Histology LAB - MIDTERMS

• Histology


  The study of tissues, in particular, normal tissues
• Histologist


  Healthcare professional that performs microscopy to study the structure of normal tissues
• Histopathology


  The study of tissues, in particular, abnormal tissues
• When you are finished with Histology

  You can move on to Histopathology
• Histopathologist


  Healthcare professional that performs microscopy to study the structure of abnormal tissues
• Importance of Studying the Normal Appearance of Tissues


  • Knowing what a normal tissue looks like will help differentiate it from abnormal tissues
  • A med tech should have a thorough understanding of the appearance of normal tissues
• Cytology


  Study of the structure and function of the plant and animal cell
• Organology


  Study of the organs and the organ system
• Microscope is the most useful tool in Histology
• Microscopy


  • One of the most basic and essential skills of a medical technologist, and one that a medical technologist should be excellent in
• Histologic Diagnosis or Microscopy is the gold standard of clinical practice
• Even though we have many advanced machines and genetic testing available, the gold standard of determining abnormalities or cancers in tissues is still histologic diagnosis using microscopy
• Cells make tissues, and tissues make organs
• Brightfield Microscope


  • Contains two lens systems for magnifying specimens – the ocular lens in the eyepiece and the objective lens located in the nose piece
  • Specimen is illuminated by a beam of tungsten light
  • Specimen appears dark against a bright background
  • Limitation – absence of contrast between the specimen and the surrounding medium, which makes it difficult to observe living cells
  • Specimens are usually nonviable, stained preparations

  
• Darkfield Microscope


  • Similar to the ordinary light microscope
  • The condenser system is modified so that the specimen is not illuminated directly
  • The condenser directs the light obliquely so that the light is deflected or scattered from the specimen
  • The specimen appears bright against a dark background
  • Living specimens may be observed more readily

  
• Phase-Contrast Microscope


  • Observation of unstained microorganisms is possible here
  • Includes special objectives and a condenser that makes visible cellular components that differ slightly in their refractive index
  • The image appears dark against a light background

  
• Fluorescent Microscope


  • Used most frequently to visualize specimens that are chemically tagged with a fluorescent dye
  • The source of illumination is ultraviolet (UV) light
  • The ocular lens is fitted with a filter that permits the longer ultraviolet wavelengths to pass, while the shorter wavelengths are blocked or eliminated
  • UV radiations are absorbed by the fluorescent label and the energy is re-emitted in the form of a different wavelength in the visible light range
  • Fluorescent dyes absorb at wavelengths between 230-350 nm and emit an orange, yellow or greenish light
  • Used primarily for the detection of antigen-antibody reactions
  • Antibodies are conjugated with a fluorescent dye that becomes excited in the presence of UV light
  • The fluorescent portion of the dye becomes visible against a black background

  
• Electron Microscope (EM)


  • Provides a revolutionary method of microscopy
  • Magnifies up to one million
  • Permits visualization of submicroscopic cellular particles and virus
  • The specimen is illuminated by a beam of electrons rather than light
  • Focusing is carried out by electromagnets instead of a set of optics
  • Components are sealed in a tube in wherein a complete vacuum is established
  • Specimens should be thinly prepared, fixed, and dehydrated to allow the electron beam to pass through freely

  
• Transmission Electron Microscope


  • Requires specimens that are thinly prepared, fixed, and dehydrated to allow the electron beam to pass through freely
  • As the electrons pass through the specimen, images are formed by directing the electrons onto photographic film – this makes the internal cellular structures visible

  
• Scanning EM


  • Used for visualizing surface characteristics rather than intracellular structures
  • A narrow beam of electrons scans back and forth, producing a three dimensional image as the electrons are reflected off the specimen's surface

  
• Body Tube


  Connects the eyepiece to the objective lens
• Arm


  Supports the body tube and connects it to the base
• Base


  Bottom of the microscope and is used for support
• Mechanical Stage


  • Fixed platform with an opening in the center allows for the passage of light from an illuminating source below to the lens system above the stage
  • Provides a surface for the placement of a slide with its specimen over the central opening
  • Can be moved vertically or horizontally by means of adjustment controls
  • Stage clip – holds the slide in place
• Ocular or Eyepiece Lens


