microscopy

Question 1

define magnification

Answer 1

how much bigger the image is compared with the actual size of the object

Question 2

what is the equation to calculate magnification

Answer 2

image size/ actual size

Question 3

how many mm in a m

Answer 3

1m=1000m

Question 4

how many um in a mm

Answer 4

1mm=1000um

Question 5

how many um in a m

Answer 5

1m= 1000000um

Question 6

how many um in a nm

Answer 6

1um=1000nm

Question 7

define resolution

Answer 7

the smallest distance two points can be separated and still be seen as separate items

Question 8

(exam q) why do we use ‘this’ unit

Answer 8

• it is the most appropriate unit
• avoids the use of excessive decimal places
• avoids the use of using unnecessary standard form

Question 9

to what limit can the human eye can see

Answer 9

+- 0.25mm

Question 10

what are the features of the light microscope (4)

Answer 10

• limited to a resolution of 0.4um
• can use living specimens
• easy to prepare specimen
• images produced in colour

Question 11

what are the features of the electron microscope (6)

Answer 11

• has a resolution up to 0.1nm
• specimen must be dry, dead and extremely thin
• extremely complex preparation of the specimen
• images produced in black and white
• slicing the sample to make it thin can damage the inter-molecular structures
• image can produce artifacts (things on the object that arent actually real) e.g. air bubbles

Question 12

describe the resolving power of the transmission electron microscope

Answer 12

it has a maximum resolution of 0.1nm which is not always achieved due to the difficult specimen preparation and the high energy (electron) beam which can destroy the specimen

Question 13

advantages of the light microscope (3)

Answer 13

• can see living specimens
• easy to prepare specimen
• variety of coloured stains

Question 14

disadvantages of the light microscope

Answer 14

• low resolution so organelle details/smaller components are not visible

Question 15

advantages of the transmission electron microscope (2)

Answer 15

• has a very high resolution at high magnification
• can identify detailed organelle/sub-organelle structure

Question 16

disadvantages of the transmission electron microscope (4)

Answer 16

• specimen not alive (in a vacuum)
• difficult preparation (very thin specimen, complex staining)
• produces a black and white image
• artefacts can spoil image

Question 17

to what resolution can the scanning electron microscope reach to

Answer 17

20nm

Question 18

advantages of the scanning electron microscope (1)

Answer 18

• 3D images show structural formation

Question 19

disadvantages of the scanning electron microscope (3)

Answer 19

• specimens are not alive
• difficult to prepare specimen
• produces a black and white image

Question 20

why do we stain specimens

Answer 20

it helps to identify specific features of cells and improves contrast within the specimen

Question 21

name the three processes to fractionate a cell (isolating and separating individual components)

Answer 21

1- homgenisation
2- filtration
3- differential ultracentrifugation

Question 22

describe the process of homogenisation

Answer 22

cells vibrated or grinded in a blender, breaking them open.

Question 23

why is a cold isotonic buffer used in homogenisation

Answer 23

-cold to stop organelle activity (hydrolytic enzymes in lysosomes)
-> lysosomes used to digest material inside the cell. contain hydrolytic enzymes which hydrolyses the components in the homogenate (biological molecules) such as polypeptides
- isotonic to prevent the movement of water in and out of the cell through osmosis (prevents organelles bursting through the movement of water in)

Question 24

why is a buffer solution used in homogenisation

Answer 24

it prevents change in PH levels, stopping important proteins from being denatured

Question 25

describe filtration

Answer 25

solution is filtered through a gauze to remove cellular debris

Question 26

describe differential ultracentrifugation

Answer 26

- solution spun in a centrifuge at a low speed - heaviest organelles fall to the bottom (form a pellet), other organelles stay suspend in the fluid above the pellet (called the supernatant) - pellet removed or supernatant depending on what you are looking for- - (pellet removed) supernataant spun at a higher speed - pellet forms. supernatant removed (these contain the lightest organelles)