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MCB 4403 > Lecture 2 > Flashcards

Lecture 2 Flashcards

1
Q

What is the resolution of the human eye?

A

150 micro meters

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2
Q

How many lenses do microscopes have and what is their magnification?

A

1 lense and 70-25 times magnification

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3
Q

How do you increase resolution? (3)

A
  • Use shorter wavelength light
  • Immerse in oil
  • Wider lense closer to specimen
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4
Q

what is the refraction index?

A

The light bending ability of a medium

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5
Q

what is immersion oil used for?

A

Immersion oil is used to keep light from bending, resulting in an increase in resolution

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6
Q

what do compound microscopes do?

A

form images using 2 or more lenses

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7
Q

What is the dark field microscope used for?

A
  • Used to observe living unstained things such as small bacteria
  • Ideal for observing bacterial motility
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8
Q

What does a Phase-Contrast microscope do?

A

Enhances the contrast between intracellular structures

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9
Q

!What does DIC stand for?

A

Differential Interference Contrast (microscope)

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10
Q

1 um = _____ m = _____ mm

A

10^-6, 10^-3

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11
Q

1 nm = _____ m = _____ mm

A

10^-9, 10^-6

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12
Q

1 um = ______ nm

A

1000

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13
Q

resolution of our eyes depends on what?

A

the distance between two photoreceptors cells in the retina

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14
Q

how many lenses does a simple microscope have?

A

one

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15
Q

what is the approximate magnification of a simple microscope?

A

~70x-25x

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16
Q

total magnification =

A

objective lens x ocular lens

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17
Q

in a compound microscope, the image from the _____ lens is magnified again by the _____ lens

A

objective, ocular

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18
Q

what are ways of increasing resolution? (3)

A
  • use shorter wavelengths of light
  • immersion oil
  • wider lens closer to specimen
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19
Q

how does immersion oil help magnification?

A

collects more light from specimen

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20
Q

refactive index

A

the light-bending ability of a medium

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21
Q

bright field illumination looks like…

A

dark objects visible against a bright background

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22
Q

in bright field illumination, life reflected off the specimen does/does not enter the objective lens?

A

does not

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23
Q

dark field illumination looks like…

A

light objects visible against a dark background

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24
Q

in dark field illumination, light scattered off the specimen does/does not enter the objective lens?

A

does

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25
Q

what is dark-field microscopy used to observe?

A

living, unstained preparations (like small bacterial)

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26
Q

dark-field microscopy is ideal for observing…

A

bacterial motility

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27
Q

phase-contrast microscopy converts…

A

difference in refractive index into detectable variations in light intensity
(enhances the contrast between intracellular structures having slight differences in refractive indexes)

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28
Q

bacterial components such as _____ and _____ _____ have refractive indices different that that of water

A

endospores, inclusion bodies

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29
Q

the differential interference contract microscope (DIC) creates images by…

A

detecting differences in refractive indices and thickness of different parts of the specimen

30
Q

why is the differential interference contrast microscope an excellent way to observe living cells?

A
  • live, unstained cells appear brightly colored and three dimensional
31
Q

florescence microscopy uses ______ that absorb high-energy light

A

fluorophores

32
Q

fluorescence microscopy is very useful for…

A

labeling molecules of interest within a cell

33
Q

how do labeled antibodies work? (2)

A
  • fluorophores covalently bound to a primary or secondary antibody
  • antibody binds that specific molecule
34
Q

fluorescently labeling molecules: DNA hybridization (3)

A
  1. fluorophores covalently bound to probe
  2. fluor-probe hybridizes with the target DNA via base-pairing
  3. the FISH assay is used to visualize
35
Q

FISH assay

A

fluorescence in situ hybridization

36
Q

fluorescently labeling molecules: gene fusion reporter (2)

A
  1. the gene of interest is fused in frame with the gene of the fluorescent protein
  2. the protein of interest is expressed in fusion with the fluorescent protein
    (think glowing bunnies?)
37
Q

confocal microscopy uses what two things?

