Topic 1

Microscopes

Light Microscopes

How do they work? Pass light through thin specimen.

Benefits Limitations

Light Microscope Compound Light

Microscope Optical lens

10x Objective lens

4x scanning power 10x 40x 100x Oil immersion

Total magnification = optical x objective mag.

Measuring length?

Look at the field of view

Using a clear ruler… 3.5 mm

OBJ 3

Measuring length?

Estimate How big is each cell?

OBJ 3

Magnification – Increase in Apparent Size

Resolution – A Measure of Clarity

Magnification vs. Resolution

Magnification - increase in apparent size Resolution - measure of clarity

Ability to show two objects as separate Human eye has limit of 0.1 mm Light microscope has limit of 0.2 micrometer (at

any magnification) Limited by light wavelength. Longer wavelengths = lower resolution

Fluorescence Confocal Microscopy Type of light microscopy: Fluorescence is used to increase resolution.•Specific molecules are tagged with fluorescent dyes•Allows you to see the specific structures of the cell•Here: cytoskeletal filaments, nucleus, etc.

Fluorescence Confocal Microscopy

Type of light microscopy.

Electron Microscopes

Use a beam of electrons instead of light

Benefits Resolution = 2nm Magnification above 50

million x Limitations SEM and TEM

OBJ 2

Transmission Electron Microscope (TEM)

Spinach Cell Chloroplast

OBJ 2

TEM

Mitochondrion

OBJ 2

TEM

Red Blood Cell in a capillary of the pancreas

OBJ 2

Scanning Electron Micrscope (SEM)

OBJ 2

SEM

OBJ 2

SEM

OBJ 2

SEM

OBJ 2

SEM

OBJ 2

Comparing microscopes

Light SEM TEM

Source Light Beam Electron Beam Electron Beam

Resolution limit

2 micrometer (smallest bacteria)

2 nanometer(0.002 micrometer)

2 nanometer(0.002 micrometer)

Properties of Specimen

Thin Small enough to fit in chamber of scope

Ultra Thin

Other Can be alive, allows you to see processes and movement

Coated in metal atoms and put in a vacuum (dead)

Coated in metal atoms and put in a vacuum (dead)