Microscopy - Midterm 1 Flashcards
How does light travel? in bright field microscopy ?
Light source -> substage condenser -> stage -> objective lens -> ocular lens
How does light travel in phase contrast microscopy?
Light source -> annular stop -> substage condenser -> stage -> objective -> phase plate -> focal plane -> ocular lens
What happens when you have too little light?
you get reduced clarity
What happens when you have too much light?
you get reduced contrast
What is resolution?
Ability of microscope to clearly distinguish two very small objects that are placed very close together as two separate entities
What is object?
the material that is being examined
What is image?
what you see with your eyes
What is field?
area on the slide that is being viewed
Total magnification is equal to
Objective x ocular (ocular is always 10)
What does parfocal mean?
when the specimen is in focus at one magnification, it will be in focus at other magnifications, requiring only minor adjustments to the stage level
Greater resolution =
the closer something can be and still be distinguished as two different things
What is the limit of resolution?
smallest distance between the two objects that will still allow them to be viewed as 2 separate objects.
Average light microscope has a limit resolution of what?
0.2 micrometer. therefore, bacterial cells of 0.5 micrometer diameter can still be viewed as separate.
What is a condenser diaphragm?
Works like the iris of the eye and controls the amount of light that enters
What happens when you open the diaphragm?
you increase the amount of focused light –> this increases clarity but decreases contrast
What happens when you close the diaphragm?
Decreasing the amount of focused light –> increasing the contrast but decreasing the clarity
True or false? Its good to use the diaphragm to control the intensity or brightness of light
False, it reduces the resolution
What is annular stop? What happens to light there?
Ring with an opaque center that blocks light in the center. this produces a hollow cylinder of lightThere is no phase shift or angle change
What happens when light passe through substage condenser after the annular stop?
there is a change in angle in the light as it bends to form a hollow cone shape. There is no change in phase
What happens the light when it hits the object?
The light that hits the cells gets diffracted and changes angle, they also change phase by -1/4 lambda
The light that hits the medium doesnt experience any change in phase shift or angle
What happens when light hits the phase plate?
The diffracted light that hits the cells are now passing through the clear center of the ring of the phase plate
The light that wasnt defracted in the medium will hit the ring of the phase plate causing it to phase shift to +1/4
