microscopy

Question 1

smallest to largest measuremtns

Answer 1

angstrom –> nanometer –> micrometer –> millimeter –> centimeter

Question 2

refractive index

Answer 2

how much a substance (i.e. water, air) slows the velocity of light

when light transitions through substances it changes speed and bends

Question 3

focal points and focal lengths

Answer 3

focal point: where the light beams are all focused onto this single point

focal lengths: distance between lens and focal point

Question 4

brightfield microscope

Answer 4

light source –> condenser lens (w object on it) –> objective lens to see through

Question 5

resolution

Answer 5

minimum distance between 2 objects so they can still be viewed as seperate from each other

-lower # is better resolution

Question 6

numerical aperture (N sin theta)

Answer 6

theta is the angle the cone of light is entering an object (max is 90 degrees)
N= refractive index

refractive index of air is 1

Question 7

oil immersion objective

Answer 7

use oil between lens and object to limit refraction (no air)

change refractive index

Question 8

Abbe equation

Answer 8

d= 0.5 lambda/ n sin theta

d=resolution (smaller is better)
lambda= wavelength
n= refractive index
theta= angle

minimize resolution: decrease wavelength, or increase n sin theta

Question 9

3 ways to visualize unpigmented microbes through light microscopy

Answer 9

1. dark-field microscopes
2. phase-contrast microscopes
3. differential interference contrast (DIC) microscopes

Question 10

dark field microscope

Answer 10

but a “poker chip” right after light source, only peripheral beams reach objective lens

Question 11

phase-contrast microcopy

Answer 11

have 2 different wavelengths through the wavelengths passing through bacteria (high refractive index) and the ones that dont pass through

-to further perturb the wavelength there is a phase plate

-condenser annulus “poker chip”
and phase plate; exaggerate difference in wavelengths

able to see motility, detailed structures like organelles

Question 12

fluorescent microscopes

Answer 12

opposite of light microscopes

light source is perpendicular to a tube; emits short wavelengths

there is a dichromatic mirror which reflects short wavelengths and transmits longer wavelengths

so short wavelengths hit mirror and go to specimen which emits longer wavelengths which bypass mirror again and go up to visualize it

fluorochromes: stain DNA

i.e. GFP

Question 13

confocal scanning laser microscopy

Answer 13

looking at 3D not flat!!!

create z stacks to get many focal planes

Question 14

light vs electron microscopy

Answer 14

electron microscopy uses electron beams instead of light

and use magnets as lens to bend the beams instead of glass

electron microscope is way higher energy wavelength then light microscope which means its more likely to hit things (i.e. jump rope and throw ball in)

but need to kill specimens and use chemicals