(exam 1) ch 3 microscopy staining

Question 1

what is light microscopy?

Answer 1

microscopes that use visible light to observe specimens

Question 2

what is a compound light microscope?

Answer 2

series of lenses for magnification which uses visible light; (no oil) 4X, 10X, 40X (oil) 100X

Question 3

in a compound microscope the image from the _______ lens is magnified again by the _______ lens.

Answer 3

objective lens; ocular lens

Question 4

total magnification =

Answer 4

object lens x ocular lens “add zero on”

Question 5

what is resolution (or resolving power) ?

Answer 5

ability to distinguish between two points; greater resolution allows finer detail and structure between two points

Question 6

what is refractive index?

Answer 6

measure of light bending ability of a medium; immersion oil has raw sane refractive index as glass and keeps light from refracting (changing direction) which increases the resolution

Question 7

what is staining?

Answer 7

coloring microorganisms with a dye that emphasizes certain structures

Question 8

what must happen before staining microbes?

Answer 8

microbes must be smeared (creating a thin film containing microbes) and fixed (attached with heat) to the glass slide

Question 9

why is fixing microbes important?

Answer 9

1) causes them to stick to the slide so they do not wash off 2) kills them 3) preserves various structures for observation

Question 10

what are three common staining techniques?

Answer 10

1) simple stain 2) differential stain 3) special stain

Question 11

what is the primary stain?

Answer 11

first dye used to color the microbe

Question 12

what is the counter stain?

Answer 12

second dye used after a wash step

Question 13

what is mordant?

Answer 13

substance used to set / bind dye and acts to intensify staining (iodine)

Question 14

what is a simple stain technique?

Answer 14

uses a single dye with a positive charge; *reveals the cells morphology* size, shape, and arrangement of bacterial cells

Question 15

what is differential stains?

Answer 15

*Used to distinguish between different bacterial species* and the differences are based on the difference in bacterial structure; usually require more than one stain

Question 16

what is the most common and most important type of differential stain in medical microbiology?

Answer 16

GRAM STAIN

Question 17

what are the two large groups that a gram stain (differential stain) places bacteria into?

Answer 17

1) gram positive 2) gram negative

Question 18

what is the structure of gram positive bacteria?

Answer 18

bacteria have a thick peptidoglycan cell wall

Question 19

what is the structure of gram negative bacteria?

Answer 19

bacteria have a thin peptidoglycan cell wall and an outer membrane composed of lipopolysaccharides

Question 20

what kind of information does a gram stain provide?

Answer 20

provides valuable information for the treatment of disease; allows for the proper selection of antibiotics; antiobiotics are typically effective against either gram positive or gram negative bacteria BUT NOT BOTH

Question 21

what is the four step procedure of a gram stain?

Answer 21

1) application of crystal violet (purple dye) *primary stain* 2) application of iodine (mordant) *seal- sets crystal violet into gram positive stain* 3) alcohol wash (decolorization) * washes purple off anything gram negative* 4) application of safranin (counter stain) *turns anything gram negative pink*

Question 22

what are the results of a gram stain (differential stain) ?

Answer 22

gram positive bacteria appear PURPLE; they retain crystal violet from the 1st stain

gram negative bacteria appear PINK; alcohol wash removes the crystal violet and bacteria appear pink after the second stain with safranin

Question 23

what is another type of differential stain that was discussed?

Answer 23

acid-fast stain

Question 24

what happens in an acid fast stain?

Answer 24

dye binds to bacteria that have a waxy material in thier cell walls - dye is not decolorized by acid acohol

Question 25

what two bacteria genera (plural of genus) would you use an acid fast stain to identify them?

Answer 25

1) _Mycobacterium_ _2) Nocardia_

Question 26

what are special stains used to distinguish and what are the three types?

Answer 26

used to distinguish specific structures of microorganisms three types 1) capsule stain 2) endospore stain 3) flagella stain

Question 27

what are bacterial capsules?

Answer 27

gelatinious outer covering; not found on all bacteria; they contribute to the ability of bacteria to cause disease (virulence)

Question 28

how do you stain bacteria capsules (special stain)?

Answer 28

bacteria capsules are difficult to stain because they are water souble and dye washes away easily; so in order to stain them we use negative staining = the background is stained instead of the cell and the dark background contrasts w capsules so they appear as a halo around the cell

Question 29

what are endospores?

Answer 29

resistant, dormant structures located inside some cells (protect bacteria from adverse environmental condition)

Question 30

how do you stain endospores (special stain)?

Answer 30

endospores cannot be stained with ordinary methods because dye cannot penetrate endospore wall; procedure is to use a 1) primary stain of malachite green w/ heat 2) decolorize cells = water and 3) counterstain = safranin result = spores appear green within red/pink cells

Question 31

what are flagella?

Answer 31

structures used for locomotion and are too small to be seen with light microscopy

Question 32

how do you stain flagella (special stain)?

Answer 32

use a mordant and carbolfuchsin dye to thicken the appearance of flagella which makes them visible under the light of a microscope