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BIOL 2330 - microbiology > preparation + staining > Flashcards

preparation + staining Flashcards

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1
Q

why do we stain bacteria

A

bacteria are colorless, transparent, and difficult to see

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2
Q

what is staining

A

coloring the microorganism with a dye for visualization

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3
Q

what can we see via staining

A

certain structures within the organism or the organism itself

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4
Q

what must we do to a microbe before staining it (1 word)

A

we must fix it

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5
Q

what is fixing

A

attaching the microbe to the slide, killing it, and preserving internal and external structures in their natural state

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6
Q

how is fixing done

A

a smear is made, the slide is air dried, and it is fixed by passing it through a flame several times

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7
Q

define smear

A

a thin film of bacteria on the surface of the slide

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8
Q

what are two ways we can make a smear

A

plate or broth

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9
Q

describe a plate smear

A

a drop of water is put onto the slide first, and then an inoculating loop is used to get the cells, then the cells are spread into the water

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10
Q

describe a broth smear

A

put inoculating loop into a broth, and then smear it onto the slide

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11
Q

describe the orientation of the slide when fixing it through a flame

A

smear side up

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12
Q

what two methods can be used for fixing microorganisms

A

heat fixation and chemical fixation

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13
Q

describe heat fixation

A

preparing the slide then passing it through a flame

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14
Q

describe chemical fixation

A

a fixative is added to the smear

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15
Q

in chemical fixation, give examples of fixative that can be added to the smear

A

ethanol, methanol, acetic acid, formaldehyde

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16
Q

which method of fixation (heat or chemical) is better for preserving the internal morphology of cells

A

chemical

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17
Q

describe the features of the dyes used to stain organisms

A

they’re ionic, and they have chromophore groups attached

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18
Q

what do chromophore groups do

A

give the dye its color

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19
Q

what two classes can ionizable dyes be divided into

A

basic and acidic dyes

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20
Q

what type of charged groups do basic dyes have

A

positive

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21
Q

what types of molecules do basic dyes bind do? give examples

A

negatively charged molecules

nucleic acids, many proteins

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22
Q

which type of dye (basic or acidic) is most used in microbiology? why?

A

basic; because they’re attracted to negative charges and bacterial cell surfaces are negatively charged

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23
Q

give examples of basic dyes

A

methylene blue, malachite green, crystal violet, safranin

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24
Q

what type of charged groups do acidic dyes have

A

negative

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25
Q

what types of molecules do acidic dyes bind to? give examples

A

positively charged things

some proteins

26
Q

describe how acidic dyes interact with bacteria

A

they repel bacteria due to having the same charge

27
Q

give examples of acidic dyes

A

eosin, acid fuchsin, congo red

28
Q

what are the two types of staining procedures

A

simple staining and differential staining

29
Q

what is simple staining

A

microbes are stained with a single agent (1 color)

30
Q

describe the steps of simple staining

A

prepare smear, air dry, fix the bacteria, apply the dye, rinse off excess, blot dry, examine

31
Q

what are the uses of simple staining

A

used to highlight the entire microbe, allows one to get info about the size, shape, and arrangement of cells

32
Q

what is differential staining

A

used to divide bacteria into separate groups based on their staining properties

33
Q

what are two types of differential staining

A

gram stain and acid fast stain

34
Q

what is a gram stain

A

divides bacteria into two groups based on their ability to take up and retain a dye

35
Q

what two groups are made in a gram stain

A

gram positive and gram negative

36
Q

what type of smear is used in a gram stain; heat or chemical

A

heat fixed

37
Q

describe the steps of a gram stain

A

apply crystal violet dye, wash, then apply iodine = everything purple

rinse, use ethanol for decolorization (gram negative will be colorless)

wash, apply safranin (counterstain), turns colorless into pink

38
Q

what dyes are used in a gram stain

A

crystal violet and safranin

39
Q

what are the results of the gram stain

A

gram negative = red

gram positive = blue

40
Q

what is an acid stain used for

A

used to stain a small group of organisms that do not readily take up stains + differentiates between 2 groups of bacteria

41
Q

what two groups of bacteria are defined in an acid fast stain

A

acid fast vs non acid fast

42
Q

define acid fast organism

A

organisms that do not readily take up stains due to high content of lipid in their cell walls

43
Q

what type of smear is used in an acid fast stain: chemical or heat fixed

A

heat fixed

44
Q

describe the steps of an acid fast stain

A

add primary stain (carbofuchsin) while slide is over steaming water

cool + rinse, add decolorizer then acid alcohol

wash, then apply counterstain (methylene blue)

45
Q

in an acid fast stain, why do we apply the primary stain while the slide is over steaming water

A

the heat is used to push the dye through the cell wall since acid fast organisms dont readily take up dye

46
Q

what two dyes are used in an acid fast stain

A

carbofuchsin red and methylene blue

47
Q

what are the results of the acid fast stain

A

acid fast = red

non acid fast = blue

48
Q

define endospore

A

a resistant structure that allows some bacteria to withstand unfavourable conditions

49
Q

what is the name for the procedure to stain an endospore

A

schaeffer-fulton procedure

50
Q

describe how we stain an endospore

A

using a heat fixed smear, add malachite green in the presence of steam (this will penetrate the endospore)

cool + rinse, add safranin as the counterstain, wash

51
Q

what are the visual results of the endospore stain

A

endospore=green

cytoplasm=red

52
Q

what does the endospore stain allow us to see

A

presence of an endospore + location of the endospore

53
Q

what are flagella used for

A

locomotion of bacterial cells

54
Q

what do we do for a flagella stain

A

add carbolfuchsin

55
Q

why would we want to do a flagella stain

A

aids in identifying bacteria based on presence and location of the flagella

56
Q

what is a negative stain

A

its when we stain the background instead of the organism

57
Q

what color will bacteria be after a negative stain

A

colorless; only the background is stained

58
Q

what dye is used in a negative stain

A

an acidic dye, ie eosin

59
Q

what is a negative stain used for

A

revealing the presence of a capsule

60
Q

what is a capsule

A

a substance that may surround some bacterial cells

61
Q

what is the charge on a capsule

A

negative

BIOL 2330 - microbiology (30 decks)

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