Ch 2

Question 1

Regardless of the type of microscope being used, what 3 elements are always needed to form an image?

Answer 1

1. a source of illumination
2. a specimen to be examined,
3. a system of lenses that focuses the illumination on the specimen and forms the image

Question 2

In a light system what is the source of illumination and system of lenses that focuses the illumination on the specific and forms the image?

Answer 2

• source of illumination: visible light

- system of lenses: series of glass

Question 3

In a light system what is the source of illumination and system of lenses that focuses the illumination on the specific and forms the image?

Answer 3

illumination: beam of electrons emitted by a heated tungsten filament

system of lenses: series of electromagnets.

Question 4

How is the image formed from light microscopy viewed

Answer 4

either viewed directly through an eyepiece or focused on a detector, such as photographic film or an electronic camera.

Question 5

How is the image formed from electron microscopy viewed

Answer 5

electron beam is focused on either a fluorescent screen or photographic film, or it is digitally imaged using a detector

Question 6

What is a wavelength

Answer 6

Distance from crest of 1 wave to crest of next wave

Question 7

What does the rope analogy demonstrate

Answer 7

The ability of an object to perturb a wave’s motion depends crucially on the size of the object in relation to the wavelength of the motion
- wavelength of light source sets limit on how small an object can be seen

Question 8

Do photons or electrons have shorter wavelengths

Answer 8

electrons

Question 9

What is interference

Answer 9

The process by which 2 or more waves combine to reinforce or cancel one another, producing a wave equal to the sum of the 2 combining waves

Question 10

What is diffraction

Answer 10

image that you see when you look at a specimen through a series of lenses is really just a pattern of either additive or canceling interference of the waves that went through the lenses,

Question 11

What two fundamental properties do light and electron microscopy have in common

Answer 11

Focal length and angular aperture

Question 12

What is focal length

Answer 12

distance btwn midline of lens & point at which rays passing thru lens converge to a focus

Question 13

What determines the focal length

Answer 13

determined by the index of refraction of the lens itself, the medium in which it is immersed, and the geometry of the lens.

Question 14

What unit is focal length measured in

Answer 14

meters

Question 15

What unit is the lens magnifying strength measured in

Answer 15

Diopters

Question 16

What is angular aperture

Answer 16

half-angle alpha of the cone of light entering the objective lens of the microscope from the specimen AKA how much of the illumination that leaves the specimen actually passes through the lens

Question 17

What determines sharpness of interference pattern & thus ability of lens to convey info about specimen?

Answer 17

angular aperture

Question 18

in best light microscopes what is the degree of angular aperture

Answer 18

70 degrees

Question 19

What is resolution

Answer 19

minimum distance 2 points can be apart & yet still remain identifiable as separate points when viewed thru microscope

Question 20

the higher the resolution, the ___ the objects that can be distinguished using a particular lens

Answer 20

smaller

Question 21

What 3 factors govern resolution

Answer 21

wavelength of light used to illuminate specimen, angular aperture, refractive index of medium surrounding specimen

Question 22

What is refractive index

Answer 22

measure of the change in the velocity of light as it passes from one medium to an- other.

Question 23

What does the constant 0.61 represent

Answer 23

the degree to which image points can overlap and still be recognized as separate points by an observer

Question 24

What is n sin alpha

Answer 24

numerical aperture

Question 25

What is the Abbe eqn

Answer 25

r = 0.61 lambda/ NA

Question 26

is smaller or larger r better?

Answer 26

smaller

Question 27

Unit for r?

Answer 27

nm

Question 28

The larger the angular aperture the more ___ info the lens can transmit

Answer 28

more

Question 29

What is useful magnification

Answer 29

.

Question 30

Greatest magnification that can be achieved with a light microscope is ____ x the numerical aperture of the lens being used

Answer 30

1000 x

Question 31

Theoretical limit of resolution of electron microscopes

Answer 31

0.002nm

Question 32

What prevents the electron microscope from achieving the theoretical limit of resolution of 0.002nm

Answer 32

electromagnets produce considerable distortion when the angular aperture is more than a few tenths of a degree. aka numerical aperture is smaller than light microscope

Question 33

What is the PRACTICAL limit of resolution of electron microscope

Answer 33

2nm

Question 34

What is a compound microscope

Answer 34

light microscope that uses several lenses in combo

Question 35

What are condenser lenses

Answer 35

direct light toward specimen mounted on glass slide

Question 36

What is stage

Answer 36

Where specimen is placed on

Question 37

what are objective lens

Answer 37

forms primary image

Question 38

what are ocular lens/eyepiece

Answer 38

enlarges primary image

Question 39

what are intermediate lens

Answer 39

positioned btwn objective and ocular lens to further enlarge image

Question 40

how do calculate overall magnification of image

Answer 40

multiply enlarging powers of objective lens, ocular lens, and intermediate lens

Question 41

Characteristics of brightfield microscopy

Answer 41

- specimen must be dead - only specimens possessing colour or other property that affects amount of light that passes thru (or else must be stained)

Question 42

Characteristics of phase-contrast microscopy

Answer 42

- improves contrast w/out sectioning or staining by exploiting differences in thickness & refractive index of various regions of cells being examined - converts phase differences into alterations in brightness

Question 43

What is a phase plate

Answer 43

optical material inserted into the light path above the objective lens.

Question 44

How does DIC (differential interference contrast microscopy diff from phase-contrast microscopy

Answer 44

DIC is more sensitive cause employs special prism to split the illuminating light beam into 2 separate rays. Then 2 beams recombined, any changes that occurred in the phase of one beam as it passed through the specimen cause it to interfere with the second beam.

Question 45

optical components required for DIC microscopy

Answer 45

PAP - polarizer, an analyzer, and a pair of Wollaston prisms.

Question 46

How does Hoffman modulation contrast increase contrast

Answer 46

increases contrast by detecting optical gradients across a transparent specimen using special filters and a rotating polarizer

Question 47

What is fluorescence

Answer 47

process that begins with the absorption of light by a molecule and ends with emission of light with a longer wavelength

Question 48

emitted photon is always of ___ energy (____ wavelength) than the original photon that was absorbed.

Answer 48

less energy, long wavelength

Question 49

Characteristics of fluorescence microscopy

Answer 49

- light microscopy that employs light to excite fluorescence in specimen

Question 50

What is a excitation filter

Answer 50

placed btwn the light source & rest of light path that transmits only light of a particular wavelength

Question 51

what is a dichroic mirror

Answer 51

reflects light below certain wavelength & transmits lights above certain wavelength, deflects incoming light toward objective lens which focuses light onto specimen

Question 52

What is immunostaining

Answer 52

based on the ability of antibodies to recognize and bind to specific molecules (antigens)