Microscopy Flashcards
What is magnification
how much bigger sample is under microscope compared to actual size.
What is resolution
Ability to distinguish between two points on image.
amount of detail
how do you calculate total magnification
objective magni x eyepiece magni
in a light microscope what are specimens exposed to ?
Light
what distance must objects be in a light microscope to be seen as one object
less than 200nm apart
what is light microscopes magnification limited to
1500x and is limited by its resolving power
what are specimens exposed to in an electron microscope
Electrons
why do electron microscopes produce images with better resolution
as electrons have shorter wavelength than light
What are the two types of electron microscopes
Scanning (S.E.M)
Transmission (T.E.M)
how are specimens prepped for S.E.M
coated in thin layer of metal (gold,platinum,tungsten)
why are specimens coated for S.E.M
improve conductivity and contrast
what happens to the electrons in S.E.M
reflected of the surface
what image does S.E.M produce
3D image of whole cells/tissues
how are specimens prepared for T.E.M
very thin slices stained with heavy metal
what happens to the electrons in T.E.M
absorbed by heavily stained parts
what image does T.E.M make
2D image of inside the cells/organelles
what are the 4 different preparation methods
1.Dry mount
2.wet mount
3.squash slide
4.smear slide
how do you prepare slides with the dry mount method
thin slice of specimen placed on slide with cover slip over it
how do you prepare slides with the wet mount method
small drops of liquid on slide specimen
Cover slip on top + ensure no air bubbles
how do you prepare slides with the squash slide method
-use soft specimen so can look at cell division.
-use wet mount method then place cover slip over specimen
how do you prepare specimen with the smear slide method
Often used for blood samples.
-use edge of slide to smear sample making it thin. At 45 degree angle
-cover slip over sample
what is an artefact
structural detail caused by processing specimen e.g air bubbles
purpose of staining
help see cell structures
what is purpose of differential staining
bind to specific cell structures and stain them differently so can be easily identified
what does methylene blue stain help see
nuclei
what does iodine help see
stains starch to see plant cell walls
what is gram staining
stain bacterial cell walls.
how do you calculate magnification
image size/actual size
what is eyepiece graticule
-fine scale
-fits inside the eyepiece lens
-units are arbitrary
what is the stage micrometer
-scale fitted on cover slip.
-size of divisions known
