Chapter 3

  1. Size of a eukaryotic cells
    10 - 500 micrometer
  2. Size of prokaryotic cells
    1 - 10 micrometer
  3. Size of a virus
    20nm - 1000nm(1mincrometer)
  4. Resolution
    ability of the lenses to distinguish between two points
  5. Describe resolution of a light microscope
    ex. virus 20 nm and microscopes's resolving power is 25nm the microscope will not be able to see it.
  6. Refractive index
    is the light - banding ability of a medium
  7. Why is there such a thing as an "oil immersion" lens on lirght micros.?
    Ther shorter waveleneghts of light provied greater resolution. Immersion oil is used to keep light from bending.
  8. Brightfield illumination
    dark objects are visible againsgt a bright backround. Light reflected off the specimen does not enter the objective lens.
  9. Electron microscope
    • uses electron insted of light
    • shorter wavelength of electrons gives greater resolution.
  10. Stains
    have postivie and negative ions
  11. Basic dye
    • color due to cation (+)
    • cristal violet, safranin... stain the bacteria
  12. Acidic dye
    • color due to anion (-)
    • stain the background not the bacteria
  13. How is a specimen prepaired?
    microbes are smeared an a slide - air dry - fix by flaiming killing microbes, than stain.
  14. Simple stain
    use of a single basic dye - stains the entire microorganism to show bgasic shape and structure.
  15. What is a mordant?
    may be used to intensify the stain, increase the affinity of the stain for the specimen, or to coat the specimen to enlarge it. ex. iodine.
  16. Differential stain
    these stians react differently with different kinds of bacteria, and allow us t otell the difference bwtween them.
  17. The Gram stain
    • Gram - positive
    • Gram - negative
  18. 1st step making a Gram stain after the bacterial smear is made and heat fixed.
    • Smea is flooed with crystal violet for 60 sec. than rinsed with distilled water
    • ~crystal violet sticks to peptidoglycan in the cell wall
    • it is a primary stain -colors all the cell and all will look purple G+ and G-
  19. 2nd Step making a Gram stain
    • flood with iodine for 60 second than rinse with distilled water
    • ~iodine is a mordant will increase the affinity of the cyrstal violet
  20. 3rd step making gram stain ( the differential step)
    • flood with an acetone/ethanol wash (decolar agent) for 5 seconds than rinse w/distilled water.
    • in Gram + cells look purple
    • Gram - look clear the ethanol washes CV-I of peptidoglycan layers.
  21. 4th step making Gram stain
    flood with safranin stain for 1.5-2 min, then rinsed with distilled water
  22. Safranin -counterstain
    • is pink in color and stain all bacteria
    • G + will not change the color and still look purple
    • G - was clear and will be stained pink.
  23. Grap positive bacteria
    • they have a thick peptidogllycan layer in their cell wall (disaccharides and amino acids) which is the part that stains purpel due to crystal violet and iodine wash.
    • Killed easily by penecilin
  24. Gran negative bacteria
    • Thin layer of peptidoglycan
    • Contain a layer of lipopolysaccharide (endotoxin) as part of their cell wall - more resistant to antibiotics which cannot penetrate this lipo layer.
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Chapter 3