Two types of microscopes
Light microscope and electron microscope
Light microscope images allow the observation of …
Cell shape and larger internal structures
Advantages of light microscopes:
they are small and relatively cheap,
specimen preparation can be straightforward enough to perform in a school laboratory,
they can be used to produce colour images,
they allow the observation of living specimens
Electron microscopes can be used to observe …
Small structures inside cells
Limitations of electron microscopes:
They are very large and expensive,
Specimens must be prepared using a highly complex process,
Specimens must be viewed in a vacuum meaning that live specimens cannot be observed,
Images are always black and white though they can be artificially coloured during processing
Two types of electron microscope:
Transmission electron microscopes (TEMs)
and scanning electron microscopes (SEMs)
How do TEMs create an image?
TEMs use electromagnets to transmit a beam of electrons through a specimen; denser parts of the specimen absorb more electrons, meaning that denser parts appear darker on the final image
How do SEMs create an image?
SEMs pass a beam of electrons across the surface of a specimen and then detect the rate at which the electrons bounce back
TEMs produce images that:
Are high-resolution,
allow the internal structures within cells and within organelles to be seen,
are two-dimensional
SEMs produce images that:
Are three-dimensional,
show the surface of specimens,
SEMs have a lower maximum resolution than TEMs
How to prepare a slide using a liquid specimen:
- Add a few drops of the sample to the slide using a pipette
- Cover the liquid/smear with a cover slip and gently press down to remove air bubbles
- Wear gloves to ensure there is no cross-contamination of foreign cells
How to prepare a microscope slide using a solid specimen:
- Take care when using sharp objects and wear gloves to prevent the stain from dying your skin
- Use scissors to cut a small sample of the tissue
- Peel away or cut a very thin layer of cells from the tissue sample to be placed on the slide (using a scalpel or forceps)
The tissue needs to be thin so that the light from the microscope can pass through - Apply a stain
- Gently place a cover slip on top and press down to remove any air bubbles
Why does staining in light microscopy make structures within the specimen visible?
The dyes used absorb specific colours of light while reflecting others
What is differential staining in light microscopy?
Certain tissues absorb certain dyes, depending on their chemical nature. Specimens or sections are sometimes stained with multiple dyes to ensure the different tissues within the specimen show up
What happens to a stained specimen using a TEM?
The dyes used for staining cause the tissues to show up black or different shades of grey
Why are heavy-metal compounds commonly used as dyes in electron microscopy?
Because they absorb electrons well
Magnification calculation:
magnification = size of image ÷ size of real object
Magnification can be defined as:
The number of times larger an image is than the actual object
Resolution can be defined as:
The ability to distinguish separate points on an image as two separate objects
How are light microscopes limited by wavelength of light?
As light passes close to physical structures it is diffracted, meaning that light waves spread out
The closer the structures are to each other, the more the light waves overlap each other as they are diffracted
Points that are closer together than half the wavelength of visible light cannot be clearly distinguished from each other
Why is resolution higher in electron microscopes?
Because electrons have a smaller wavelength than visible light
The objects past which the electrons travel can therefore be much closer together before the diffracted beams overlap
Eukaryotic cell definition-
A type of cell that contains a nucleus along with membrane bound organelles
Prokaryotic cell definition-
A type of cell that does not contain any membrane bound organelles or a nucleus
Nucleus definition-
An organelle found in eukaryotic cells that stores the genetic information of the cell
as chromosomes and is surrounded by a membrane called the nuclear envelope
