T2: Microscopy

Question 1

magnification

Answer 1

how many times bigger the image produced by the
microscope is than the object

Question 2

resolution

Answer 2

the ability to distinguish between objects that are close together
(the ability to see two structures that are very close together as two separate structures)

Question 3

types of microscopes

Answer 3

optical
electron - SEM and TEM

Question 4

magnification formula

Answer 4

magnification = size of image / size of object
IAM triangle

all units must be the same

Question 5

why can’t all organelles be seen with a optical microscope?

Answer 5

the resolution is too low because the wavelength of light is long

Question 6

how do optical microscopes work?

Answer 6

use visible light that passes through the specimen and two condensers

max resolution - 0.2pm
max magnification - x1500
produce a 2D image in colour
require thin specimens

Question 7

how do transmission electron microscopes work?

Answer 7

use a beam of electrons that passes through the specimen, area that absorbs electrons appears darker

Question 8

how do scanning electron microscopes work?

Answer 8

use a beam of electrons that pass across the surface and scatter, the pattern of scattering builds a 3D image based on the shape of the specimen

produces a 3D image in black and white

Question 9

magnification and resolution of electron microscopes

Answer 9

resolution = 0.2nm (1000x greater than optical)
can observe ribosomes, ER and lysosomes
magnification = 1,500,000x (1000x greater)

Question 10

advantages of TEM’s

Answer 10

give hi-res images (more detail), allows internal structures within cells to be seen

Question 11

disadvantages of TEM’s

Answer 11

can only be used with very thin specimens
can’t be used on live specimens (has to be in a vacuum so all water gone)
lengthy treatment to prep specimens means artefacts can be introduced
don’t produce a colour image

Question 12

advantages of SEM’s

Answer 12

can be used on thick, 3D specimens
allow the external 3D structure of cells to be seen

Question 13

disadvantages of SEM’s

Answer 13

give low-res images (less detail) than TEMs
can’t be used on live specimens (has to be in a vacuum so all water gone)
don’t produce a colour image

Question 14

what is an eyepiece graticule?

Answer 14

disc with engraved scale present in the eyepiece, has no units

Question 15

how to calibrate a eyepiece graticule

Answer 15

1. Align the eyepiece graticule with the stage micrometer
2. Count how many divisions on the graticule correspond with the stage micrometer
3. Calculate how long one EPU is, check the size of your micrometer

you have to calibrate for every magnification (LP,MP,HP)

Question 16

how to make a temporary wet mount

Answer 16

add a drop of water to the slide
balance specimen atop the water drop
stain with iodine in KI or a suitable stain
lower coverslip gently using forceps

Question 17

what is a stage micrometer?

Answer 17

ruler that you place on the microscope’s stage