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VMCB 121 Lecture > 3: Bacterial Demonstration > Flashcards

3: Bacterial Demonstration Flashcards

1
Q

why do we stain

A

direct visualization of the organism

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2
Q

process in which bacteria are stained to give color to them (because they are colorless and transparent).

A

bacterial staining

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3
Q

types of dyes

A
  • Cationic/basic - combines with acidic cellular elements
    - e.g., crystal violet, methylene blue, malachite green, safranin
  • Anionic/acidic - combines with alkaline cell elements
    - e.g., acid fuchsin, eosin, nigrosine

!!! Bacteria have an affinity to BASIC dyes due to the acidic nature of their protoplasm

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4
Q

2 components of stains

A

positive ions
negative ions

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5
Q

section of a molecule in a given material that absorb particular wavelengths of visible light causing us to see color

A

chromophore

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6
Q

3 types of stains

A

simple stains
differential stains
special stains

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7
Q

a type of stain that use only one and stains all cells with the same color

give 3 examples

A

simple stain - e.g., methylene blue, crystal violet, safranin, carbol fuchsin

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8
Q

a type of stain that distinguish two kinds of microorganisms

give 2 examples

A

differential stains - e.g., gram stain, acid-fast stain

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9
Q

a type of stain that highlight specific cell structures

give 2 examples

A

special stains - e.g., Hansen’s, Anthony’s, Schaeffer-Fulton

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10
Q

– inactivate enzymes that might disrupt cell morphology and toughens cell structures so they will
not change during staining and observation
- microorganisms are killed and attached firmly to the slide during this method

A

fixation

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11
Q

2 fundamental types of fixation

A

heat fixation - gently flame heating an air-dried film of bacteria and adequately preserve overall morphology but not structures within the cells

chemical fixation - to protect fine cellular substructure and the morphology of larger and more delicate microorganisms

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12
Q
  • differential stain that allows the determination of to protect fine cellular substructure and the morphology of larger and more delicate microorganisms
  • devised by Hans Christian Gram (1884)
A

gram staining

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13
Q

name the reagents in gram staining
- primary stain
- mordant
- decolorizer
- counterstain

A

primary stain - crystal violet
mordant - Lugol’s iodine
decolorizer - acetone alcohol
counterstain - safranin

!!! study procedure !!!

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14
Q

3 mechanisms involved in gram staining

A

thickness of the cell wall
pore size
permeability of the intact cell

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15
Q

4 limitations of gram staining

A
  • high number of microorganisms required
  • liquid samples with low numbers of microorganisms (e.g., cerebrospinal fluid)
  • requires centrifugation to concentrate pathogens
  • pellet then examined after staining
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16
Q

a differential stain introduced by Paul Ehrlich (1882) and modified by Ziehl and Neelsen (1883)

used for microorganisms that are not staining
by simple or Gram staining method

A

acid-fast staining

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17
Q

name the reagents in acid-fast staining
- primary stain
- mordant
- decolorizer
- counterstain

A

primary stain - carbol fuchsin
mordant - heat
decolorizer - acid alcohol
counterstain - methylene blue

!!! study the procedure !!!

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18
Q

the substance behind why some bacteria retain the color of the primary stain (carbol fuchsin) in acid-fast staining

A

mycolic acid - a waxy substance that does not allow the decolorizer to enter into the cell wall due to its
waxy nature

19
Q
  • a special stain for bacterial capsules wherein colorless bacteria are observed against a dark background
  • distortions of cell size and shape are minimized because heat fixing is not necessary and the cells do not pick up the stain
A

negative stain

20
Q
  • a special stain method for endospores wherein the endospore are stained with malachite green; heat is used to provide stain penetration
  • rest of the cell is then decolorized, and counter stained a light red safranin
A

Schaffer-Fulton Method

!!! study method !!!

