Light Microscopy Flashcards
1
Q
Define refraction
A
- change in direction of light when passing from one
medium to another
2
Q
Explain the principle of magnification using a convex lens
A
- as parallel light rays from object pass t/ magnifying glass, convex lens causes refraction and changes direction of light path
- passes through a 2nd lens in eye, hits the retina and
converted to image by brain.
3
Q
How to microscopes achieve higher mag (>5x)?
A
- objectives (air and oil)
4
Q
What is a parafocal objective?
A
- if focus image correctly at particular magnification, can move to next higher magnification objective and remain close to the focal plane
5
Q
Define refractive index?
A
- extent materials refract light
- affects how path of light travels t/ sample
6
Q
What is light scatter?
A
- diff in refractive index between air and glass causes light refraction and radiates out from sample
7
Q
Why use oil-immersion microscopy?
A
- continuous physical contact between the slide and oil objective
- immersion oil has same refractive index as glass, so prevents light scatter and increases light collection by objective
- significantly increase the resolution of an
image when performing high-magnification microscopy
8
Q
Define resolution
A
- min distance at which two points can be distinguished
within a defined space
9
Q
Define exposure time
A
- length of time light is collected to produce image
10
Q
What is image saturation?
A
- if exposure times too long, areas of image appear white, show no detail and becomes saturated, where sensor chip cannot absorb anymore light
11
Q
What is gain?
A
- digital amp of data collected at particular exposure time, can boost weak signals
- but excessive gain reduces res and loses detail
12
Q
How would you balance exposure and gain?
A
- ideally to get v detailed image would have exposure just below saturation (quite high) and no gain
- practically would start w/ lower exposure w/ a little bit of gain, to make the process a bit faster
