Lecture 7: Electron Microscopy

What is the resolution limit of a light microscope?

0.2 micrometers (µm)

What is the resolution limit of an electron microscope (EM)?

0.2 nanometers (nm) or pico meters

Why do electron microscopes have a higher resolving power than light microscopes?

electrons have wavelengths about 100,000 times shorter than visible light

Who developed the first Transmission Electron Microscope (TEM)?

• Max Knoll

• Ernst Ruska

• in Germany 1931

What type of lenses do electron microscopes use?

Electromagnetic lenses

How do EM’s work

use electrons instead of light

How does the source of illumination differ between light microscopes and electron microscopes?

Light microscopes use visible light, while electron microscopes use electron beams

What are the magnification capabilities of an electron microscope compared to a light microscope?

EMs can magnify up to 10,000,000x or more, while light microscopes magnify 1,000-1,500x

How do Transmission Electron Microscopes work?

• has a focused beam of electrons to “see through” the specimens

• incredible internal detail

What is a key sample preparation requirement for TEM analysis?

Samples must be ultrathin to allow electrons to pass through, achieving electron transparency.

What is the main advantage of Scanning Electron Microscopy (SEM)?

• solves TEM’s slicing problem

• produces 3D images of the specimen's surface, ideal for studying external features

When was the first commercial Scanning Electron Microscope (SEM) introduced?

Around 1965

What is the appearance of TEM micrographs compared to SEM micrographs?

TEM provides 2D images showing internal structures, while SEM provides 3D images of surfaces

What does a TEM image reveal about the specimen?

Internal details like ribosomes, nucleoid regions, and cell wall layers.

Why are SEM images often artificially colored?

To highlight specific areas of the specimen for better visualization

What biological structures can be studied using TEM?

Structures as small as ribosomes and macromolecules.

What biological structures can be studied using SEM?

Surface features of intact cells, viruses, and microbial biofilms

What does negative staining in TEM help reveal?

• the texture of cell surfaces, viruses, and isolated organelles

• light (elections pass through)

• dark (dense area of the object)

How does TEM analysis need to be prepared?

• samples need to be sliced thin enough for e- to pass through

• this is electron transparency

What is an electron micrograph?

An image produced using an electron microscope.

What is the effect of magnification on electron microscopes compared to light microscopes?

EMs provide approximately 1,000-fold greater magnification than light microscopes.

How are beard hairs affected by shaving with an electric shaver compared to a razor blade?

Electric shavers cause ragged ends, while razor blades produce cleaner cuts.

What feature of Bacillus bacteria is clearly visible under SEM?

Terminal endospores

What is shown in SEM images of pollen grains?

The intricate external structures and patterns of pollen grains

What does a SEM image of E. coli show when adhering to human epithelial cells?

The formation of a biofilm, a sticky, stringy mass of cells.

What preparation is unnecessary for SEM compared to TEM?

Thin sectioning or slicing of the specimen

What is a common use of TEM in microbiology?

Examining the internal structure of pathogens like Bacillus anthracis

What colors are seen in raw electron micrographs?

Black and white; colors are added artificially for detail emphasis