Unit 7: Review Microscopy and Cytology Lab

Part I: Microscopy A. Parts of the compound light microscope

A
Examine the labeled microscope in your lab manual. Make sure you can identify all the parts.

B C E

F G H I J K L Continue Continue Side view Front view M N

Part I: Microscopy B. Principles of microscopy

Magnification the factor by which a specimen is enlarged. For each specimen viewed, the total magnification is calculated by multiplying the magnification of the ocular lens by the magnification of the objective. The light microscopes ocular magnification is 10X. Parfocal and parcentral imaging Parfocal imaging the ability of the microscope image to remain in focus when switching from one objective to the next. Fine adjustment using the fine adjustment knob might be needed, and the amount of light may need to be adjusted using the iris diaphragm lever and rheostat. Parcentral imaging the image will remain in the center of view when objective lenses are changed.

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Field of view
Field of view the microscope field that appears as a circle when viewing a slide through the ocular and objective lenses. As the magnification is increased, the diameter of the field will decrease. You will need to determine the width of this circle so that you can make estimates about the size of an object observed. A ruler is placed across the stage. You are viewing the image through the scanning objective (4X). The diameter of the field is 4.2 mm. Convert to m. Using the above information you can calculate the field of view for the low power (10X), high power (40X), and oil immersion objectives (100X). You should also record area of your field of view.

1 mm

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Depth of focus
Depth of focus also known as depth of field. It is the vertical distance or thickness of an object that remains in focus of one time. A 3-colored thread slide was focused. Note that one thread is in focus and the others are not. The clearly focused thread lies on top of the mount.

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Image orientation
Due to the optics involved with the light microscope, the image seen will be real, inverted, and magnified by the objective. The image is then magnified again by the ocular lens.

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40X

400X

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40X

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Resolving power
Resolving power the degree at which two adjacent points on a specimen are seen as separate detailed images.

Part I: Microscopy C. Other types of microscopes

Dissecting microscope
The dissecting microscopes are useful when working with larger specimens. The large distance between the objective and the specimen allows for viewing thicker mounts and dissections. The illumination is from both above and below. Unlike the compound microscope, the image you see is not inverted.

Contrast
Contrast how well the details of a specimen stand out against a background. Stains and lighting are used to increase contrast to see detail. See the phase contrast micrograph in your lab manual.

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Part II: Cytology A. Preparing wet mounts

To prepare a wet mount you need to a clean slide.

Part II: Cytology B. Eukaryotic cells

Elodea leaf wet mount

A leaf from a sprig of Elodea is used as the specimen for making a wet mount.

Cell wall Central vacuole location Cell membrane

Add a drop of water onto the center of a microscope slide. Then take a thin sample of your specimen and place it in the drop of water.

Examine the parts of the cell in this micrograph.

Cytoplasm
Add a cover slip to your prepared slide.

Chloroplasts

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Onion leaf cell wet mount
A fleshy leaf from white onion was removed. The leaf was folded back to remove the thin layer of epidermis. A piece of the epidermis was placed on a microscope slide containing a drop of iodine. Examine the parts of the cell in this micrograph.

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Cell membrane
Human cheek cells wet mount The inside of the mouth was scraped with a toothpick. The scrapings were stirred in a drop of methylene blue stain. Examine the parts of the cell in this micrograph.

Cell wall

Cytoplasm Nucleus

Cell membrane

Cytoplasm

Location of central vacuole Nucleolus

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Part II: Cytology C. Prokaryotic cells

Bacteria have diverse shapes. The most common bacterial shapes include coccus (cocci, pl.), bacillus (bacilli, pl.), and spirillum (spirilla, pl.). Bacteria are stained with Grams stain. Pink to red color indicate Gram negative and purple to blue color indicate Gram positive.

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Bacillus Gram positive cocci

Gram negative spirilla

(The arrow shows an aggregate of about 3 bacilli.)

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Coccus Spirillum

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(You see clusters of spirilla in this view.)

(The arrows are pointing to single cocci. You also see clusters of cocci at other areas.)

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Part II: Cytology D. Models

C A Animal cell model #1 B
A. Cell membrane B. Golgi apparatus C. Mitochondria D. Cytoplasm E. Endoplasmic reticulum F. Nucleus G. Pair of centrioles

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Animal cell/Plant cell Model #2

A. Nuclear envelope B. Nucleolus C. DNA (chromatin) D. Nuclear pore

D E G F

B C

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A Plant cell Model #3
A. Plasma membrane B. Mitochondria C. Golgi apparatus D. Cell wall E. Endoplasmic reticulum F. Cytoplasm G. Central vacuole H. Chloroplasts I. Nucleus

B E F G C I H

End of Lab Review