  Certain terms are used to distinguish the staining reaction of a cell.  The term basophilic indicates that the structure can be stained with the basic dye hematoxylin. All nuclei are basophilic.  Cytoplasmic elements may be either basophilic, acidophilic or, neutral.  Eosin is the most commonly used acid stain and any acid components that stain positive with eosin are termed eosinophilic.
 
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8.  '''Other stains''' used in preparing slides – Most slides for histology and pathology are stained with Hematoxylin and Eosin (H&E).  Additional staining techniques are utilized to demonstrate specific characteristics of tissues.  In any staining process variations in the tissue and the technical procedure may lead to minor color modifications in individual slides, but, in general, the reactions are as stated below.
 
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a. '''Masson’s Trichrome stain''' (hematoxylin, acid fuchsin, and aniline blue): nuclei stain black or dark blue; cytoplasm stains red by the acid fuchsin; reticular and collagen fibers stain blue with aniline blue.
 
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b. Gomori’s Trichrome stain: Another version of a trichrome stain that stains nuclei - red-purple; normal muscle myofibrils - green-blue with distinct A and I bands; intermyofibrillar muscle membranes – red; and interstitial collagen - green
 
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c. '''Periodic acid-Schiff’s reagent (PAS)'''.  The PAS method stains glycogen, mucin, connective tissue fibers, and other structures that contain carbohydrates, pink, red, or maroon.  The periodic acid converts adjacent 1, 2 glycol groups to aldehydes and the basic Fuchsin of Schiff’s reagent stains the aldehydes.  Sometimes Hematoxylin is used as a counter stain giving you a PASH.
 
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c. '''Silver stain'''.  This special procedure employs silver nitrate to specifically demonstrate reticular fibers, neurofibrils of neurons and granules in enteroendocrine cells.  These structures are stained black whereas other tissue components may take on a faint gray background stain without revealing detail.
 
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d. '''Toluidine blue'''.  Used to demonstrate granules in mast cells.  Nuclei are deep blue; mast cell granules are reddish-purple.
 
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e. '''Verhoeff-Van Gieson stain (VVG)'''.  This method is used for identifying elastic fibers in tissues such as skin, aorta, etc.  The elastic fibers will be stained blue-black and background will be stained yellow.
 

Revision as of 19:59, 30 May 2014

Chapter 1: Histology

Chapter 2: The Cell

Chapter 3: Epithelial Cells

Chapter 4: Support Cells and the Extracellular Matrix

Chapter 5: Contractile Cells

Chapter 6: Nervous Tissue

Chapter 7: Blood Cells

Chapter 8: Immune System

Chapter 9: Blood and Lymphatic Circulatory Systems and Heart

Chapter 10: Respiratory System

Chapter 11: Alimentary Tract

Chapter 12: Liver

Chapter 13: Musculoskeletal System

Chapter 14: Endocrine System

Chapter 15: Urinary System

Chapter 16: Male Reproductive System

Chapter 17: Female Reproductive System

Chapter 18: Skin and Breast

Chapter 19: Special Senses


Acknowledgements

This Virtual Microscopy Histology Laboratory Manual is a derivative work from the laboratory teaching materials produced over many years by anatomists from the University of Alabama at Birmingham School of Medicine. The instructors who designed the curriculum, acquired the teaching slide sets and developed this laboratory manual were: George Hand, Ph.D., Jim Sheetz, Ph.D. and Laura Cotlin, Ph.D.

The virtual microscopy slides described in this manual are primarily scans of original glass slides used in the University of Alabama at Birmingham School of Medicine Cell Biology and Histology teaching program. Additional virtual microscopy slides were kindly contributed by: James L. Fishback, MD, University of Kansas School of Medicine; Mary Ann Sens, MD, PhD, University of North Dakota School of Medicine; and Richard M. Conran, M.D., Ph.D., Uniformed Services University of the Health Sciences.




Peter G. Anderson, D.V.M., Ph.D. Professor & Director of Pathology Undergraduate Education Department of Pathology University of Alabama at Birmingham Volker Hall, 213 1670 University Boulevard Birmingham, Alabama 35294-0019

Phone: 205-934-2414 Email Address: pga@uab.edu Web page: http://peir.net

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