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MBIO 1010 > Cell Structure and Function > Flashcards

Cell Structure and Function Flashcards

1
Q

Principles of Light Microscopy

A

compound light microscope uses visible light to illuminate cells

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2
Q

Types of Light Microscopy

A
  • Bright-field
  • Phase-contrast
  • Dark-field
  • Fluorescence
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3
Q

Bright-field scope?

A
  • Specimens are visualized because of differences in contrast between specimen
    and surroundings
  • Two sets of lenses form the image
  • Objective lens (usually 10x -100x mag.) & ocular lens (usually 10x – 20x mag.)
  • Total magnification = objective magnification ✕ ocular magnification
  • Maximum magnification is ~2,000✕
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4
Q

Resolution

A

The ability to distinguish two adjacent objects as separate and distinct
- Limit of resolution for light microscope is about 0.2 µm

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5
Q

Magnification

A

The ability to make an object larger

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6
Q

How to calculate Magnification?

A
  • Magnification = ocular x objective
  • ex. Ocular = 10x, Objective = 40x
  • Magnification = 10 x 40 = 400x
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7
Q

Resolution

A
  • The ability of a lens to distinguish small objects that are close together
    - Ex) resolving power of 0.2µm
  • Two points can be distinguished if they are at least 0.2 µm apart
  • Light must pass between two points for them to be viewed as separate objects
  • As wavelength decreases resolution improves
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8
Q

Throw ink-covered objects at target (“E”)

A
  • Basketballs: Cannot fit between arms, poor resolution.
  • Tennis Balls: Fit between arms, resolution improves.
  • Jelly Beans & Beads: As diameter of objects thrown decreases, greater numbers pass between the arms & the resolution increases.
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9
Q

How does improving Contrast in light Microscopy help?

A

Improving contrast results in a better final image

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10
Q

How to improve Contrast?

A
  • Staining improves contrast
  • Dyes are organic compounds that bind to specific cellular materials.
  • Examples of common stains are methylene blue, safranin, and crystal violet
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11
Q

What are the types of staining

A
  • Simple Staining
  • Differential Stains
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12
Q

Simple Staining

A
  • One dye used to color specimen.
  • Chromophore - colored portion of a dye
  • Two types:
    • Basic Dye
    • Acidic Dye
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13
Q

What are the two types of simple staining?

A

Basic Dye: positively charged chromophore.
- Binds to negatively charged molecules on cell surface.
Acidic Dye: negatively charged chromophore
- Repelled by cell surface
- Used to stain background
- Negative stain

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14
Q

Preparing Samples for Staining

A
  1. Preparing a smear: spread culture in thin film over slide -> Dry in air.
  2. Heat fixing and staining: Pass slide through flame to heat fix -> Flood slide with stain; rinse and dry.
  3. Microscopy: -> place drop of oil on slide; examine with 100x objective lens.
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15
Q

Differential Stains

A

The Gram Stain
- Separates bacteria into 2 groups based on cell wall structure .
- Gram Positive
- Gram Negative

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16
Q

Gram Positive

A

Cells that retain a primary stain
- Purple

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17
Q

Gram Negative

A

Cells that lose the primary stain
- Take color of counterstain
- Red or Pink

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18
Q

What are the two differential stains?

A
  • Acid Fast Stain
  • Endospore Stain
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19
Q

Acid Fast Stain

A
  • Detects mycolic acid in the cell wall of
    the genus Mycobacterium
    - Mycobacterium – retains primary stain
    - Fuchsia (pink)
    - Anything else on slide – color of counterstain
    - Blue
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20
Q

Endospore Stain

A
  • Endospores retain primary
    - Green
  • Cells counterstained
    - Pink
  • Ex. Bacillus anthracis spores.
21
Q

Phase-Contrast Microscopy

A
  • Phase ring amplifies differences in the refractive index of cell and
    surroundings.
  • Improves the contrast of a sample without the use of a stain.
  • Allows for the visualization of live samples.
  • Resulting image is dark cells on a light background.
22
Q

Dark Field Microscopy

A
  • Specimen is illuminated with a hollow cone of
    light
  • Only refracted light enters the objective
  • Specimen appears as a bright object on a dark
    background
  • Used to observe bacteria that don’t stain well
    - Ex) Treponema pallidum: the causative agent of syphilis
23
Q

Fluorescence Microscopy

A
  • Used to visualize specimens that fluoresce
  • Emit light of one color when illuminated with
    another color of light
24
Q

Cells in Fluorescence Microscopy?

A
  1. Cells may fluoresce naturally
    - Ex. Photosynthetic Cyanobacteria have
    chlorophyll
    - Absorbs light at 430 nm (blue-violet)
    - Emits at 670 nm (red)
  2. After staining with Fluorescent dye
    - Ex) DAPI specifically binds to DNA
25
Q

Differential interference contrast (DIC) microscopy

A
  • Uses a polarizer to create two distinct beams of polarized light
  • Gives structures such as endospores, vacuoles, and granules a three-
    dimensional appearance
  • Structures not visible by bright-field microscopy are sometimes visible by
    DIC
26
Q

Confocal scanning laser microscopy (CSLM)

A
  • Uses a computerized microscope coupled
    with a laser source to generate a three-
    dimensional image
  • Computer can focus the laser on single
    layers of the specimen
  • Different layers can then be compiled for a
    three-dimensional image
  • Resolution is 0.1 μm for CSLM
27
Q

Electron Microscopy

A
  • Electron microscopes use electrons
    instead of photons to image cells
    and structures
  • Wavelength of electrons is much
    shorter than light → higher
    resolution
28
Q

What are the two types of electron Microscopes?