21
Q

a special stain method where thin cells and structures are thickened with silver on the surface so they become visible under ordinary microscope

e.g., demonstration of spirochetes and bacterial flagella and the sue of FONTANA’S SILVER STAIN

A

impregnation

22
Q

any technique for producing visible images of structures or details too small to be seen by the human eye using a microscope or other magnification too

A

microscopy

23
Q

3 main branches of microscopy

A
  • optical/light - use of any kind of microscope that uses visible light to observe specimens
  • electron - uses electromagnetic lenses, electrons, and a fluorescent screen
    - image captured on photographic film to create an ELECTRON PHOTOMICROGRAPH
    - use 0.005 nm wavelength, 100, 000 SHORTER THAN VISIBLE LIGHT
  • scanning probe microscopy - photographic film to create an electron photomicrograph
    - higher resolution than electron and sample does not need special preparation
24
Q

3 things a microscope should provide

A

magnification
resolution
clarity

25
Q

name the two phases of magnification in compound microscopes

A

objective lens initially forming the REAL IMAGE
ocular lens forming the VIRTUAL IMAGE

26
Q

how to compute for total magnification

A

power objective * power of ocular lens
e.g., 10x LPO * 10x = 100X

27
Q
  • ability of a lens to separate or distinguish two adjacent objects
  • function of wavelength of light and objective lens
A

resolution or resolving power

28
Q

how does wavelength affect the resolution

A

shorter wavelength, better resolution

29
Q

mathematical constant that describes the relative efficiency of a lens in bending light rays

A

numerical aperture (NA)

the higher the NA, the better the resolution

30
Q

differences in intensity between objects or between
an object and its background

A

contrast

31
Q

effect of staining on contrast

A

staining increases contrast

32
Q

5 types of optical/light microscopy

A
  • Bright-field (light) microscopy
  • Dark-field microscopy
  • Phase-contrast microscopy
  • Fluorescent microscopy
  • Confocal microscopy

Boy Doesn’t Put Fucking Condoms

!!! SEE MODULE TO COMPARE IMAGES SEEN !!!

33
Q

most common type of light microscopy that is easiest to use and evenly illuminates the field of view

A

bright-field (light) microscopy e.g., compound light microscope

!!! see module for images

34
Q

type of light microscopy valuable when the specimen that cannot be observed with direct light

however, because the light passes around rather than through organisms, INTERNAL STRUCTURES CANNOT BE STUDIED

A

dark-field microscopy

!!! see module for images

35
Q

type of light microscopy that has one set of light rays come directly from the light source and the other from light that is reflected or diffracted from a particular structure in the specimen

based on the wave nature of light

A

phase-contrast microscopy

!!! see module for images

36
Q

type of light microscopy that use the *ability of substances to absorb short wavelengths of light (ultraviolet) and give off light at a longer wavelength (visible)

specimens are stained with fluorochromes so they will emit, or return light

A

fluorescent microscopy

*flourescence

!!! see methods in module !!!

37
Q

type of light microscopy wherein lenses focus a laser beam to illuminate a given point on one vertical plane of a specimen

uses:
- to obtain three-dimensional images
- to evaluate cellular physiology

A

confocal microscopy

!!! see module for images

38
Q

2 types of electron microscopy

A

transmission electron microscopy (TEM)
scanning electron microscopy (SEM)

39
Q

a type of electron microscopy wherein electrons pass through the specimen and are scattered

magnetic lenses then focus the image onto a fluorescent screen or photographic plate

A

transmission electron microscopy (TEM)

!!! see module for images

40
Q

a type of electron microscopy wherein primary electrons sweep across the specimen and knock electrons from its surface

secondary electrons are picked up by a collector, amplified, and transmitted onto a viewing screen or photographic plate

A

scanning electron microscopy (SEM)

!!! see module for images

41
Q

microscopy that map the bumps and valleys of a surface on an ATOMIC SCALE

A

scanning-probe microscopy

42
Q

2 types of scanning-probe microscopy

A

scanning tunneling microscopy (STM)
atomic force microscopy (AFM)

43
Q

type of scanning-probe microscopy used for detailed viewing of molecules such as DNA

A

scanning tunneling microscopy (STM)

!!! see module for images

44
Q

a type of scanning-probe microscopy that produce 3D images of the SURFACE of a molecule

A

atomic force microscopy (AFM)

!!! see module for images

VMCB 121 Lecture (5 decks)

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