Light microscopy (lecture 2)

Question 1

Structures in which we can see with a light microscope?

Answer 1

frog egg 
eu cells 
nucleus 
most bacteria 
mito

Question 2

Resolution is determined by?

Answer 2

wavelength of radiation source used for illumination

Question 3

Size in which is able to be seen with the naked eye (good eye sight)

Answer 3

0.2mm

Question 4

Magnify an image what is needed?

Answer 4

Magnifier or lens

Question 5

Cornea?

Answer 5

focus light onto the retina

Question 6

Magnification?

Answer 6

Magnification: the ratio of the size of the image to that of the object.

Question 7

Resolution?

Answer 7

Resolution: the clarity of the image; illumination and quality of the optics.

Question 8

Contrast?

Answer 8

Contrast: the contrast between the lightest and darkest areas of the sample.

Question 9

How to calculate total magnification?

Answer 9

Magnification of the objective lens and multiple by the eyepiece

Question 10

Robert hooke? Time, what he discovered and how

Answer 10

17th century
- the cell
cork sections - tree

Question 11

Antony van leeuwenhoek?

Answer 11

Collaborated with Hooke in which he improved the production of the lenses and observed different cell types (eg red blood cells)

Question 12

Specimen holder?

Answer 12

Now known as a stage

Question 13

In the 17th century, a lens was?

Answer 13

water flask

Question 14

Zeiss?

Answer 14

1866
lens better
less diffraction
greater level of detail (improved the resolution)

Question 15

Prism allows

Answer 15

to sit at a desk whilst observing structures under a microscope

Question 16

If we adjust the condenser lens?

Answer 16

locus light onto a specimen

Question 17

Objective lens

Answer 17

collects cone of light rays to create an image

Question 18

5 different types of light microscopy?

Answer 18

1) Bright Field
2) Phase contrast
3) Differentail inference contrast (DIC)
4) Fluorescence
5) Confocal

Question 19

Bright field miscopy image type?

Answer 19

Poor images
not much contrast
a lot of light passes through cells

Question 20

Phase contrast, (high contrast due to?)

Answer 20

The diffracted and un-diffracted rays give rise to a change in brightness.

Question 21

Can we see mitotic divisions occurring in phase contrast microscopy?

Answer 21

yes

Question 22

Light first moves though _____ before passes onto the specimen phase contrast microscopy?

Answer 22

Condenser

Question 23

From the light source, the light then passes through the _________ and creates?

Answer 23

Annulgar ring&raquo_space; creates a hollow cone of light

Question 24

From the angular ring, light passes through ______(phase contrast microscopy?)

Answer 24

the specimen

Question 25

Diffraction depends on (phase contrast microscopy?)

Answer 25

Density of the specimen

Question 26

Where is light collected (phase contrast microscopy?)

Answer 26

Phase ring

Question 27

How is a greater detail achieved (phase contrast microscopy?)

Answer 27

Change in amplitude

Question 28

DIC stands for?

Answer 28

Differentail inference contrast

Question 29

DIC uses?

Answer 29

Polarised light

Question 30

What kind of prism is required and purpose? (DIC)

Answer 30

Beam spliter prism Generating two beams of light which travel close to eachother

Question 31

If one of the beams is diffracted by the specimen (DIC)??

Answer 31

when they are re-combined by the top Wollaston prism they will interfere and generate contrast ranging from black to white

Question 32

If the beams are not diffracted by the specimen (DIC)

Answer 32

They are not recombined and produces a pale grey colour

Question 33

Wollaston prism is use for which light miscopy method and what is it?

Answer 33

DIC | - 2 quartz wedges, causes a beam of polarised light into 2 beams which travel close to eachother

Question 34

Single cells can be ______ ______

Answer 34

viewed directly

Question 35

Tissues may need ________

Answer 35

Sectioning

Question 36

Detection of cellular components or cells types may need?

Answer 36

Dyes or chromogenic enzyme substrates

Question 37

Example of plant roots cells becoming stained

Answer 37

plants produce phenolic compounds if they become damaged, which can be died and detected

Question 38

Lignin in cells walls ???

Answer 38

can be dyed and detected

Question 39

example of enzymes being used as dyes

Answer 39

Artificial substrate reveals protease activity from apathogenic fungus infecting insect cuticle. Apply a substrate>>> colour change when in the presence of proteases can see where the fungus is localised

Question 40

Dyes also allow the detection of

Answer 40

life or death in cells

Question 41

Example of neutral red dye use??

Answer 41

in dead cells (infected by a fungi) dye is not retained by vacuole (as the vacuole membrane is damaged) Living cells take up dye

Question 42

Fluorescence is the property of??

Answer 42

Fluorescence is the property of absorbing light of a particular wavelength and then emitting light of a different wavelength.

Question 43

Visible light is defined by?

Answer 43

Visible light is usually defined as having wavelengths in the range of 400–700 nm.

Question 44

What type of filters used for Fluorescence light miscopy ??

Answer 44

Selective filters (illuminate at a particular wavelength of light)

Question 45

First light passes through ?? Causes??

Answer 45

Filter (particular wavelength of light produced) Excitement of a fluorescent compound

Question 46

Light is then focused (from filter) Fluorescence light miscopy

Answer 46

Onto the objective lens

Question 47

From the objective lens.. light _____ (Fluorescence light miscopy )

Answer 47

light passes through a beam splitter

Question 48

How is data viewed (Fluorescence light miscopy)

Answer 48

By eye or computer

Question 49

Applications of Fluorescence light miscopy

Answer 49

Visualising organelles within cells e.g. by using immunofluorescence microscopy. locations of molecules within cells (multiple probes multiple structures)

Question 50

Confocal microscopy purpose?

Answer 50

Increases resolution - standard fluorescence microscopy

Question 51

Confocal microscopy research options ?

Answer 51

1) co-localisation 2) intra-cellular studies 3) examination of thick specimens 4) studies where 3-D structure of the sample is important.

Question 52

Confocal microscopy allows for?

Answer 52

Optical sections to be made without actually having to form sections within a specimen

Question 53

Image volume for confocal microscopy?

Answer 53

DEEP

Question 54

where is light collected from confocal miscopy?

Answer 54

Exclusively collect light from the focal plane

Question 55

GFP derived from?

Answer 55

Aequorea Victoria- GFP

Question 56

Expression profiles allow??

Answer 56

see where a gene is localised