Microscopy

Question 1

definition of magnification

Answer 1

how much bigger the image is compared to the specimen

Question 2

definition of resolution

Answer 2

ability to distinguish between 2 points that are close together as separate structures

Question 3

formula for magnification

Answer 3

image/actual

Question 4

definition of cell fractionation

Answer 4

process whereby cells are broken up + cell organelles separated out (homogenisation + ultracentrifugation)

Question 5

definition of ultracentrifugation

Answer 5

process by which cellular fragments in a homogenate are separated out in a centrifuge at different speeds

Question 6

before breaking up cells you must put it in….

Answer 6

a cold, buffered solution

Question 7

why does the solution need to be cold?

Answer 7

so enzymes don’t denature, slows metabolic activity = reduces enzyme activity that could break down organelles.

Question 8

why does the solution need to be buffered?

Answer 8

stabilizes pH so does not damage cell organelles/ enzymes

Question 9

why does the solution need to be isotonic?

Answer 9

so cells don’t shrivel/burst from osmotic pressure

Question 10

what is the speed of centrifugation to separate out nuclei?

Answer 10

1000 min-1

Question 11

what is the speed of centrifugation to separate out mitochondria?

Answer 11

3500 min-1

Question 12

what is the speed of centrifugation to separate out lysosomes?

Answer 12

16500 min-1

Question 13

what is the solution at the bottom of the centrifuged called?

Answer 13

sediment/pellet

Question 14

what is the solution at the top of the pellet called which is then centrifuged again?

Answer 14

supernatant

Question 15

what are artefacts?

Answer 15

things that result from the way a specimen was being prepared (not part of the natural specimen)

Question 16

does optical microscopes have a long or short wavelength?

Answer 16

long

Question 17

what is the magnification of an optical microscope?

Answer 17

x1500

Question 18

what is the magnification of TEM?

Answer 18

x500000

Question 19

what is the magnification of SEM?

Answer 19

x100000

Question 20

TEM: shine an electron beam ________thin specimens = __D image.
SEM: shines an electron beam _____ the _______ of specimens = __D image.

Answer 20

through, 2D

onto the surface, 3D

Question 21

advantages and limitations of using Optical microscope?

Answer 21

advantages: easy to use/lightweight, cheap, does not require radiation
disadvantages: poor/low resolution

Question 22

advantages/limitations of TEM

Answer 22

advantages: better resolving power than SEM
limitations: living specimens can’t be viewed, need to be in vacuum, specimen extremely thin, complex preparation (artefacts), only 2D images

Question 23

advantages/disadvantages of SEM

Answer 23

advantages: specimens don’t need to be thin, preparation is less complex, less chance of artefacts
limitations: lower resolving power than TEM, whole system in vacuum, no living specimens

Question 24

eyepiece graticule must be calibrated by using…

Answer 24

a stage micrometer

Question 25

in ultracentrifugation, the _______ organelle becomes the sediment/pellet first at the ______ speed

Answer 25

heaviest, lowest

Question 26

both TEM and SEM have ____ wavelengths

Answer 26

short

Question 27

How to calibrate a microscope?

Answer 27

stage micrometer units/eyepiece graticule units

this gives the conversion number

Question 28

how to calculate specimen size from EPGUs?

Answer 28

1. observe specimen i.e. extends 8-88 EPGUs
2. calculate size of specimen i.e. 88-8=77 EPGUs
3. find actual size by multiplying it by conversion no. i.e. 77 EPGUs x 2.5 micrometers = 192.5 micrometers