Microscopy Chapter 3 Biology Mr. Gilbertson

DISCOVERY OF CELL PRECEDED BY THE INVENTION OF THE MICROSCOPE ROBERT HOOKE - NAMED “CELLS” BECAUSE THEY LOOKED LIKE LITTLE ROOMS FOUND IN A MONASTERY ABOUT 1665 –LOOKING AT CORK BARK FROM MEDITERRANEAN CORK OAK –ALSO OBSERVED STEMS OF ELDER, CARROT, AND FERN –ALL WERE FOUND TO BE SIMILAR –ACTUALLY HE OBSERVED ONLY DEAD CELLS ANTOINE VAN LEEWENHOEK - –DUTCH LENS AND MICROSCOPE MAKER –FIRST TO OBSERVE LIVING CELLS ABOUT 1675

Looking at Cells Cells are too small to be viewed with the naked eye. (most smaller than a grain of sand)Cells are too small to be viewed with the naked eye. (most smaller than a grain of sand) Requires the use of a microscope to view.Requires the use of a microscope to view. Measurement is made using the SI system.Measurement is made using the SI system. –Based on powers of ten – like the money system –Uses common prefixes – makes conversion easy –Truly international system Micrometer is the unit used for cell size (one millionth of a meter, 1000 th of a mm)Micrometer is the unit used for cell size (one millionth of a meter, 1000 th of a mm)

Microscopes Light microscopes – also known as compound microscopesLight microscopes – also known as compound microscopes –Use light and lenses to focus images of very small objects. –Maximum magnification about 2000X Electron MicroscopesElectron Microscopes –Use a beam of electrons focused by magnetic fields to form images of objects. –Two types – TEM (transmission electron microscope) mag. up to 200,000XTEM (transmission electron microscope) mag. up to 200,000X SEM (scanning electron microscope) max mag 50,000XSEM (scanning electron microscope) max mag 50,000X

Microscopic Images Usually labeled with the type of microscope used.Usually labeled with the type of microscope used. Magnification – making the apparent image larger than the actual objectMagnification – making the apparent image larger than the actual object Resolution – a measure of the clarity of image (ability to distinguish fine detail)Resolution – a measure of the clarity of image (ability to distinguish fine detail) As magnification increases the resolution decreases, ultimately determined by the wavelength of light usedAs magnification increases the resolution decreases, ultimately determined by the wavelength of light used Electron microscopes can achieve greater magnifications because they operate at shorter wavelengths.Electron microscopes can achieve greater magnifications because they operate at shorter wavelengths.

Viewing the Microscopic World

The Simple Microscope Hand lens or a magnifying glass is a simple microscope.Hand lens or a magnifying glass is a simple microscope. Used to magnify, allows us to see fine details of structureUsed to magnify, allows us to see fine details of structure Power of Magnification is the number of times the apparent size of an object is increased. (10X – ten times)Power of Magnification is the number of times the apparent size of an object is increased. (10X – ten times) Focusing – the process of moving the position of the lenses to bring the image into clear focusFocusing – the process of moving the position of the lenses to bring the image into clear focus

The Compound Light Microscope Max magnification about 1000XMax magnification about 1000X As magnification increases often resolution decreases, must use more precise light source.As magnification increases often resolution decreases, must use more precise light source. Eyepiece – usually 10XEyepiece – usually 10X Objective lenses – 4X scanning, 10X low, 40X high, 100X oil immersionObjective lenses – 4X scanning, 10X low, 40X high, 100X oil immersion

A Compound Light Microscope

Parts of a Light Microscope

Light train is made up of the condenser, diaphragm, slide, specimen, objective, body tube, and eyepiece.Light train is made up of the condenser, diaphragm, slide, specimen, objective, body tube, and eyepiece. Coarse adjustment, brings the sample into basic focus.Coarse adjustment, brings the sample into basic focus. Fine adjustment brings sample into clear focus (hopefully).Fine adjustment brings sample into clear focus (hopefully). Always start out on low power and work up.Always start out on low power and work up. To view sample under oil immersion, oil must be used between objective and cover slip.To view sample under oil immersion, oil must be used between objective and cover slip.

Dissecting Scope Used to look at objects that are too big for a light microscope but to small to see clearly with the naked eye.Used to look at objects that are too big for a light microscope but to small to see clearly with the naked eye. As name implies used for dissection to view fine structure.As name implies used for dissection to view fine structure. Stereoscopic – giving a 3-D viewStereoscopic – giving a 3-D view Often with built in illumination and large stage for specimensOften with built in illumination and large stage for specimens

Research Microscope May have additional featuresMay have additional features StereoscopicStereoscopic 4 or more objectives4 or more objectives Built-in illuminationBuilt-in illumination Video adaptersVideo adapters Computer interfaces (Digitizers)Computer interfaces (Digitizers) Advanced optics (infrared, ultraviolet)Advanced optics (infrared, ultraviolet) Phase contrast opticsPhase contrast optics

Electron Microscopes Uses an electron beam and detectors to form an image of the specimen.Uses an electron beam and detectors to form an image of the specimen. Must be fixed (immovable) dead and coated with a reflective material (gold) in a vacuumMust be fixed (immovable) dead and coated with a reflective material (gold) in a vacuum Can magnify up to 2 million times.Can magnify up to 2 million times. TEM – Transmission Electron MicroscopeTEM – Transmission Electron Microscope SEM – Scanning Electron MicroscopeSEM – Scanning Electron Microscope STM – Scanning Tunneling MicroscopeSTM – Scanning Tunneling Microscope

TEM Photos Always in black and white, cells must be dead and fixed Electrons pass through the specimen giving a flat image – must be very thin.

SEM Photos False color can be added by a computer Electrons bounce off of the Object and give a 3-D image.

STM Images