Analysis of Cell Components

Question 1

Magnification

Answer 1

how many times bigger the image produced by the microscope is, compared to the real object under the microscope

Question 2

Resolution

Answer 2

the ability to distinguish between objects that are close together

Question 3

Optical microscopes

Answer 3

use light to form an image

Resolution is limited (max. Resolution of ~0.2 micrometres)

used to observe eukaryotic cells, nuclei and sometimes mitochondria & chloroplast

The maximum useful magnification of optical microscopes is about ×1500

Question 4

Electron microscopes

Answer 4

use of electrons to form an image

Increased resolution - electron beams have a smaller wavelength than light, so can resolve objects close together

Max. resolution of 0.2nm
Can be used to observe small organelles - ribosomes, endoplasmic reticulum or lysosomes

Max magnification is ~ x1,500,000

Question 5

Transmission electron microscopes

Answer 5

use electromagnets to focus a beam of electrons → the beam is transmitted through the specimen

The denser parts of the specimen absorb more electrons → appear darker

Give high resolution images so you see the internal structure of organelles

Only used on thin specimen

Question 6

Scanning Electron Microscope

Answer 6

Scan a beam of electrons across the specimen → beam bounces off the surface of the specimen → electrons are detected → forms an image

SEMs produce three-dimensional images that show the surface of specimens

Used on thick specimens

But give a lower resolution image than TEMs

Question 7

How to prepare an optical microscope slide?

Answer 7

Pipette a small drop of water onto the slide
Then using tweezers to place a thin section on top of drop of water
Add drop of stain
Add the cover slip, carefully tilt and lower reducing chances of air bubbles

Question 8

Magnification calc

Answer 8

Size of image/ size of real object

Question 9

Cell fractionation

Answer 9

the process of separating cell organelles from each other - often for imaging or research purposes

Question 10

What happens during Homogenisation?

Answer 10

breaking up cells using a homogeniser that grinds up cell plasma membrane and releases organelles into solution. Tissue sample must be in cold (reduce the activity of enzymes) , isotonic buffer (prevent cells being broken down, prevent cells undergoing osmosis and maintain pH). The product is called homogenate

Question 11

Filtration

Answer 11

Homogenate is filtered through a gauze which separates the large cell debris and tissue debris that wasn’t broken down

Question 12

Ultracentrifugation

Answer 12

the filtrate (filtered product) is placed into a tube and the tube is placed in a centrifuge

The filtrate is first spun at low speed → largest, heaviest organelles (e.g., nuclei) settle at the bottom of the tube → forms a pellet (thick, sediment-like material) + the supernatant (rest of the solution, containing organelles)

The supernatant is then centrifuged again → process repeated at increasing speeds until all different types of organelles present are separated out OR the desired organelles is found

Question 13

Artefacts

Answer 13

things you may see in a prepared slide that aren’t part of the specimen, and often occur during the preparation of a sample