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Bacte LAB > STAINING TECHNIQUES > Flashcards

STAINING TECHNIQUES Flashcards

1
Q

-> organic or aqueous preparations of dyes that impart a variety of color to tissue or microorganisms

• Differentiate or identify structures and characteristics of bacteria

A

Dyes or stains

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2
Q
  • ion that imparts the color
A

Chromophore

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3
Q

The chromophore is a positive ion -›____
• methylene blue, crystal violet, carbol fuchsin, malachite green, safranin

A

BASIC STAIN

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4
Q
  • The chromophore is a negative ion -› _______
    eosin, nigrosin, picric acid
A

ACIDIC STAIN

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5
Q

• -> ion not giving off the color

A

Counterion

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6
Q

• directly stains bacteria and bacterial structure
bacteria absorbs the stain

A

DIRECT STAINING

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7
Q

• No special structures are seen
Stain evenly dyes the bacteria
The stain is flooded across the smear

A

Simple Staining

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8
Q

• Only one type of dye is used
• No special structure of a cell is distinguished
• eg. methylene blue

A

Simple Staining

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9
Q

• Can visualize bacterial structures
• Can isolate specific parts of bacterial structure

A

Selective/Special Staining

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10
Q

-> Not easily colorized, not easily decolorized
-> Impenetrable

A

Endospore Stain or Schaeffer-Fulton stain

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11
Q

Differential Staining
• Two Types:

A

Gram Stain
Acid Fast Stain

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12
Q

-> The first stain used in the procedure
-> A stain that will impart the color

A

Initial/Primary Stain

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13
Q

•
-> allow stains to stick to the bacteria
- for a bridge between cell and stain

A

Mordant

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14
Q

-› no chemical eg. heat or cold
-› ferrous sulfate, iodine, tannic acid

A

Physical Mordant

Chemical Mordant

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15
Q

-> Removes color
-> Bacteria in question retain the color
-> Bacteria without the specific structure or composition in them will not retain the color

A

Decolorizer

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16
Q
  • Bacteria that were decolorized will absorb the color
A

Counterstain/Secondary Stain

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17
Q

• Negative/Relief Staining
• The background is stained instead for contrast
- Demonstrate the HALO EFFECT
• DRY INDIA INK/NIGROSINE STAIN

A

INDIRECT STAINING

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18
Q

TYPES OF IONIZABLE DYES USED IN STAINING BACTERIA

A

Basic dyes
Acidic dyes

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19
Q

▪ Commonly used in bacteriology
▪ These are cationic dyes with positively charged groups that adhere to negatively charged molecules.

A

BASIC DYES

positive

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20
Q

Basic dyes

A

Includes:
METHYLENE BLUE
MALACHITE GREEN

BASIC FUCHSIN
CRYSTAL VIOLET
SAFRANIN

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21
Q

▪ These are anionic dyes with negatively charged group that bind to positively charged cell structures

A

ACIDIC DYES

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22
Q

ACIDIC DYES

A

EOSIN
ACID FUCHSIN
ROSE BENGAL

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23
Q

▪ Example: METHYLENE BLUE, IODINE

A

SIMPLE STAINING

24
Q

▪ Directed towards the forms and shapes of the cells

A

SIMPLE STAINING

25
Q

▪ Basic dyes have greater affinity for nuclei; Acid dyes have greater affinity for cytoplasm

A

SIMPLE STAINING

26
Q

▪ Application of single staining solution

A

SIMPLE STAINING

27
Q

▪ Example: GRAM STAINING, ACID FAST STAINING

A

DIFFERENTIAL STAINING

28
Q

▪ Makes use of primary stain, mordant, decolorizer and counterstain

A

DIFFERENTIAL STAINING

29
Q

▪ Application of 2 or more stains

A

DIFFERENTIAL STAINING

30
Q

▪ Example: INDIA INK, NIGROSIN STAIN

A

▪ NEGATIVE STAINING

31
Q

▪ Results the bacteria appearing as light colored bodies against the dark background

A

▪ NEGATIVE STAINING

32
Q

GRAM STAINING

▪ Named after________ (Danish Scientist), worked with______ in City hospital morgue in Berlin.

A

HANS CHRISTIAN GRAM

CARL FRIEDLANDER

33
Q

▪ PRINCIPAL STAIN in the laboratory for microscopic examination of bacteria

A

Gram stain

34
Q

▪ Divides bacteria into two large groups: GRAM POSITIVE and GRAM NEGATIVE.

