Chapter 3

Question 1

What is the path of light through a compound microscope?

Answer 1

Light from the illuminator passes through the condenser, from there to the objective lenses, and finally the ocular lenses.

Question 2

Describe Dark Field Microscopy

Answer 2

It is used to examine live organisms that cannot be stained. Uses a dark condenser that contains an opaque disk. Specimen looks light against a dark background. Frequently used to examine unstained microorganisms suspended in liquid. Good for examining very thin spirochetes such as Treponema palladum (syphilis). A disadvantage is that it is not very useful for examining small organisms.

Question 3

Describe Phase Contrast microscopy

Answer 3

PERMITS DETAILED EXAMINATION OF INTERNAL STRUCTURES in living microorganisms. It is not necessary to fix or stain. Two sets of light rays combine to form an image of the specimen in gray scale.

Question 4

Describe Fluorescence Microscopy

Answer 4

Takes advantage of substances’ ability to absorb short wavelength (UV) light and give off longer, visible wavelength light. If substances do not naturally fluoresce, they are treated with fluorochromes. This type of microscopy is used TO VIEW ANTIBODY SPECIFICITY.

Question 5

Describe Bright Field Microscopy

Answer 5

Under normal conditions, in a compound light microscope

Question 6

Explain how electron microscopy is different from light microscopy.

Answer 6

Electron microscopy uses electrons instead of light which provides for shorter wavelengths and is focused with magnets. Substances have to be stained in hard metals and the sample has to be in a vacuum and very cold. Provides 3D images. Very high resolution, but specimens are killed in the process.

Question 7

What is a simple stain?

Answer 7

An aqueous or alcohol solution of a single basic dye. The primary purpose of a simple stain is to highlight the entire microorganism so that cellular shapes and basic structures are visible.

Question 8

What are the two types of dyes?

Answer 8

Basic and acid dyes

Question 9

What are the basic dyes?

Answer 9

crystal violet, methylene blue, malachite green, and safranin

Question 10

What are the acid dyes?

Answer 10

eosin, acid fuschin, nigrosin

Question 11

What is the difference between acid and basic dyes?

Answer 11

The color of basic dyes is in the positive ion, and the color of acid dyes is in the negative ion.

Question 12

How do differential stains work?

Answer 12

Differential stains react differently with different kinds of bacteria. This allows for them to be used to distinguish between different bacteria. Examples are gram and acid fast stains.

Question 13

What are some common special stains?

Answer 13

Negative stains (to identify capsules), gram stains, and endospore stains.

Question 14

Describe negative stains in reference to capsules.

Answer 14

With a negative stain, only the background is colored, and the capsules appear as unstained halos around bacterial cells, and stand out against the contrasting background.

Question 15

Describe endospore stains.

Answer 15

They are used to detect endospores in bacteria. Malachite green is applied to a heat fixed smear, the stain penetrates the endospore. When safranin is applied, it stains the rest of the cell red.

Question 16

What makes Gram Stains work?

Answer 16

Different kinds of bacteria react differently to gram stains due to structural differences in their cell walls. These differences affect the retention or escape of the crystal violet-iodine combination known as CV-I. Gram+ have a thicker peptidoglycan cell wall than Gram- which have lipopolysaccharide as part of their cell wall. Crystal violet and iodine readily enter both types and combine to form CV-I. This complex is larger than what originally entered the cells, and because of its size, cannot be washed out of the intact peptidoglycan layer of gram + cells, retaining the crystal violet dye. In gram- cells, alcohol wash disrupts the outer lipopolysaccharide layer and CV-I is washed out through the thin layer of peptidoglycan, which is why gram- cells are colorless until counterstained with safranin.

Question 17

Misc Gram facts :)

Answer 17

Gram+ tend to be killed easily by penicillins and cephalosporins. Gram- are generally more resistant because the antibiotics cannot penetrate the lipopolysaccharide layer.