Chapter 3 - Observing microorganisms through a microscope

Question 1

What objective lenses does a compound microscope have?

Answer 1

4x
10x
40x
100x (oil immerison)

Question 2

Magnification is

Answer 2

enlarging the appearance of an object
- can be changed by rotating the nosepiece
- total magnification is 1000x bc it includes the 10x ocular lens

Question 3

Resolution is

Answer 3

the ability to distinguish between 2 objects
- resolution is lost if magnification increases beyond 1,000x
- cannot be adjusted

Question 4

Focus is

Answer 4

adjusting the plane of the specimen so the image can be clearly resolved
- accomplished by raising/lowering the stage
- focus can be adjusted using the coarse and fine focus adjustment knobs
- trying to find that perfect clear image

Question 5

what is refractive index?

Answer 5

• refers to how light bends as it moves through the material
• can also cause problems since glass and air have different refractive indexes
• immersion oil is used to minimize refraction at 1000x

Question 6

Describe what needs to be done for a bacterial smear

Answer 6

• stained since most bacteria is colorless
• bacteria must be attached to the slide and then heat fixed

Question 7

What does heat fixed mean?

Answer 7

• kills the bacteria so it stops moving
• dead bacteria picks up stain better

Question 8

What is a simple stain?

Answer 8

involves a single dye so that all cells desire differences in physiology all stain the same color
- methylene blue is common
- this is allows us to observe cell morphology which is cell shape and arrangement

Question 9

What is cell morphology?

Answer 9

refers to the appearance of cells under the microscope
- two main attributes are cell shape and arrangement

Question 10

What are the 3 cell shapes?

Answer 10

1. Spherical - cocci
2. rod-shaped - bacillus
3. other shapes such as curved and spiral

Question 11

What are the types of arrangement of cell morphology (NOT COLONY)?

Answer 11

1. single cells
2. chains = streptop
3. clusters = staphylo
4. other arrangements = diplo

Question 12

What is a gram stain?

Answer 12

1. named after Hans Christian Gram
2. based on cell wall composition (thickness of the peptidoglycan)
3. gram negative - single layer
4. gram positive - multiple layers

Question 13

What is gram stain process?

Answer 13

1. all cell walls are stained purple with crystal violet regardless of negative or positive
2. then once locked in iodine is applied - thick cell walls
3. purple color is washed out with a thin cell - alcohol - decolorizer
4. finally ALL cells receive a pink stain called safranin as a counter stain

Question 14

What is an Acid Fast stain?

Answer 14

• since some bacteria have a waxy cell due to mycolic acid in the cell wall acid fast stain is needed
• appears as pink
• primary stain with heat
• decolorizer - acid alcohol
• counterstain with methylehene blue

Question 15

What is an endospore stain?

Answer 15

• when bacteria form hardy structures called endospores, this cell surrounds its DNA with several protective layers
• Primary stain - malachite green with heat
• decolorizer - water
• counter stain - safranin

Question 16

What is a capsule stain?

Answer 16

• some bacteria are covered in a gelatinous material called a capsule
• capsules are common to bacterial pathogens as they function in immune evasion so immune system doesn’t recognize it so doesn’t respond to it
• doesn’t stain well so have to stain everything NEGATIVE except the capsule

Question 17

Describe the process of the capsule stain

Answer 17

1. india ink - provides color to the background
2. counterstain - stains the cell and could be any color

Question 18

List the two types of other microscopy

Answer 18

1. electron
2. fluorescence

Question 19

What is electron microscopy?

Answer 19

• uses electrons instead of photons (light)
• shortwave length = greater resolution
• used for viruses
  two types
  1. transmission TEM - internal structures can be seen
  2. Scanning SEM - external structures and topography can be seen

Question 20

What is fluorescence microscopy?

Answer 20

since some molecules have the ability to absorb light at one wavelength and emit at a different wavelength we use fluorescence