A

Gram stain

35
Q

Gram staining

PRINCIPLE:

Based on the differences in composition between gram positive cell walls which contain thick_____ with NUMEROUS_______ and gram negative cell walls which consist of______.

A

peptidoglycan; TEICHOICE ACID CROSS-LINKAGES

THINNER LAYER OF PEPTIDOGLYCAN

36
Q

If the______ is rinsed too vigorously prior to the application of iodine, it will not be retained and will leave the gram negative bacteria unstained

A

crystal violet dye

37
Q

If the decolorization is prolonged, the ———-will be removed and the gram positive bacteria will not be stained.

A

Gram – positive complex

38
Q

If the decolorization is insufficient, the organism may_______

A

falsely appear as gram positive cells.

39
Q

If the______ is left applied for more than a minute, the ________will be eliminated from the previously stained cells

A

safranin dye

Gram positive complex

40
Q

Failure to leave the safranin dye for a sufficient time will result in ______and background materials.

A

unstained gram negative bacteria

41
Q

All cocci are gram – positive except for

A

Neisseria

Moraxella

Veilonella

42
Q

All bacilli are gram – negative except for

A

Actinomadura, Arcanobacterium, Bacillus, Clostridium, Corynebacterium, Erysipelothrix, Gordonia, Kuthria, Listeria, Mycobacterium, Nocardia, Rhodococcus, Streptomyces, Tropheryma whipplei, and Tsukamurella.

All Awesome Biologists Collect Cool Exotic Germs Knowing Lab Microbes Need Real Scientific Testing and Training.

Actinomadura
Arcanobacterium
Bacillus
Clostridium
Corynebacterium
Erysipelothrix
Gordonia
Kuthria
Listeria
Mycobacterium
Nocardia
Rhodococcus
Streptomyces
Tropheryma whipplei
Tsukamurella

43
Q

REASONS WHY GRAM – POSITIVE BACTERIA BECOME GRAM NEGATIVE BACTERIA

A
  1. Removal of MgRNA
  2. Aged, dying, and autolyzing cells
  3. By using acidic iodine during staining
  4. Due to technical error or the wrong use of stains
44
Q

EXCEPTIONS IN GRAM STAINING

A
  1. Organisms that exist almost exclusively within host cells (Chlamydia)
  2. Organisms that lack cells walls (Mycoplasma and Ureaplasma)
  3. Organisms with insufficient dimension to be resolved by light microscope (Spirochetes)
45
Q

Acid fast

▪ Heat is applied as a mordant in the ______while tergitol is used in the______

A

Ziehl Neelsen Method

Kinyoun Method

46
Q

▪ In this procedure, the cell wall of acid – fast bacteria resists the acid –
alcohol (hydrochloric acid – ethanol mixture) decolorization step.

A

ACID FAST STAINING

47
Q

▪ It utilizes carbolfuchsin as the primary stain and methylene blue or malachite green as the secondary stain.

A

ACID FAST STAINING

48
Q

▪ It is used to stain bacteria that have high lipid contents in their cell walls.

A

ACID FAST STAINING

49
Q

Acid fast

▪ It utilizes______ as the primary stain and _____ or _____ as the secondary stain.

A

carbolfuchsin

methylene blue or malachite green

50
Q

Acid fast principle

A

PRINCIPLE:

• The primary stains binds to mycolic acid in the cell walls of acid fast bacteria, and is retained after decolorizing with acid alcohol.

• Acid Fast Bacilli (AFB) retain the primary stain and deep-pink or red-colored while non – AFB are either blue or green colored.

51
Q

– Hot staining method

A

Ziehl Neelsen

52
Q

– Cold staining method

A

Kinyoun

53
Q

– differentiates Mycobacterium smegmatis from Mycobacterium tuberculosis.

A

Pappenheim Method

54
Q

– differentiates Mycobacterium leprae from Mycobacterium tuberculosis.

A

Baumgarted Method

55
Q

– selective for the cell wall of AFB.

A

Auramine – rhodamine Method

56
Q

WAYS TO FACILITATE ACID – FAST STAINING

A
  1. The use of heating or steaming process for five to seven minutes to temporarily remove the mycolic acid, while the smear is flooded with stain.
  2. By increasing the concentration of dye and phenol in the staining reagent.
  3. A prolonged contact of the specimen with the primary stain.
  4. The addition of wetting agent like tergitol

Bacte LAB (15 decks)

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