Microscopy

Microscopy

Technique to study cells, tissues, and small organisms

Morphology

Size, shape, and structure of cells, tissues, and small organisms

Zacharias Janssen

Dutch maker of one of the earliest compound microscopes

Antonie van Leeuwenhoek

Maker of simple microscopes with one lens

Light

Visible electromagnetic radiation detected by the human eye

Optical Microscopy

Uses glass lenses to magnify and focus light

Scanning Probe Microscopy

Uses a mechanical probe to feel the surface of the specimen

Uses of Optical Microscopes

Illuminate, magnify, generate contrast, resolve, and capture images

Principle of Optical Microscopy

Uses lenses to focus light onto the specimen and collect the image

How Does a Light Microscope Work?

Light is focused onto the sample and magnified through lenses

Components of Light Microscopy

Light source, diaphragm, condenser, stage, focus controls, objectives, eyepiece, camera

Example Uses of Light Microscopy

Determining cell presence, cancer stage, subcellular structures, molecules, and proteins

Magnification

Combination of eyepiece and objective lenses

Resolution

Minimum distance at which two distinct points can be seen

Numerical Aperture

Measure of light-collecting and resolving power of a lens system

Refraction

Bending of light between different media

Refractive Index

Matching refractive indices to increase resolution

Limits of Resolution

Anything smaller than 200 nm cannot be resolved using light microscopy

Fluorescence Microscopy

Using specific spectra to detect fluorescent molecules

Epifluorescence Microscopy

Floods the entire sample with light to capture both in-focus and out-of-focus light

Confocal Fluorescence Microscopy

Exposes a specific part of the sample to light and uses a pinhole to focus emitted light

Electron Microscopy

Uses electromagnetic/electrostatic lenses to magnify and focus electrons

Transmission Electron Microscopy (TEM)

Images thin samples by passing electrons through the sample

Scanning Electron Microscopy (SEM)

Images thicker samples by detecting emission of secondary electrons from the surface

Disadvantages of electron microscopes

Bulkier, expensive, and cannot image live cells