3.2.1.3 Methods of studying cells Flashcards
what is magnification?
how much the image can be increased in size
what is resolution?
the ability to determine between two separate points
what is the equation for magnification?
magnification = image / object
what is another name for an optical microscope?
a light microscope
what are the principles of an optical microscope?
directs light waves through a sample (or reflects light off the surface of a live sample) into the eye of the observer
what is the purpose of staining a specimen?
it is used to colour cell structures so that they can be seen as most cells are transparent / don’t contain pigment
what is the maximum magnification of an optical microscope?
1500 x
what is the maximum resolution of an optical microscope?
0.2 µm
why is the resolution of an optical microscope low?
because light rays have a longer wavelength
optical microscopes can view ____ specimens?
live
why does the specimen need to be extremely thin for an optical microscope?
so that the light rays can pass through
what is the resolution of an electron microscope?
upto 0.1 nm
why is the resolution of an electron microscope better than that of an optical microscope?
the electron beam has a shorter wavelength than light
what are the two types of electron microscopes?
- transmission
- scanning
what are the principles of a TEM?
- electrons fired at the specimen
- denser parts absorb more electrons and so appear darker
what are the principles of a SEM?
uses a focused beam of high-energy electrons which reflect off the surface of structures
what are the advantages of using a TEM over a SEM?
- can view internal structures
- higher resolution
what are the limitations of electron microscopes?
- the whole system must be in a vacuum so living specimens cannot be observed
- a complex staining process is required which may introduce artefacts (e.g. dust or air bubbles) into the image
- specimens have to be very thin, especially for the TEM so that electrons can pass through
- SEM has a lower resolving power than TEM
what is cell fractionation?
the process by which cells are broken up and the different organelles are separated out based on their mass using gravity
what are the three main stages of cell fractionation?
- homogenisation
- filtration
- centrifugation
what type of solution does the tissue need to be placed in before cell fractionation and why?
- ice-cold - reduces enzyme activity to prevent digestion of organelles
- isotonic - prevents osmosis so no lysis / shrinkage of organelles
- buffer - maintains pH so that enzymes / proteins are not denatured
what happens during homogenisation?
breaking up the tissue to break open cell membranes and release organelles
what happens during filtration?
homogenate is filtered to remove large debris as this will be heavy and will sink to the bottom of the test tube
what happens during centrifugation?
- faster spinning = a greater force
- at a slower speed, large fragments collect at the bottom forming a pellet whilst, at the top, smaller fragments remain suspended in a liquid called the supernatant
- the supernatant is transferred to another test tube and re-spun at higher speeds
- smaller organelles collect at the bottom forming a new pellet
