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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
what is dark-field microscopy used to observe?
living, unstained preparations (like small bacterial)
26
dark-field microscopy is ideal for observing...
bacterial motility
27
phase-contrast microscopy converts...
difference in refractive index into detectable variations in light intensity (enhances the contrast between intracellular structures having slight differences in refractive indexes)
28
bacterial components such as _____ and _____ _____ have refractive indices different that that of water
endospores, inclusion bodies
29
the differential interference contract microscope (DIC) creates images by...
detecting differences in refractive indices and thickness of different parts of the specimen
30
why is the differential interference contrast microscope an excellent way to observe living cells?
- live, unstained cells appear brightly colored and three dimensional
31
florescence microscopy uses ______ that absorb high-energy light
fluorophores
32
fluorescence microscopy is very useful for...
labeling molecules of interest within a cell
33
how do labeled antibodies work? (2)
- fluorophores covalently bound to a primary or secondary antibody - antibody binds that specific molecule
34
fluorescently labeling molecules: DNA hybridization (3)
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
FISH assay
fluorescence in situ hybridization
36
fluorescently labeling molecules: gene fusion reporter (2)
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
confocal microscopy uses what two things?
- fluorochromes - laser light
38
what does the laser light in confocal microscopy do?
illuminated each plane in a specimen to produce a three-dimensional image
39
the highest resolution light microscope is about _____
200 nm
40
You are observing a bacterium using a 10x ocular lens and a 45x objective lense. What would the total magnification be?
450x
41
in an electron microscope, electron behave like...
light waves
41
Which microscope uses a laser as a light source?
confocal microscope
41
the high frequency of electron waves allows for what in electron microscopy?
very great resolution
42
how is a sample prepared for transmission electron microscopy? (4)
- sample is fixed with aldehydes to prevent protein movement - flash frozen - sliced very thick - stained with heavy metals to increase contrast
42
what heavy metals can be used to stain to increase contrast? (2)
uranium and osmium
42
for an electron microscope to work, the sample must? (3)
- absorb electrons - be coated with heavy metal - be placed in a vacuum
43
transmission EM can detect molecular complexes like... (3)
- ribosomes - flagellar bases - strands of DNA
44
how is a sample prepared for scanning electron microscopes? (2)
- coated in heavy metals - not sliced
45
in scanning EM, what two things are true?
- the 3D structure is maintained - only the surface is examined
46
preparation methods: negative stain
heavy metals do not penetrate the specimen and render a dark background
47
what is negative stain used for? (2)
the study of viruses, and bacterial gas vacuoles
48
preparation methods: shadowing
coating of a specimen with a thin film of a heavy metal on only one side
49
what is shadowing useful for? (3)
viral morphology, flagella, and DNA
50
cryoEM tomography
rapid freezing technique provides a way to preserve the native state of structures examined in a vacuum
51
how does cryoEM tomography create 3D images?
it records images from many different directions that are pieced together forming the 3D image
52
cryoEM tomography provides extremely high resolution of things like...(5)
- cytoskeletal elements - magnetosomes - inclusion bodies - flagellar motors - viral structures
53
gram-positive bacteria tend to be killed by _____ and _____
penicillin, detergents
54
which bacteria are generally more resistant to antibiotics? gram positive or negative?
gram-negative
55
steps of the gram stain (4)
1. application of crystal violet (purple dye) 2. application of iodine (mordant) 3. alcohol wash 4. application of safranin (counterstain)
56
when stained using the gram stain, gram positive cells appear...
purple
57
when stained using the gram stain, gram negative cells appear...
red
58
acid-fast stain is primarily used for...
bacteria that have waxy material in their cell wall
59
tuberculosis is caused by what bacterium?
mycobacterium tuberculosis
60
leprosy is caused by what bacterium?
mycobacterium leprae
61
negative staining is useful for _____
capsules
62
_____ is required to drive stain into endospores
heat
63
flagella staining requires a _____ to do what?
mordant; make the flagella wide enough to see
64
in what two phases of the gram stain do gram+ and gram- cells appear different? what do they look like?
- 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
What color are gram- bacterial at the completion of the gram staining process?
pink/red
66
What is the correct order of reagents in the gram stain?
1. crystal violet 2. iodine 3. ethanol/alcohol 4. safranin
67
which microscopes allow the best view of bacterial movement?
dark-field microscope

MCB 4403 (7 decks)

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