A
  1. Transmission electron microscopes (TEM)
  2. Scanning electron microscopes (SEM)
29
Q

Transmission Electron Microscopes

A
  • High magnification and resolution (0.2 nm)
  • Specimen must be very thin (20 – 60 nm)
  • Unstained cells do a poor job of scattering electrons
    • Must be stained with metals → lead or uranium
    • Bind to cell structures to make them more electron dense
    • Enables visualization of structures at molecular level
30
Q

Scanning Electron Microscopy (SEM)

A
  • Specimen is coated with a thin film of heavy metal (e.g., gold)
  • An electron beam scans the object
  • Scattered electrons are collected by a detector, and an image is
    produced
  • Allows an accurate 3D image of specimen’s surface.
31
Q

Prokaryotes (Before Nucleus)

A
  • No membrane bound nucleus or organelles
  • Generally smaller than eukaryotes
  • Simple internal structure
  • Divide by binary fission
  • Most are unicellular
32
Q

Bacteria (Eubacteria)

A
  • Diverse metabolism
  • Live in a broad range of
    ecosystems
  • Pathogens and non-pathogens
33
Q

Archaea (Archaebacteria)

A
  • Diverse metabolism
  • Live in extreme environments
  • Non-pathogens
34
Q

What are the Cell Morphology?

A
  1. Coccus (pl. cocci)
    • Roughly spherical
    • EX. Streptococcus pyogenes
  2. Bacillus (pl. bacilli)
    • Rod shaped
    • EX. E. coli
  3. Spirillum (pl. spirilla)
    • Spiral shaped
    • EX. Spirillum volutans
35
Q

Different Cells

A
  • Cells with unusual shapes -> Spirochete -> Treponema pallidum
  • Budding & appendaged bacteria -> Caulobacter crescentus
  • Filamentous bacteria -> Streptomyces griseus
36
Q

Prokaryote Size

A
  • Average:
    - E. coli ~ 1.0 x 3.0 µm
    - Staphylococcus aureus ~ 1.0 µm diameter
  • Very small:
    - Mycoplasma genitalium ~ 0.3 µm
  • Very large:
    • Epulopiscium fishelsonii ~ 80 x 600 µm.
37
Q

Membrane Structure (Cytoplasmic Membrane)

A
  • Cell or plasma membrane
  • Thin structure that surrounds the cell
  • Vital barrier that separates cytoplasm from environment
  • Highly selective permeable barrier; enables concentration of specific
    metabolites and excretion of waste products
38
Q

Membrane Structure (Composition of membranes)

A
  • General structure is phospholipid bilayer
  • Contain both hydrophobic (fatty acid) and hydrophilic (glycerol-phosphate)
    components
  • Can exist in many different chemical forms as a result of variation in the
    groups attached to the glycerol backbone
  • Fatty acids point inward to form hydrophobic environment; hydrophilic
    portions remain exposed to external environment or the cytoplasm
39
Q

Phospholipid Structure (Ester Phospholipids)

A
  • Consist of: glycerol
    • 2 Fatty acids
    • Phosphate
    • Side chain (optional)
40
Q

Phospholipid Structure (Amphipathic)

A

Has both polar and non-polar characteristics

41
Q

Phospholipid Structure (Polar)

A
  • Molecule carries full or
    partial charge
    • Hydrophillic
42
Q

Phospholipid Structure (Non-Polar)

A
  • Molecule is uncharged
    - Hydrophobic
43
Q

Membrane Structure (Cytoplasmic Membrane)

A
  • 8–10 nm wide
  • Embedded proteins
  • Stabilized by hydrogen bonds and hydrophobic interactions.
  • Mg2+ and Ca2+ help stabilize membrane by forming ionic bonds with
    negative charges on the phospholipids
  • Somewhat fluid
44
Q

Membrane Structure (Membrane Proteins)

A
  • Integral membrane proteins
    • Firmly embedded in the membrane
  • Peripheral membrane proteins
    • One portion anchored in the membrane
45
Q

Membrane Structure (Archaeal Membranes)

A
  • Ether linkages in phospholipids of Archaea
  • Bacteria and Eukarya that have ester linkages in phospholipids
46
Q

Membrane Structure (Archaeal Membranes) Part 2

A
  • Archaeal lipids lack fatty acids; have isoprenes instead
  • Major lipids are glycerol diethers and tetraethers
  • Can exist as lipid monolayers, bilayers, or mixture
47
Q

Membrane Function (Permeability Barrier)

A
  • Polar and charged molecules must be transported
  • Transport proteins accumulate solutes against the concentration gradient
  • Prevents leakage and functions as a gateway for transport of nutrients into, and wastes out of, the cell
48
Q

Membrane Function (Protein Anchor)

A
  • Holds transport proteins in place
  • Site of many proteins that participate in transport, bioenergetics, and chemotaxis
49
Q

Membrane Function (Energy Conservation)

A
  • Generation of proton motive force
  • Site of generation and dissipation of the proton motive force

MBIO 1010 (13 decks)

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