methods of studying cells

Question 1

what are the principles and limitations of optical microscopes

Answer 1

light focused using glass lenes
light passes through thin speciman
generates 2D image

x low resolution 200nm due to long wavelength of light
x low magnification x1500

+ can view living organisms x only thin speciman
+ simple prep
+ can show colour

Question 2

what are the princliples and limitations of transmission electron microscope

Answer 2

electrons focused using electromagnets
2D cross section
denser areas= more electrons absorbed = darker

+ v. high resolution 0.2nm as electrons have short wavelength
+ high magnification x1,000,000

x dead organisms
x complex prep so artefacts present
x no colour

Question 3

what are the principles and limitations of a scanning electron microscope

Answer 3

electrons focused using electromagnets
scan speciman surface
3D image

+ high resolution 2nm due to short wavelength of electrons
+ high magnification x1,000,000

x dead speciman
x no colour
x complex prep so artefacts likely

Question 4

how did scientists distinguish artefacts

Answer 4

artefacts often occur during prep of sample
scientists prepared specimens in different ways
if an object could be seen with one technique but nit another its more likely to be an artefact

Question 5

what is magnification

Answer 5

number of times greater the image is that the size of the real object

Question 6

how to calculate magnification

Answer 6

size of image/ size of real object

Question 7

what is resolution

Answer 7

minimum distance apart 2 objects can be for them to be distinguished as seperate objects
limited by wavelength

Question 8

how to convert units

Answer 8

m
mm
um
nm

x1000 every time going down
/ 1000 every time going up

Question 9

what is an eyepiece graticule

Answer 9

where - in the eyepiece, spans across entire field of view
units- no fixed units, must be callibrated to work out the sixe of divisions at a particular magnification
use - remains in the field of view when viewing speciman so is used to measure size of the objects

Question 10

what is a stage micrometer

Answer 10

where - on a microscope slide placed on the stage
units- does have fixed units, 1mm long micrometer is normally divided into 100 units of 10 um
use- used to calibrate the eyepiece graticule at a particular magnification

Question 11

how to measure the size of objects viewed with optical microscopes using eyepiece graticule and stage micrometer

Answer 11

1. line up the eyepiece graticule with the stage micrometer
2. calibrate the eyepiece graticule- use stage micrometer to calculate size of divisions on the eyepiece graticule
3. take the micrometer away and use the graticule to measure how many divisions make up the object
4. calculate the size of the object by multiplying the number of divisions by size of divisions
5. recalibrate at different magnifications

Question 12

what are the principles of cell fractination and ultracentrifugation

Answer 12

1. homogenise tissue using blender- disrupts cell membrane and breaks open cell to release contents
2. keep in cold, isotonic and buffered solution
3. filter homogenate to remove large unwanted debris
4. ultracentrifugation to seperate organelles in order of density
   - centrifuge homogenate in a tube at low speed
   - remove pellet of heaviest organelle and respin at higher speed

Question 13

what properties dies the solution that the homogenate is kept in need to be and why

Answer 13

cold - to reduce enzyme activity so organelles arent broken down

isotonic- so water doesnt move in or out by osmosis so they dont burst

buffered - to keep pH constant so enzymes dont denature

Question 14

what is the order that the organelles are seperated in ultracentrifugation

Answer 14

nuclei
chloroplast/ mitochondria
lysosomes
endoplasmic reticulum
ribosomes