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60 Cards in this Set

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Define Light Microscopy(e)
microscope that uses light to observe specimens
What's another name for compound light microscope?
brightfield microscope
Condenser (lenses)
focuses light through specimen
Diaphragm
controls the amount of light entering the condenser
Ocular lens
eyepiece - remagnifies the image formed by the objective lens
objective lenses
primary lenses that magnify the specimen
Define Resolution
The ability of lenses to distinguish between two points at a specific distance apart.
What is the maximum magnification of a compound light / brightfield microscope?
2000 X
Why use immersion oil?
Without immersion oil, most light is refracted and lost. Has the same refractive index of glass microscope slide.
used for examining live microorganisms that do not stain easily, or are distorted by staining, or are invisible in brightfield microscopy.
Darkfield microscopy
In a Darkfield microscope, a specimen appears ___________ against a _____________ background.
light, dark
A ____________ microscope uses an opaque disc that blocks light from entering objective directly
Darkfield
What is Flourescence microscopy? Brief answer.
uses an ultraviolet source of illumination that causes fluorescent compounds to emit light.
What is the name of the fluorescent dye used in Florescence microscopy
fluorochromes
Describe fluorescent-antibody technique?
1. Combine a fluorochrome to a specific antibody for a specific antigen.

2. When the antibody combines with the specific antigen, the fluorochrome fluoresces
_____________ microscopy Is useful in diagnosing syphilis and rabies or detecting specific cell types or tissues.
Fluorescence
___________ microscopy uses a beam of electrons instead of light.
Electron
What are the 2 types of Electron microscopy?
1) Transmission electron microscopy (TEM)

2) Scanning electron microscopy (SEM)
Define and describe the term: TEM
Transmission Electron Microscopy

- magnification from 10,000 X - 100,000 X

- not in 3D, very thin specimen must be used
Why does Transmission Electron Microscopy (TEM) open the possibility of artifacts?
-Specimens must be fixed, dehydrated, and viewed under vacuum therefore shrinkage and distortion may occur
Define and describe the term SEM:
Scanning Electron Microscope

- 3 Dimensional image
- can study surface of cells and viruses
-1,000 X to 10,000 X
What is the purpose of staining?
Staining microorganism with a dye makes them more visible for study under a microscope.
Prior to staining, organism must be ___________ to microscope slide.
fixed
Explain how to fix an organism
smear of microorganism is air dried then heat fixed by passing the slide quickly above an open flame a few times.
__________ are salts composed of a positive and negative ion.
Stains
What are the 2 types of dyes for staining?
Basic dyes (positive colored ion)
& Acidic dyes (negative colored ion)
Bacteria are slightly ______________ charged at pH 7.0
negatively
Basic dyes are attracted to the ________ charged bacteria.
negatively
Name 3 basic dyes and their color.
Crystal violet (purple)
Methylene blue (blue)
safranin (pink)
Carbolfuchsin (red)
Acidic dyes are...describe 3 brief points.
- not attracted to most bacteria (bacteria also negatively charged)
- stains the background
- valuable in observing cell shapes, sizes, capsules
In ____________ staining, bacteria are colorless against a stained background.
negative
Name 3 acidic dyes
eosin (pink)
nigrosin (dark purple)
india ink (purple)
Simple Stain
- An aqueous or alcohol solution of a single basic dye

- used to highlight the entire microorganism so that cellular shapes and structures are visible.
What is a mordant? and how is it used in Simple staining technique?
Chemical added to solution to intensify the stain. Used to coat a structure like the flagellum to make it thicker and easier to see.
Why doesn't a negative stain color bacteria?
Because it's an acidic dye that is negatively charged. Bacteria is also slightly negative so it repells the stain.
Name 4 simple (basic) stains and their color.
1) methylene blue (blue)
2) carbolfuchsin (red)
3) crystal violet (purple)
4) safranin (pink)
Differential Stains...
...react differently with different bacteria. Can be used to distinguish them
What are the two types of Differential Stains?
1) Gram Stain
2) Acid Fast Stain
Short description of Gram Stain:
classifies bacteria into 2 large groups:

a) gram positive (+)
b) gram negative (-)
Long description of Gram Stain:
1) heat fixed smear is covered with purple die.

2) purple dye is washed off, then smear covered with iodine, a mordant. Bacteria appear dark violet / purple.

3) Slide is washed with alcohol solution which is a decolorizing agent. Removes purple from some species but not others.

4) Alcohol rinsed off and slide stained with safrin (red dye). Washed again and blotted dry.

Bacteria that retain purple / violet color are gram-positive.

bacteria that loose dark violet or purple are called gram-negative.
What drugs easily kill gram (+) bacteria?
penicillin and sulfonamide
What drugs easily kill gram negative bacteria?
Tetracycline and drugs that interfere with protein synthesis.
What are the 4 liquids used in Gram stain procedure in order?
1) Crystal Violet (purple)
2) Iodine (mordant)
3) Alcohol wash (decolorization)
4) Safranin (counterstain)
Gram positive organisms have ______ membrane covering peptidoglycan wall
no
Gram negative organisms differ from Gram positive in that the outer membrane covers the _______________.
peptidoglycan layer
What stain binds to bacteria that have a waxy material in their cell walls and is used to identify all bacteria in the genus Mycobacterium?
Acid-Fast Stain
(carbolfuchsin) RED
Acid-Fast Stain is used to identify all bacteria in the ________ ___________.
genus Mycobacterium
What two well known diseases can be detected by Acid-Fast Stain?
Mycobacterium tuberculosis
Mycobacterium leprae
Special Stains are used for?
- used to color and isolate specific parts of microbes such as flagella, endospores, and capsules.
What is a capsule?
a gelatinous covering of some microbes; confers virulence.
What makes staining capsules difficult?
capsules are soluble in water and may be removed during washing.
Capsule Staining
1) Mix bacteria with Nigrosine, (dark purple background)

2) spread dye as in negative staining

3) stain slide with crystal violet to stain bacteria (purple)

4) Capsules will appear as colorless halos surrounding purple bacterial cells against a dark purple background.
Resistant, dormant structures within a cell which protects the microbe from adverse environmental conditions.
Endospore
Do endospores stain with ordinary dyes? Explain.
No because ordinary dyes do not penetrate the walls of the endospore.
What are the 2 staining methods used for Endospores?
A) Schaeffer-Fulton endospore stain

B) Dorner endospore stain
Schaeffer-Fulton Endospore Stain

Describe the steps.
a) Apply malachite green to heat-fix smear on slide and steam heat for 5 minutes. Spores stain green, cells will be colorless

b) Wash

c) Counterstain with safranin, which will stain the cells pink. Spores will be green.

d) Wash
Dorner Endospore Stain:

Describe the steps
a) put carbolfuchsin, red, in a test tube

b) Add several loopfuls of organism

c) Boil in a beaker of water for 10 minutes. Spores will be red, cells colorless.

d) Add several loopfuls of the carbolfuchsin-bacteria mixture to a drop of nigrosine on a slide.

e) Smear mixture
- bacterial cells will be colorless against dark purple background (Negative Staining)
-Endospores inside the cells will be red.
Describe flagella staining
Staining flagella involves using a mordant and carbolfuchsin (red) to build up the diameters of the flagella until they become visible under the light microscope.

Tedious and delicate staining procedure.
Which method of Endospore staining uses Negative Staining?
The Dorner Endospore Stain
What method of staining would you use to observe capsules?
Negative Staining