Microscopy & Simple Stains

Your Light Microscope TAKE CARE OF YOUR SCOPE: GETTING YOUR SCOPE OUT: Your responsibility to take care of your scope and learn to use it properly. GETTING YOUR SCOPE OUT: When transporting your scope, always hold it with one hand under the base, and one hand around the arm. PUTTING SCOPE AWAY: Whenever your are getting ready to put your scope away: • Use alcohol swab to clean stage and lens paper to clean lenses. • Shortest lens (the one with the red band) should be facing down toward stage. • Use course focus to position stage as low as it can go. ARM BASE

Bright-field Compound Microscope Microscopy – Light Microscopes Bright-field Compound Microscope

Microscopy – Light Microscopes Bright-Field Microscope Light microscope produces a dark image against brighter background. Commonly used to view stained cells. Simple microscopes have single magnifying lens (like a magnifying glass). Compound microscopes have two sets of lenses for magnification. Lens closer to the eye = ocular lens (magnifying power of 10x). Lenses closer to the object being viewed = objective lens. (Most light microscopes used in biology have three or four objective lenses). Ocular lenses Objective Lenses OBSERVATION OF MICROORGANISMS

Objective Lenses Scanning Objective Lens Has red band around it. Magnifies objects 4x. Total magnification = 40xTM This lens is of no use to us in looking at bacterial stains.

Objective Lenses Low Power Objective Lens Has yellow band around it. Magnifies objects 10x. Total magnification = 100xTM Start with this lens to get your bacterial smear into crisp focus. You will not see individual bacteria with this lens, you are just using it to focus so that you can move up to the next magnification.

Objective Lenses High Dry Objective Lens Has blue band around it. Magnifies objects 40x. Total magnification = 400xTM Move up to this lens after focusing your smear at 100xTM. You will not be able to clearly see individual bacteria with this lens. Just get the image in focus as much as possible.

High Dry Objective Lens After you focus the image at 400xTM, you need to cover this lens with a finger cot so that it does not get oil on it. Do not move the focus knob or the stage when placing the finger cot on the high dry lens or you will take the image out of focus! NEVER use coarse focus with high dry or oil immersion lenses!!!

Oil Immersion Objective Lens Has black and a white band around it. Magnifies objects 100x. Total magnification = 1000xTM Move up to this lens after focusing your smear at 400xTM and covering the 400xTM lens with a finger cot. NEVER use coarse focus with high dry or oil immersion lenses!!!

Microscopy – Electron Microscopes Two types: Both huge, expensive machines. Transmission Electron Microscope (TEM): 2-D image: Transmission Electron Micrograph Scanning Electron Microscope (SEM): 3-D image: Scanning Electron Micrograph A transmission electron micrograph of Escherichia coli (E.coli). SEM AIDs virus attacking T4 lymphocyte

Wrap-up When putting away scope: Make sure lenses are clean (wipe with alcohol swab) Have scanning power lens in position (4x) Make sure stage is in lowest position Return scope to its proper “address”