2.1.3 Methods to Study Cells Flashcards
which methods help to determine the internal structure of cells
-microscopes
-cell fractionation
-ultracentrifugation
what are the three key types of microscopes
-optical microscopes
-transmission electron microscopes
-scanning electron microscopes
define magnification
-how many times greater the image is compared to the object
define resolution
-the minimum distance between two objects in which they can still be distinguished as separate objects
features of an optical (light) microscope
-light focused using glass lenses
-light passes through specimen, different structures absorb different amounts & wavelengths
-generates a 2D image of a
cross-section
-low resolution due to long
wavelength of light
-can’t see internal structure of
organelles or ribosomes
-specimen = thin
-low magnification (x 1500)
-can view living organisms
-simple preparation
-can show colour
why must samples be in a vacuum for electron microscopes
- electrons are absorbed by air
how do transmission electron microscopes work
-electrons focused using
electromagnets
-electrons pass through specimen, denser parts absorb more and appear darker
-generates a 2D image of a
cross-section
-very high resolution due to short wavelength of electrons
-can see internal structures of
organelles and ribosomes
-specimen = very thin
-high magnification (x 1,000,000)
-can only view dead / dehydrated specimens as uses a vacuum
-complex preparation so
artefacts often present
-does not show colour
how do scanning electron microscopes work
-electrons focused using
electromagnets
-electrons deflected / bounce
off specimen surface
-generates a 3D image of surface
-high resolution due to short
wavelength of electrons
-can’t see internal structures
-specimen does not need to be thin
-high magnification (x1,000,000)
-can only view dead / dehydrated specimens as uses a vacuum
-complex preparation so artefacts often present
-does not show colour
formula for size of structures viewed under an optical microscope
image size = actual size x magnification
order for units
metre
millimetre
micrometre
nanometre
order for conversion (largest to smallest)
x1000
x1000
x1000
order for conversion (smallest to largest)
divide by 1000
divide by 1000
divide by 1000
conditions needed during homogenisation
-cold
-isotonic
-buffered
what happens when the solution is centrifuged
the solution is placed into a centrifuge which spins at high speed to separate organelles depending on their density due to the centrifugal force
suggest how the scientific community distinguished between artefacts (eg. dust, air bubbles occurring during preparation) and cell organelles
-scientists prepared specimens in different ways
-if an object was seen with one technique but not another, it was more likely to be an artefact than an organelle
describe how the size of an object viewed with an optical microscope can be measured
- line up (scale of) eyepiece graticule with (scale of) stage micrometre
- calibrate eyepiece graticule - use stage micrometre to calculate size of divisions on eyepiece graticule
- take micrometre away and use graticule to measure how many divisions make up the object
- calculate size of object by multiplying number of divisions by size of division
- recalibrate eyepiece graticule at different magnifications
explain why tissue is homogenised in cell fractionation
-disrupts the cell membrane, breaking open cells to release
contents / organelles
explain why cell contents are placed in a cold solution in cell fractionation
-to reduce enzyme activity
○ so organelles not broken down / damaged
explain why cell contents are placed in an isotonic solution in cell fractionation
-so water doesn’t move in or out of organelles by osmosis
○ so they don’t burst
explain why cell contents are placed in a buffered solution in cell fractionation
-to keep pH constant
○ so enzymes don’t denature
explain why the homogenate is filtered
-remove large, unwanted debris eg. whole cells, connective tissue
explain why ultracentrifugation is used
-separates organelles
in order of density / mass
describe ultracentrifugation
-centrifuge homogenate in a tube at a low speed
-remove pellet of heaviest organelle and respin supernatant at a higher speed
-repeat at increasing speeds until separated out, each time the pellet is made of lighter organelles
order of density in pellets in ultracentrifugation
(nuclei → chloroplasts /
mitochondria → lysosomes → ER → ribosomes)
