microscopy Flashcards
Magnification =
Image size
——-———
Actual size
Magnification
How enlarged an image is
Resolution
How well you can distinguish two separate points to see in detail
Resolution: light-TEM-SEM
200nm-0.2nm-2nm
which microscope has the best resolution
TEM
Magnification light-TEM-SEM
1500-1,000,000-500,00
How does a light/optical microscope work?
Light passes from the condenser lens and through the specimen where certain wavelengths are filtered to produce an image. then light passes through the objective lens then eyepiece where magnification can be altered to be seen
What are the advantages of light microscopes?
They are cheap.
You can observe living organisms.
You can produce a coloured or stained image.
It is easy to use.
It is small and portable.
What are the advantages of electron microscopes?
More detail is seen because electrons have a smaller wavelength than light
They have a high magnification.
They have a higher resolution.
How does a transmission electron microscope TEM work?
Uses electromagnets to force a beam of electrons transmitted through the specimen to produce 2-D images. Denser parts of the specimen absorb more electrons which makes them look darker.
How does scanning electron microscope SEM work?
It Scans a beam of electrons across the surface of the specimen, this knocks of electrons from the specimen which are gathered in a cathode ray tube to form an image showing the surface of the specimen so it can be 3-D and has to be sliced to show the inside
Why can electron microscopes not observe living organisms?
Microscope has to have a vacuum to ensure the electron beams travel in straight lines
What are light microscopes used to observe?
Whole cells and tissues
What are electron microscopes used to observe?
Organelles
How can drawings be improved?
No shading
No overlapping
No arrowheads
Add a scale or magnification
Add a title
How to make a sample for a light microscope
Place stain at the edge of sample
Lower covers slip at an angle using a mounted needle
Use blotting paper to remove excess stain
Use multiple stains to improve contrast
Two different stains are used while making a slide to observe through a light microscope one stains the cytoplasm, one stains the nucleus, Why are they both needed?
Stain is needed for the cytoplasm for it to be visible against the background and a stain is needed for the nucleus to differentiate it from the cytoplasm
How to view a sample through a light microscope
- Select the lowest powered objective lens
- Use the course adjustment knob to bring the stage up to just below the objective lens
- Look down the eye piece and use the course adjustment knobs to move the stage down until image is roughly in focus
- Adjust the focus with the fine adjustment knob until there is a clear image of the slide
- If greater magnification was required, refocused the microscope using a higher powered objective lens
What microscope would you use to observe the inside of a mitochondria and why 2 mark
An electron microscope because a high resolution and magnification is needed to study internal structures of mitochondria
How to put a stain on a wet mount side
Put a drop of stain next to one edge of the covers sip and a piece of paper towel next to the opposite edge. The paper towel will draw the stain under the slip across the specimen.
Safety precautions
Wear goggles/ gloves
take care not to break the glass
take care of the sharp tools
Identify the microscope used to produce this image (blank and white and 2D)
Tem/ transmission electron microscope
What is the difference between the images of light microscope and electron microscope
It has higher resolution so individual organelles can be observed better
Which microscope has the best resolution and magnification
TEM
How to measure the diameter of the nucleus of a cell using a light microscope
Use the eyepiece graticule and calibrate it by aligning two scales to find the length of one division
Measure the diameter using graticule units
And use to calculate actual diameter
How can improvements be made to making a sample slide
Use a sharp blade to cut sample thin so individual cells can be observed
Choose a thinly sliced sample so maximum light penetrates sample
Squash slide to avoid air bubbles
Benefits of using stains in microscopy
Higher contrast
Internal organelles more visible
So clearer image is observed
Why is it important to use different stains when observing blood smear
To differentiate cells, organelles and have a higher contrast especially for white blood cells so they can be counted
Examples of stains
Methylene blue
Giesma stain
Haematoxylin
Eosin
What is a stage micrometre?
A microscope slide with an accurate scale that is used to work out the value of the divisions on the eye piece graticule at particular magnification
What are scanning electron microscopes used to observe?
The surface of cells
