MIDTERM Histo Lab

• Types of microscopes


  • Brightfield microscope
  • Darkfield microscope
  • Phase-contrast microscope
  • Fluorescent microscope
  • Electron microscope

  
• 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 an ultraviolet (UV) light
  • High pressure mercury lamp or hydrogen quarts lamp – light source
  • 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 EM


  • 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

  
• Parts of a compound brightfield microscope


  • Body tube
  • Arm
  • Base
  • Mechanical stage
  • Ocular or eyepiece lens
  • Illuminator or light source
  • Condenser
  • Iris diaphragm
  • Revolving nosepiece or turret

  
• 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 is cannot be lowered to the point of contacting with 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 is 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

  
• 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 or resolution


  • 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

  
• The epithelial tissues cover external body surfaces and provide lining of the internal surfaces
• Categories of epithelial tissues


  • Simple epithelium
  • Stratified epithelium
  • Glandular or pyramidal epithelium
• Simple epithelium


  • Composed on one layer of cells all attached to the basement membrane
• Types of simple epithelium

  • Simple squamous epithelium
  • Simple cuboidal epithelium
  • Simple columnar epithelium
  • Pseudostratified columnar epithelium
• Goblet cell


  Column-shaped cell found in the respiratory and intestinal tracts that secrets the main component of mucus
• Stratified epithelium


  • Composed of more than one layer of cells
  • Only the basal cells are attached to the basement membrane
• Types of stratified epithelium


  • Stratified squamous epithelium
  • Stratified cuboidal epithelium
  • Stratified columnar epithelium
  • Transitional epithelium
• Stratified squamous (non-keratinized)


  Opening of the mouth, esophagus and vagina
• Stratified squamous (keratinized)


  Epidermis of the skin
• Stratified cuboidal epithelium


  Sweat gland ducts
• Stratified columnar epithelium


  Male urethra, conjunctiva, excretory ducts
• Transitional epithelium


  Excretory passage of urinary tract (ureter, urinary bladder and urethra)
• Types of glandular or pyramidal epithelium


  • Serous glandular epithelium
  • Mucous glandular epithelium
  • Mixed glandular epithelium
• Serous glandular epithelium


  Pancreas, parotid glands
• Mucous glandular epithelium


  Sublingual glands
• Mixed glandular epithelium


  Submaxillary glands
• Types of connective tissues


  • Proper – connective tissue
  • Cartilage – connective tissue
  • Bone – connective tissue
  • Blood – connective tissue
• Mesenchyme


  • Loose and fluid type of embryonic tissues
  • Common origin of all connective tissues
• Characteristics of connective tissues


  • Different degrees of vascularity (blood flow)
  • Its cells are separated by a large amount of intercellular space
• Components of connective tissues


  • Extracellular Matrix
  • Non-living component of connective tissues
  • Inert matrix
  • Ground Substance
  • Fibers
• Ground substance


  Watery, rubbery, unstructured mat