A
  • fluorochromes
  • laser light
38
Q

what does the laser light in confocal microscopy do?

A

illuminated each plane in a specimen to produce a three-dimensional image

39
Q

the highest resolution light microscope is about _____

A

200 nm

40
Q

You are observing a bacterium using a 10x ocular lens and a 45x objective lense. What would the total magnification be?

A

450x

41
Q

in an electron microscope, electron behave like…

A

light waves

41
Q

Which microscope uses a laser as a light source?

A

confocal microscope

41
Q

the high frequency of electron waves allows for what in electron microscopy?

A

very great resolution

42
Q

how is a sample prepared for transmission electron microscopy? (4)

A
  • sample is fixed with aldehydes to prevent protein movement
  • flash frozen
  • sliced very thick
  • stained with heavy metals to increase contrast
42
Q

what heavy metals can be used to stain to increase contrast? (2)

A

uranium and osmium

42
Q

for an electron microscope to work, the sample must? (3)

A
  • absorb electrons
  • be coated with heavy metal
  • be placed in a vacuum
43
Q

transmission EM can detect molecular complexes like… (3)

A
  • ribosomes
  • flagellar bases
  • strands of DNA
44
Q

how is a sample prepared for scanning electron microscopes? (2)

A
  • coated in heavy metals
  • not sliced
45
Q

in scanning EM, what two things are true?

A
  • the 3D structure is maintained
  • only the surface is examined
46
Q

preparation methods: negative stain

A

heavy metals do not penetrate the specimen and render a dark background

47
Q

what is negative stain used for? (2)

A

the study of viruses, and bacterial gas vacuoles

48
Q

preparation methods: shadowing

A

coating of a specimen with a thin film of a heavy metal on only one side

49
Q

what is shadowing useful for? (3)

A

viral morphology, flagella, and DNA

50
Q

cryoEM tomography

A

rapid freezing technique provides a way to preserve the native state of structures examined in a vacuum

51
Q

how does cryoEM tomography create 3D images?

A

it records images from many different directions that are pieced together forming the 3D image

52
Q

cryoEM tomography provides extremely high resolution of things like…(5)

A
  • cytoskeletal elements
  • magnetosomes
  • inclusion bodies
  • flagellar motors
  • viral structures
53
Q

gram-positive bacteria tend to be killed by _____ and _____

A

penicillin, detergents

54
Q

which bacteria are generally more resistant to antibiotics? gram positive or negative?

A

gram-negative

55
Q

steps of the gram stain (4)

A
  1. application of crystal violet (purple dye)
  2. application of iodine (mordant)
  3. alcohol wash
  4. application of safranin (counterstain)
56
Q

when stained using the gram stain, gram positive cells appear…

A

purple

57
Q

when stained using the gram stain, gram negative cells appear…

A

red

58
Q

acid-fast stain is primarily used for…

A

bacteria that have waxy material in their cell wall

59
Q

tuberculosis is caused by what bacterium?

A

mycobacterium tuberculosis

60
Q

leprosy is caused by what bacterium?

A

mycobacterium leprae

61
Q

negative staining is useful for _____

A

capsules

62
Q

_____ is required to drive stain into endospores

A

heat

63
Q

flagella staining requires a _____ to do what?

A

mordant; make the flagella wide enough to see

64
Q

in what two phases of the gram stain do gram+ and gram- cells appear different? what do they look like?

A
  • the third (decolorizing agent/alcohol) and fourth (counterstain/safranin) steps
  • gram+ look purple in both
  • gram- are colorless in step three and red in step 4
65
Q

What color are gram- bacterial at the completion of the gram staining process?

A

pink/red

66
Q

What is the correct order of reagents in the gram stain?

A
  1. crystal violet
  2. iodine
  3. ethanol/alcohol
  4. safranin
67
Q

which microscopes allow the best view of bacterial movement?

A

dark-field microscope

MCB 4403 (7 decks)

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