Chapter 2 Flashcards
Aims of the examiner when observing specimen
Observation of:
- overall structure
- identification
- any movement?
- thicker or thinner specimen
Wet mounds
consists of a drop or two of culture placed on a slide and overlaid with a coverslip
Hanging Drop
- a drop of culture is placed in a concave (depression slide)
- Vaseline adhesive or sealant, and coverslip are used to suspend the sample
When using wet mounds or hanging drops to observe specimens, cells are suspended in what substances to maintain viability and to help see them move as they naturally would
water, broth, or saline
Procedure that applies colored chemical dyes to specimens
Staining
Basic dyes have a ______ charge
positive +
Acidic dyes have a _____ charge
negative -
Bacteria have numerous negatively charged substances and ______ basic dyes
attract (because basic dyes are positive)
Cells ____ acidic dyes
repel
Positive stains
dye sticks to specimen and gives it color
Negative stains
Dye does not stick to the specimen but settles some distance from its outer boundary forming a silhouette
; can be used to accentuate a capsule
; Nigrosin/India Ink is a negative stain
Simple stains
only require a single dye and are used for uncomplicated procedures
- cause all cells in the smear to turn more or less the same color
- reveal shape, size, and arrangement
- positive type of staining
- won’t help differentiate between cells
Differential stains
- utilizes two differently colored dyes: a primary dye and counterstain
- distinguish cells types or parts (especially CELL WALL)
- requires additional chemical reagents to produce the desired reaction
- developed in 1884 by Hans Christian Gram: THE GRAM STAIN
- Crystal Positive (primary dye)
- Gram’s Iodine (stabilizer/mordant)
- Alcohol Rinse (wash/cleanse)
- Safranin (counterstain)
The Gram Stain (differential stain)
remains the universal basis for bacterial classification and identification
a practical aid in diagnosing infection and guiding treatment
Differential Stains: Acid-Fast Stain
differentiates acid-fast bacteria (acid resistant - turns pink) from non acid-fast bacteria (non acid resistant - turns blue)
*originated as a method to detect mycobacterium tuberculosis
Differential Stain: The Endospore stain
similar to acid-fast stain in that dye is forced by heat into resistant bodies called endospores
- distinguishes between endospores and vegetative (inactive) cells
- significant in identifying gram-positive, spore-forming members of the genus Bacillus and Clostridium
Special Stains
used to emphasize cell parts that are not revealed by conventional staining methods
Special Stains: Capsular Staining
observes microbial capsules
-negatively stained with India ink
Special Stains: Flagellar staining
used to reveal tiny slender filaments used by bacteria for locomotion
-flagella are enlarged by depositing a coating on the outside of the filament and then staining it
Outcome: List and describe the three elements of good microscopy.
Magnification: occurs in two phases, the real image (slide) and the virtual image (what we see through lens)
-oil immersion total mag: 1000x
Resolution: resolving power; the capacity of an optical system to distinguish two adjacent objects or points from one another
- resolution oh human eye: 0.2mm
- Res. of oil immersion lens: 0.2 micrometers
Contrast: darkness of the background relative to the specimen placed on the stage
-measured in refractive index; the degree of bending that light undergoes as it passes from one medium to another
Outcome:
Differentiate between light and electron microscopy
Light:
- uses light as an illuminating source
- lower resolution
- colored image
Electron:
- used electron beam as illuminating source
- higher resolution
- black and white imaging
Outcome:
Convert among different units of metric system
1 micrometer = 1000 nanometers
1000 micrometers = 1 millimeter
Outcome: Give an example of a SIMPLE stain
A crystal violet stain
A methylene blue stain
Outcome: Give an example of a DIFFERENTIAL stain
Gram stain (gram pos turns purple and gram neg turns pink)
Acid-fast stain
Endospore stain
Outcome: Give an example of a SPECIAL stain
A flagellar stain
A capsular stain with India Ink
Outcome: Explain what the 5 I’s are and what each step entails
- Inoculation
-infection
-introduction of an inoculum (small sample of microbes) to a medium - Incubation
-applying special needs of the microbes to grow
-ie. keeping certain temperatures or introducing different gases like oxygen to help the microbes grow
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