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Microscopy

In this exercise you will become familiar with a bright field microscope that you'll be using throughout the semester. 

Bright Field Microscope Components

A: Eyepiece or ocular lenses. Magnify 10X

B: Nosepiece

C: Specimen holder

D: Condenser. Focuses the light onto the specimen

E. Aperture Iris Diaphragm. Controls the amount of light that enters the objective

F: Field iris diaphragm. Controls the amount of light leaving the source.

G: Light Source

H: X-axis control knob. Moves specimen left and right

I: Oil-immersion objective lens. Magnifies 100X

J: High-dry objective lens. Magnifies 40X

K: Low-power objective lens: Magnifies 10X

L: Power switch

M: Knob to control light intensity

N: Coarse adjustment focusing knob. Makes large changes in focus

O: Fine adjustment focusing knob. Makes small changes in focus

P: Y-axis control knob. Moves specimen up and down

Bright field microscope

Care and Use

  • Never slide a microscope across a bench surface.  Always carry a microscope with both hands.  One hand should be placed on the arm and the other should support the base.

  • Microscopes should be cleaned both before and after use.  Use ONLY lens paper and lens cleaner.  Kleenex, paper towels and even Kimwipes can scratch the lenses.

  • ONLY use oil when using the100X oil immersion lens.  DO NOT get oil on the other objective lenses.

  • Store microscopes with the 10X (low power) objective in position.  Bring the stage all the way up so that there is as little distance as possible between the stage and the objective lens.

  • DO NOT wrap the cord around the microscope.  Instead, fold the cord and place it between the arm and the stage.

  • Replace the dust cover before putting the microscope away

Terms and Definitions

  • Microscope: a device for magnifying objects that are too small to be seen with the naked eye.

  • Simple microscope: single lens magnifier

  • Compound microscope: employs two or more lenses

  • Parfocal: the objective lenses are mounted on the microscope so that they can be interchanged without having to appreciably vary the focus.

  • Resolving power or resolution: the ability to distinguish objects that are close together.  The better the resolving power of the microscope, the closer together two objects can be and still be seen as separate.

  • Magnification: the process of enlarging the size of an object, as an optical image. 

  • Total magnification: In a compound microscope the total magnification is the product of the objective and ocular lenses (see figure below).  The magnification of the ocular lenses on your scope is 10X. 

  • Objective lens X Ocular lens = Total magnification

    • For example: 

      • low power: (10X)(10X) = 100X

      • high dry: (40X)(10X) = 400X

      • oil immersion: (100X)(10X) = 1000X

  • Immersion Oil:  Clear, finely detailed images are achieved by contrasting the specimen with their medium.  Changing the refractive index of the specimens from their medium attains this contrast.  The refractive index is a measure of the relative velocity at which light passes through a material.  When light rays pass through the two materials (specimen and medium) that have different refractive indices, the rays change direction from a straight path by bending (refracting) at the boundary between the specimen and the medium.  Thus, this increases the image’s contrast between the specimen and the medium. One way to change the refractive index is by staining the specimen.  Another is to use immersion oil.  While we want light to refract differently between the specimen and the medium, we do not want to lose any light rays, as this would decrease the resolution of the image.  By placing immersion oil between the glass slide and the oil immersion lens (100X), the light rays at the highest magnification can be retained.  Immersion oil has the same refractive index as glass so the oil becomes part of the optics of the microscope.  Without the oil the light rays are refracted as they enter the air between the slide and the lens and the objective lens would have to be increased in diameter in order to capture them.  Using oil has the same effect as increasing the objective diameter therefore improving the resolving power of the lens.

Results

You will view and draw pictures of the pond water wet mounts observed on the 10 and 40X objective lenses.

dinoflagellate at 100x total magnificationpond water at 100x total magnificationpond water organism at 100x total magnificationpond water organism at 400x total magnificationpond water organism at 400x total magnificationpond water organism at 400x total magnification

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