Module 2

Question 1

What is the goal of a microscope?

Answer 1

to accurately enlarge or magnify the image of an object

Question 2

What is light refraction?

Answer 2

when light passes from one medium to another, it is refracted or bent, at the interface

Question 3

What is the refractive index?

Answer 3

the measure of how much a substance slows the velocity of light

Question 4

what focal length will magnify an object more?

Answer 4

a lens with a shorter focal length

Question 5

what focal length will magnify an object less?

Answer 5

a lens with a longer focal length

Question 6

What is microscope resolution?

Answer 6

the ability of a lens to separate or distinguish small objects that are close together

Question 7

What is the objective lens?

Answer 7

the lens closest to the specimen

Question 8

What is the Abbe Equation?

Answer 8

d = 0.5λ/nsinΘ

Question 9

what is the d in the Abbe Equation?

Answer 9

min. distance between 2 objects to distinguish them

Question 10

what is the nsinΘ in the Abbe Equation?

Answer 10

numerical aperture (NA, ability to gather light)
n = refractive index of medium
Θ = amount of light entering the lens

Question 11

How can we get the best resolution using Abbe’s Equation?

Answer 11

want the smallest d
- small λ
- large nsinΘ

Question 12

What is a light microscope?

Answer 12

compound microscopes, where the image is enlarged by the action of 2 or more lenses

Question 13

what are the 2 parts of the 2 lens system

Answer 13

1. Objective
2. Eye-piece

Question 14

Explain the Objective part of the 2 lens system

Answer 14

• nearer the specimen
• magnifies the specimen
• shorter the focal length, shorter the working distance

Question 15

Explain the Eye-piece part of the 2 lens system.

Answer 15

• eye-piece is the ocular
• produces virtual image seen by the eye

Question 16

How do you find the Total Magnification using a light microscope?

Answer 16

ocular magnification * objective magnification

Question 17

What is fixing in sample preparation?

Answer 17

fixing preserves internal and external structures (organisms killed and firmly attached to microscope slides)

Question 18

What are the two types of fixation in sample preparation?

Answer 18

1. heat fixation
2. chemical fixation

Question 19

When is heat fixation used?

Answer 19

routinely used with prokaryotes; preserves overall morphology but not internal structures

Question 20

When is chemical fixation used?

Answer 20

used with larger, more delicate organisms; protects fine cellular structures + morphology

Question 21

What is staining in sample preparation?

Answer 21

• staining increases visibility and accentuates specific morphological features

Question 22

What are the two common features of dyes in staining?

Answer 22

1. Chromophore groups (chemical groups with conjugated double bonds that give color)
2. Ability to bind with cells (ionic, covalent, hydrophobic bonding)

Question 23

Explain simple staining

Answer 23

• a single stain is used
• useful for determining size shape and arrangement of cells
• most common with binding by ionic interactions

Question 24

What do basic dyes bind to?

Answer 24

negatively charged molecules (DNA, proteins, cell surfaces)

Question 25

What do acidic dyes bind to?

Answer 25

positively charged molecules

Question 26

What is differential staining?

Answer 26

differential staining is used to differentiate organisms based on their staining properties

Question 27

What type of staining is gram staining?

Answer 27

differential staining

Question 28

What is gram staining?

Answer 28

the most widely used differential stain that differentiates based on differences in the cell wall structure

Question 29

Gram (+) gives a _______ colour

Answer 29

dark purple

Question 30

Gram (-) gives a ________ colour

Answer 30

pink/red

Question 31

What is a brightfield microscope?

Answer 31

produces a dark image against a bright background

Question 32

What can a brightfield microscope observe?

Answer 32

- viewing organizations of microorganisms - stained bacterial cells (ex. gram staining)

Question 33

What does a darkfield microscope show?

Answer 33

produces a bright image against a dark background

Question 34

What is a darkfield microscope used for?

Answer 34

- used to observe living, unstained preparations (bacteria, algae) - used to observe internal structures in eukaryotes - thin, delicate samples

Question 35

What is a phase contrast microscope?

Answer 35

enhances contrast in transparent/colorless samples by amplifying small differences in the refractive index of the sample

Question 36

What can a phase contrast microscope observe?

Answer 36

- excellent way to observe living cells (esp. eukaryotes) - can view cell division (mitosis and meiosis) without needing to kill or stain

Question 37

What is a fluorescence microscope?

Answer 37

microscope that uses fluorescent dyes (fluorochromes) to label specific structures in a sample

Question 38

What can a fluorescence microscope observe?

Answer 38

- protein localization - tracking live cells over time - studying cell organelles

Question 39

How are images produced in electron microscopy?

Answer 39

by beams of electrons

Question 40

What is different about the wavelength in electron beam vs light?

Answer 40

the wavelength is much shorter with the electron beam

Question 41

What are the advantages to a shorter wavelength in electron microscopy?

Answer 41

- significantly higher resolution - useful for different studies of microorganisms

Question 42

What is a TEM?

Answer 42

transmission electron microscopy: - the electrons that don't scatter when passing through a specimen are transmitted and used to produce an image

Question 43

What happens in the denser regions of a TEM?

Answer 43

the denser regions scatter more electrons and appear darker

Question 44

What can a TEM observe?

Answer 44

produces images of internal structures

Question 45

What is a SEM?

Answer 45

Scanning Electron Microscopy: - uses a focused beam of electrons reflected from the surface to create an image

Question 46

What can a SEM observe?

Answer 46

produces a 3-D image of the surface of a cell/sample

Question 47

What must be done to the sample before observing in an electron microscope?

Answer 47

samples must be fixed, dried and coated in metal (therefore, they must be dead)

Question 48

What is a confocal microscope?

Answer 48

scanning laser microscope that creates high resolution, 3-D images of the species

Question 49

What can a confocal microscope observe?

Answer 49

- live, 3-D cell imaging is possible

Question 50

What study is a confocal microscope used for?

Answer 50

useful for the study of biofilms.

Question 51

What are the two types of scanning probe microscopy?

Answer 51

1. Scanning tunneling microscope 2. Atomic force microscope

Question 52

What is Scanning tunneling microscope?

Answer 52

sharp probe moves over the surface of a specimen at a constant current to create an image

Question 53

What does a scanning tunneling microscope show?

Answer 53

allows scientists to view atoms on the surface of a solid

Question 54

What is an atomic force microscope?

Answer 54

sharp probe moves over surface of specimen at a constant distance to produce an image

Question 55

What does an atomic force microscope show?

Answer 55

used to study the interaction of pr-, to follow the behavior of living bacteria + other cell, and to visualize membrane pr- such as aquaporins

Question 56

What do Scanning Probe Microscopes use to create images of samples?

Answer 56

they use mechanical interactions between the probe and sample surface to gather info