  • Lens at the top of the microscope
  • Usually 10x-15x

  
• Illuminator or Light Source


  • Positioned in the base of the instrument
  • Built in light source – provides direct illumination
  • Mirror (one side flat and other concave) – indirect illumination
  • Uses an external light source (lamp or sunlight) – placed in front of the mirror to direct the light upward into the lens system
  • Flat side – artificial light
  • Concave side - sunlight
• Condenser


  • Found directly under the stage and contains two sets of lenses that collect and concentrate light passing upward
  • Illuminates the object with rays of light that pass through the specimen obliquely as well as directly

  
• Iris Diaphragm


  • Rotating disk under the mechanical stage
  • Can be adjusted to change the intensity and size of the cone of light that is projected upwards into the slide
  • Used to create contrast on cellular components
  • Tissues – open iris diaphragm more
  • Cells or urinary sediments – close iris diaphragm for better contrast
• Revolving Nosepiece or Turret


  • Holds the objective lenses
  • Allows easy rotation of objective lenses
• Scanner Objective (4x)


  • Shortest objective
  • Useful for getting an overview of the slide
  • For WHOLE ORGANS like a section of the spinal cord, lung, digestive tract, etc.
  • Safe to use since it cannot be lowered to the point of contacting the slide
• Low Power Objective (10x)


  • The most useful lens for viewing slides
  • Almost any feature can be featured in this objective
  • Also safe to use since it cannot be lowered to the point of contacting with the slide
• High Power Objective (40x)


  • Also called high-dry objective
  • Used for observing fine detail such as striations in skeletal muscle, types of nerve cells in the retina, etc.
• Oil Immersion Objective (100x)


  • The longest objective
  • Used for observing details of the individual cells such as white blood cells, red blood cells and immature cells
  • Must be used together with immersion oil
• Immersion Oil


  • Light that passes through the glass slide, passes through air, then the lens – this light gets refracted
  • At high magnification, this refraction can blur the image
  • To eliminate refraction between the slide and lens – replace the air gap with immersion oil
  • The immersion oil has the same refractive index as glass
  • Oil Refractive index – 1.518
  • Refractive index – ability to bend light
  • Glass from the slide bends light more than air
  • Light is bent away from the lens as it passes through specimen
  • Immersion oil – same refractive index as glass
  • More light is gathered by the lens – no blurring of image
• Magnification


  • Enlargement or magnification of a specimen is the function of a two-lens system
  • The ocular lens is found in the eyepiece
  • The objective lens is situated in the revolving nose-piece
  • The lenses are separated by the body tube
  • The objective lens is neared the specimen and magnifies it
  • The real image is projected up into the focal plane and then magnified by the ocular lens to produce the final image
• Resolving Power


  • Unlimited enlargement is not possible by merely increasing the magnifying power of the lenses or by using additional lenses
  • Because lenses are limited by a property called RESOLVING POWER
  • Resolving power is the ability of a lens to show two adjacent objects as discrete entities
  • It is dependent on the wave-length of light used and the numerical aperture
• Numerical Aperture


  • Function of a diameter of the objective lens in relation to its focal length
  • It is doubled by use of the substage condenser
• Microscope Care - Once the microscope is on the table


  1. Remove all unnecessary materials (books, papers, purses,etc.)

  2. Uncoil the microscope's electric wire and plug into the outlet

  3. Clean all lens systems – ocular and objectives

  4. Methanol or xylol (xylene) – lens cleanser
• Microscope Care - After using the microscope


  1. Clean all the lenses with a dry, clean lens paper with a drop or two of methanol

  2. Xylol can be used to clean the mechanical stage

  3. Place the objective back to LPO

  4. Center the mechanical stage

  5. Coil the electric wire are the body tube

  6. Return the microscope
• Types of Cells


  • Prokaryotes
  • Eukaryotes