Biology module 2.1

• All living organisms are made of cells, there are several different types of cells, some of them sharing some common features
• Humans are made up of eukaryotic cells
• All eukaryotic cells contain a nucleus and membrane bound organelles
• Ultrastructure


  Detailed structure of cells that can be obtained by using a microscope
• Nucleus


  • Surrounded by a double membrane called the envelope containing pores which enable molecules to enter and leave the nucleus
  • Contains chromatin and a nucleolus which is the site of ribosome production
• Rough endoplasmic reticulum (RER)


  • Series of flattened sacs enclosed by a membrane with ribosomes on the surface
  • Folds and processes proteins made on the ribosomes
• Smooth endoplasmic reticulum (SER)


  • System of membrane bound sacs
  • Produces and processes lipids
• Golgi apparatus


  • Series of fluid filled, flattened & curved sacs with vesicles surrounding the edges
  • Processes and packages proteins and lipids
  • Produces lysosomes
• Mitochondria


  • Usually oval shaped, bound by a double membrane called the envelope
  • Inner membrane is folded to form projections called cristae with matrix on the inside containing all the enzymes needed for respiration
• Centrioles


  • Hollow cylinders containing a ring of microtubules arranged at right angles to each other
  • Involved in cell division
• Ribosomes


  • Composed of two sub units
  • Site of protein production
• Lysosome


  • Vesicle containing digestive enzymes bound by a single membrane
• Cytoskeleton


  • Plays an important role in providing mechanical strength as well as aiding transport within cells and enabling cell movement
• Protein transport


  1. Proteins produced on ribosomes

  2. Proteins produced on surface of RER folded and processed in RER

  3. Proteins transported from RER to Golgi apparatus in vesicles

  4. Proteins modified in Golgi apparatus

  5. Golgi apparatus packages proteins into vesicles to be transported around the cells
• Prokaryotic cells


  • Cell wall - Rigid outer covering made of peptidoglycan
  • Capsule - Protective slimy layer which helps the cell to retain moisture and adhere to surfaces
  • Plasmid - Circular piece of DNA
  • Flagellum - a tail like structure which rotates to move the cell
  • Pili - Hair-like structures which attach to other bacterial cells
  • Ribosomes - Site of protein production
• Microscopy


  The use of microscopes to analyse cell components and observe organelles
• Magnification


  How many times bigger the image produced by the microscope is than the real-life object you are viewing
• Resolution


  The ability to distinguish between objects that are close together (i.e. the ability to see two structures that are very close together as two separate structures)
• Types of microscopes


  • Optical (light) microscopes
  • Electron microscopes
  • Laser scanning confocal microscopes

  
• Optical (light) microscopes


  • Use light to form an image
  • Have a maximum resolution of around 0.2 micrometres (µm) or 200 nm
  • Can be used to observe eukaryotic cells, their nuclei and possibly mitochondria and chloroplasts
  • Cannot be used to observe smaller organelles such as ribosomes, the endoplasmic reticulum or lysosomes
  • Have a maximum useful magnification of about ×1500

  
• Electron microscopes


  • Use electrons to form an image
  • Have a maximum resolution of around 0.0002 µm or 0.2 nm (i.e. around 1000 times greater than that of optical microscopes)
  • Can be used to observe small organelles such as ribosomes, the endoplasmic reticulum or lysosomes
  • Have a maximum useful magnification of about ×1,500,000

  
• Types of electron microscopes


  • Transmission electron microscopes (TEMs)
  • Scanning electron microscopes (SEMs)

  
• Transmission electron microscopes (TEMs)


  • Use electromagnets to focus a beam of electrons
  • The beam of electrons is transmitted through the specimen
  • Denser parts of the specimen absorb more electrons, making these denser parts appear darker on the final image produced
  • Can give high-resolution images (more detail)
  • Can only be used with very thin specimens or thin sections of the object being observed
  • Cannot be used to observe live specimens
  • Do not produce a colour image
• Scanning electron microscopes (SEMs)


  • Scan a beam of electrons across the specimen
  • The beam bounces off the surface of the specimen and the electrons are detected, forming an image
  • Can produce three-dimensional images that show the external, 3-D structure of specimens
  • Give lower resolution images (less detail) than TEMs
  • Cannot be used to observe live specimens
  • Do not produce a colour image

  
• Laser scanning confocal microscopes


  • The cells being viewed must be stained with fluorescent dyes
  • A thick section of tissue or small living organisms are scanned with a laser beam
  • The laser beam is reflected by the fluorescent dyes
  • Multiple depths of the tissue section/organisms are scanned to produce an image
  • Can be used on thick or 3-D specimens
  • Allow the external, 3-D structure of specimens to be observed
  • Produce very clear, high resolution images
  • Can see the structure of the cytoskeleton in cells
  • It is a slow process and takes a long time to obtain an image
  • The laser has the potential to cause photodamage to the cells
• Optical microscopes cannot resolve (distinguish between) two objects that are closer than half the wavelength of visible light (500-650 nanometres)
• Optical microscopes have a maximum resolution of around 0.2 micrometres (µm) or 200 nm
• Electron microscopes have a maximum resolution of around 0.0002 µm or 0.2 nm (i.e. around 1000 times greater than that of optical microscopes)
• The maximum useful magnification of optical microscopes is about ×1500
• The maximum useful magnification of electron microscopes is about ×1,500,000
• Preparing a slide using a liquid specimen


  1. Add a few drops of the sample to the slide using a pipette

  2. Cover the liquid/smear with a coverslip and gently press down to remove air bubbles

  3. Wear gloves to ensure there is no cross-contamination of foreign cells
• Preparing a microscope slide using a solid specimen


  1. Take care when using sharp objects and wear gloves to prevent the stain from dying your skin

  2. Use scissors to cut a small sample of the tissue

  3. Peel away or cut a very thin layer of cells from the tissue sample to be placed on the slide (using a scalpel or forceps)

  4. Apply a stain

  5. Gently place a coverslip on top and press down to remove any air bubbles
• Preparing a microscope slide using a solid specimen (alternative method)


  1. Treat the tissue sample with chemicals to kill/make the tissue rigid

  2. Fix the specimen using formaldehyde (preservative), dehydrate it using a series of ethanol solutions, impregnate it in paraffin/resin for support then cut thin slices from the specimen using a microtome

  3. Remove the paraffin from the slices/specimen, apply a stain and mount using a resin

  4. Gently place a coverslip on top and press down to remove any air bubbles
• Preparing a microscope slide using a solid specimen (alternative method)


  1. Freeze the specimen in carbon dioxide or liquid nitrogen

  2. Cut the specimen into thin slices using a cryostat

  3. Place the specimen on the slide and add a stain

  4. Gently place a coverslip on top and press down to remove any air bubbles
• When using an optical microscope always start with the low power objective lens
• Adding a drop of water to the specimen (beneath the coverslip) can prevent the cells from being damaged by dehydration
• Using a graticule to take measurements of cells


  1. Place a graticule (engraved ruler) into the eyepiece of the microscope

  2. Calibrate the graticule using a stage micrometer (engraved scale on a microscope slide)

  3. This allows the number of micrometers each graticule unit is worth to be determined

  4. The calibrated graticule can then be used as a ruler in the field of view
• The size of cells or structures of tissues may appear inconsistent in different specimen slides due to the 3D nature of cells and tissues being cut at different planes
• Optical microscopes do not have the same magnification power as other types of microscopes and so there are some structures that cannot be seen
• The treatment of specimens when preparing slides could alter the structure of cells