2-D image: Transmission Electron Micrograph
A transmission electron micrograph of Escherichia coli (E.coli).
Scanning Electron Microscope (SEM):
3-D image: Scanning Electron Micrograph
• 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.
Welcome to Lab!
Getting Ready for Today’s Lab • Stash backpacks in cubbies on right side of room. • • Choose your seat. Put on your “lab coat” and set goggles, a „dirty‟ copy of lab exercise/report #1 and your lab card on bench next to you. (The other copy of the lab exercise and report should be in your backpack. This is your „clean‟
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.
Contrast:
• • • What is it? Difference between the object and the background Easiest ways to improve contrast are to use dye and/or manipulate light
Microscopy – Light Microscopes
copy.)
Байду номын сангаас
•
TURN IN: Pre-Lab #1 BEFORE LAB BEGINS!
•
Come get the following handouts:
- “Lab Cards” that you will be using each lab for sign in and sign out. - Signature page.
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.
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:
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!!!
Resolution:
• • •
•
What is it? The ability to distinguish between objects that are close together. The optometrist‟s eye chart is a test of resolution at a distance of 20 feet. Limits maximum magnification
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.
Onion Cells
Elodea
Cheek Cells
4. Cheek cell Wet mount using NaCl Try to view, then add stain. Contrast!
Before you get started, I want you to prepare a slide with a bacterial sample.
Bright-field Compound Microscope
AT THIS POINT, GO GET A SCOPE AND SIGN IT OUT.
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).
Objective Lenses Ocular lenses
Lenses closer to the object being viewed = objective lens. (Most light microscopes used in
biology have three or four objective lenses).
BASE
Microscopy
General Principles Magnification:
• • • What is it? Apparent increase in size of an object. Indicated by a number and “X”, which is read “times”.
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.
Remember the term parfocal?
NEVER use coarse focus with high dry or oil immersion lenses!!!
Microscopy – Electron Microscopes
Two types: Both huge, expensive machines. Transmission Electron Microscope (TEM):
End of Today’s Lab IMPORTANT!!!! Before next week, print out two copies of all labs (exercise & report), so that you have a “clean” copy and a “dirty” copy and keep these in a binder that you bring with you to class/lab. Each week I will tell you which exercises you will need to have out in lab. •
TAKE CARE OF YOUR SCOPE: