Observing Cells Under the Microscope

Brightfield (Light) Microscopes

Light microscopes were the first to be invented

• Light microscopes use glass lenses and light to magnify cells
• Allowed scientists to see the size, shape and contents of cells
• Led to the discovery that plants and animals have different cellular structures

1. Magnification: The image size produced by the microscope is much larger than the actual size of the object
2. Resolution: Ability to distinguish between two adjacent objects
3. Clarity: Ability to identify different structures

Parts of a brightfield (light) microscope

• Ocular lens: the eyepiece lens. It usually magnifies 10x.
• Objective lens: there are usually 3 objective lenses on a microscope that magnify 4x, 10x, 40x (sometimes there is a 4th lens that magnifies 100x).
• Stage: platform the microscope slide is placed on.
• Slide holder: holds the slide to the platform.
• Condenser: focuses the light onto the specimen.
• Condenser aperture lever: controls how far the light condenser is from the slide. Not all microscopes have this.
• Light / Illuminator: produces the light that passes through the specimen.
• Coarse adjustment: this is always the larger of the two adjustment knobs.
• Fine adjustment: the smaller of the two adjustment knobs. Used to make smaller changes to the focus.

Electron Microscopes

• Electron microscopes use beams of electrons instead of light to produce images.
• Compared to light, this beam has a very short wavelength, allowing it to pass between two cell features less than 0.2 µm apart.

Transmission Electron Microscopes (TEM)

• The beam of electrons is passed through a sample that has been sliced into very thin sections
• Produces a 2-dimensional image

Scanning Electron Microscopes (SEM)

• The beam of electrons is scans the surface of a sample
• Produces a 3-dimensional image

Scanning Tunnelling Microscopes (STM)

• A tiny electrical probe is placed near the surface of the sample,
• Produces a topographical image
• Used for atomic-level imaging

Determining Magnification & Size

Calculating Magnification

• The eyepiece usually magnifies the specimen 10x
• There are three objective lenses you can use:
• Scanning (4x)
• Low (10x)
• High (40x)
• To determine the total magnification, multiple the magnification of the eyepiece by the magnification of the objective lens used

Estimating Size

Brightfield Microscopy: Calibrating the Ocular Scale

How do you calibrate an ocular scale?

• The ocular scale is in the eye piece of the bright-field microscope
• You need a stage micrometer with known/defined division lengths in order to calibrate your ocular scale
• The ocular scale should be calibrated for each magnification

• You must align the ocular scale to the stage micrometer
• Next, determine the number of ocular divisions that span a single stage micrometer division
• In the above example 1 stage division = 10 ocular divisions
• If each stage division is 0.1 mm, then 10 ocular divisions also equals 0.1 mm
• If you need to convert between magnifications, then you need to determine the difference in magnification from the calibrated magnification to the one being used