Laboratory Tips and Microscope Use

Setting Up: Microscope Carrying and Bench Layout

Always hold the arm of the microscope while supporting the

base with your other hand

Microscope

Lab

Manual

Student B

Microscope

Lab

Manual

Student A

Work Area

Test tubes

Microscope Parts and Tips for Use

Close the iris diaphragm until it gets too dark in order to improve contrast

Keep the condenser lens in the full “up” position by rotating the condenser adjustment knob.Close the iris diaphragm until it gets too dark in order to improve contrast

Adjust the interpupillary distance for your eyes by widening/narrowing the ocular distance.

Always lower the stage before mounting a new slide. Be sure to always begin by getting the specimen focused and centered using the 4X objective first. If you can’t see the specimen at 4X, you’ll never see it at higher powers.

Magnification is Inversely Related to Field Diameter

Objective Ocular Lens Total Apparent Actual

Lens Power Power Magnification Field Diameter Field Diameter

4X 10X 40X

10X 10X 100X

40X 10X 400X

100X 10X 100X

1800 µm

450 µm

180 µm

45 µm

Refraction Becomes Significant When Using a High Power (100X) Objective

When the objective lens is very close to the slide, the loss of light, resolution, and clarity are so severe that immersion oil is needed to prevent this. Immersion oil has the same index of

refraction as glass so the light path continues straight from the slide to the lens.

• Resolution is the ability of microscope lenses to distinguish between two points very close together.

Resolution (Resolving Power) and Microscope Limits

• A microscope with a resolving power of 0.4 nm can distinguish between two points ≥ 0.04 nm apart. If they are closer than 0.4 mm, they will be seen as one point.

• Resolution wavelength ()

• How does the color (wavelength) of light affect resolution?

• Resolution is good when it is a very low value

Electron wave,

R O Y G B V

Electrons,

Resolution Limit With Light Also Limits Max. Useful Magnification

R O Y G B V

Limited resolution with light effectively

limits magnification

to 1000X

Electrons,

Light micrograph at 1000X

Transmission electron micrograph at 10,000X

Scanning electron micrograph at 10,000X

Much better resolution allows for high, sharp,

magnification at ~ 30,000X

Using the 100X and Cleaning Up Afterwards

NEVER go back to the 40X objective after setting up the 100X on oil. You will get oil on the 40X, which ruins it. If you need to refocus at a lower power, clean off the

lens and the slide, or swing to the 10X, focus, and go back.

1. Find your specimen, get it centered and in focus using the 4X, then 10X, then 40X. Don’t touch the focus after this.

2. Swing the 100X carefully into position over the light hole3. Use the fine focus knob to get the focus sharp using the 100X.

Removea sheet

Clean the 100X, your slide, and excess oil on the stage

Before Putting Away the Microscope….

Remember the Following Steps For Lab…

1. Put away all inessential books and coats under the lab bench; food and drinks stay packed up or are left outside the classroom.

2. Put on your lab coat and tie back your hair, if necessary

3. Disinfect the lab bench top in your area.

4. Wash your hands.

5. Take notes on any information added by the teacher regarding the day’s lab session. Plan your day’s activities with your lab partner.

6. Together with your lab partner, get the necessary supplies or incubated cultures and begin work.

7. When done, put away microscopes, dispose of cultures, & put away supplies properly. Make sure your lab drawer and the supply area are neat for the next lab session.

8. Disinfect your lab desk.

9. Remove your lab coat, hang on the rack.

10. Wash your hands.

11. Push in your chair, and you’re done!