studying cells

Question 1

define magnification

Answer 1

how many times larger the image is compared to the object
(mag. = image / actual)

Question 2

define resolution

Answer 2

the minimum distance between two objects in which they can be views as separate

Question 3

how is resolution determined in an optical and electron microscope?

Answer 3

optical = wavelength of light
electron = wavelength of beam of electrons

Question 4

what creates the image in an optical (light) microscope?

Answer 4

a beam of light is released by the lamp which is shone up and then condensed which creates the imagew

Question 5

what creates the image in an electron microscope?

Answer 5

a beam of electrons is condensed using electromagnets to create the image

Question 6

what are the advantages of using an optical microscope?

Answer 6

• colour images are produced
• living samples can be used
• cheaper

Question 7

what are the disadvantages of using an optical microscope?

Answer 7

• poor resolution due to light having a longer wavelength
• lower magnification
  ==> this means you cannot see the inside of organelles in detail or for some small organelles, they aren’t visible at all

Question 8

what are the advantages of using an electron microscope (general)

Answer 8

• higher magnification
• higher resolution due to electron beams having a shorter wavelength

Question 9

what are the disadvantages of using an electron microscope (general)

Answer 9

• sample must be in a vacuum so therefore only a dead specimen can be used
• complex staining process which may contain artefacts
• black and white image produced only

Question 10

what are the two types of electron microscopes?

Answer 10

transmission and scanning

Question 11

explain how transmission electron microscopes produce an image?

Answer 11

1) extremely thin specimens are stained and placed in a vacuum
2) an electron gun produces a beam of electrons that pass through
==> the areas which electrons pass through appear light on the image
==> some areas absorb the electrons and these will appear dark - the more electrons absorbed means the darker the area
3) the image produced is 2D and shows detailed image on the internal structure
==> Tem = Two d

Question 12

explain how scanning electron microscopes produce an image

Answer 12

the specimens do not need to be thin as the electrons aren’t transmitting / passing through
1) the electrons are beamed onto the surface where the electrons scatter and reflect un different ways depending on the contours / depth
2) the different depths create a 3D image and details to do with texture and 3D depths are shown of either cells or organelles

Question 13

what are the key differences between SEM and TEM?

Answer 13

• SEM produces 3D image, TEM produces 2D image

-SEM image is views on a screen as there is no eyepiece, TEM image is viewed with eyepiece

• SEM has lower resolution, TEM has higher resolution

-SEM electrons are deflected off the specimen, TEM electrons pass through the specimen

Question 14

what is the equation for magnification?

Answer 14

magnification = image size
—————
actual size

Question 15

how do you convert units
m - mm - um - nm

Answer 15

m - nm = x1000^3
nm - m = divide 1000^3

Question 16

in a magnification calculation question, what does the scale bar represent and how would you use it to calculate the image size?

Answer 16

the actual size
- measure the scale bar to find the image size

Question 17

what is cell fractionation?

Answer 17

the process where cells are broken open to release the contents and the different organelles within are then separated

Question 18

what type of solution must the cells be prepared in before cell fractionation?

Answer 18

cold, isotonic, buffered

Question 19

why must a solution for cell fractionation be cold?

Answer 19

(reason and explanation must both be stated for the mark)
1- to reduce enzyme activity
2- when the cell is broken up enzymes are released which could damage the organelles

Question 20

why must a solution for cell fractionation be isotonic?

Answer 20

(reason and explanation must both be stated for mark)
1- to prevent organelles shrinking or bursting
2- due to osmosis

Question 21

why must a solution for cell fractionation be buffered?

Answer 21

(reason and explanation must both be stated for mark)
1- a constant pH
2- preventing proteins denaturing and damaging the organelles

Question 22

what are the two stages in cell fractionation?

Answer 22

stage 1 - homogenisation
stage 2 - ultracentrifugation

Question 23

explain the process of homogenisation

Answer 23

-The cells are broken up (homogenised) to make a homogenate by using a blender and blending the cells in a cold, isotonic, and buffered solution
- This homogenate is then filtered to remove debris and whole cells

Question 24

what is ultracentrifugation?

Answer 24

the process by which the fragments in the filtered homogenate are separated according to their density by a centrifuge machine
- this enables separation of organelles by high speed spinning

Question 25

explain the process of ultracentrifugation

Answer 25

• a test tube of filtrate is spun at a lower speed to begin with
• the heaviest organelle, the nuclei, are forced to the bottom of the tube where they form a pellet
• the fluid at the top of the tube, supernatant, is removed and transferred to another tube where it is spun at a faster speed and for longer
• the next heaviest organelles, mitochondria and chloroplasts, are forced to the bottom of the tube
• this process is continued with increasing speed so that each organelle is isolated from the cell (differential centrifugation)

Question 26

what is the order of organelle fractionation?

Answer 26

nuclei
mitochondria / chloroplasts
endoplasmic reticulum
lysosomes
ribosomes

Question 27

why does the speed of the centrifuge increase as the organelles get lighter?

Answer 27

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