Mod 3 Test

Question 1

A micrometer is defined as:

Answer 1

10 to the -6

Question 2

Resolution and contrast are two critical factors that influence your ability to see an object. Explain each.

Answer 2

Resolution refers to the distance between two objects at which the objects still can be seen as separate. Poor or low resolution means two (or more) objects may appear as one.

Contrast is the difference in light absorbance between two objects. Poor contrast gives a high background and makes the visualization of multiple objects difficult. For instance, trying to identify 2 dark colored objects at night (low light = low contrast) versus the same 2 objects in the middle of a sunny afternoon (bright light against 2 dark objects = high contrast).

Question 3

Assuming a fixed ocular, identify the part of the microscope you would adjust to enhance the magnification of a sample.

Answer 3

Objective
Only the oculars (eyepiece) and the objectives contribute to the magnification of the sample. Since the eyepiece is fixed, only the objectives could be altered.

Question 4

What is the total magnification (relative to your eye) of a sample imaged with a 60x objective and a 10x eyepiece? Show your math.

Answer 4

60 x 10 = 600x magnification

Question 5

True or False: Staining is often required to image a cell that is adherent and flat (thin).

Answer 5

True
Adherent, flat and unstained cells are almost invisible due to the limits on both resolution and contrast. Therefore, cell staining is often required to adequately image the sample.

Question 6

Which of the following could NOT be seen clearly by the unaided eye? Select all that apply.
Protozoa with diameter of 250 μm
Virus with a diameter of 0.2 μm
Skin cell with diameter of 1500 μm
Bacteria with diameter of 24 μm

Answer 6

Virus with a diameter of 0.2 μm
Bacteria with diameter of 24 μm
The unaided eye can, on average, clearly resolve objects > 200 μm

Question 7

Label the following unmarked microscope components (numbered arrows) by matching it with the components provided (letters).

A. Stage
B. Fine Adjustment Knob
C. Iris Diaphragm
D. Neck
E. Condenser Lens
F. Eyepiece
G. Objective
H. Base
I. Coaxial Controls

Answer 7

1 F
2D
3B
4G
5A
6H

Question 8

This type of microscope is best suited for visualizing GFP, RFP, and YFP proteins.

Answer 8

Fluorescence

Question 9

This type of microscope can provide detailed images of live cells without staining.

Answer 9

Phase-Contrast

Question 10

This type of microscope is used to greatly increase the contrast between samples and background by reflecting light off of the specimen.

Answer 10

Dark Field

Question 11

This type of microscope uses neither halogen nor UV light sources but rather lasers to illuminate stained cells in high resolution.

Answer 11

Confocal

Question 12

Identify what type of electron microscope was used to capture the following image and explain your choice.

Answer 12

The image was captured using a Scanning Electron Microscope (SEM). The above image shows the trademark ‘shell’ image (no subcellular organelles are visible) reminiscent of SEM. Only TEM is capable of visualizing subcellular substrucutres.

Question 13

Gram-Positive cells appear [answer1] in color due to a [answer2] peptidoglycan layer in the cell wall.

Answer 13

1. Purple
2. Thick

Question 14

Gram-Negative cells appear [answer1] in color due to a [answer2] peptidoglycan layer in the cell wall.

Answer 14

1. Pink
2. Thin

Question 15

True or False: The distinguishing characteristic of Gram-Negative bacteria is the presence of LPS in the outer membrane.

Answer 15

True

Question 16

Name one substance capable of chemically fixing cells to a slide.

Answer 16

Any of the following are true: Paraformaldehyde, ethanol or methanol.

Question 17

You want to observe the size and shape of a cell. What is the easiest staining technique that you could perform? Name at least one dye you would use during this process.

Answer 17

Simple stain. You could use any of the following: methylene blue, crystal violet, safranin or fuschin.

Question 18

You suspect a patient may have TB. Once a sample has been obtained, it is sent off to the lab for an acid-fast stain. If the patient were infected with TB, describe what you would expect to see on the stained slide.

Answer 18

You would expect to see red cells (TB+) on a blue background (TB negative).

Question 19

True or False: If a patient is suspected of having malaria, a Giemsa stain would be an appropriate differential test to perform.

Answer 19

True
Giemsa stains are often used in the clinical setting to aid in the diagnosis of blood parasites.

Question 20

Identify what type of electron microscope was used to capture the following image and explain your choice.

Answer 20

The above image is captured via a Transmission Electron Microscope (TEM). Even at 20nm resolution (inset image), subcellular substructures are still visible. The image lacks the outside ‘shell’ only appearance of SEM.

Question 21

A simple stain has been attempted on an unknown sample with no success. Given this property, which stain should be attempted next?

Answer 21

Acid-fast staining

Question 22

Select the type of microscopy used to generate the image:

Answer 22

Fluorescence microscopy

Question 23

As the lead technician in the Microbiology Department’s core Microscopy lab, you are given a list what different scientists would like to observe in their samples. You must match these samples with appropriate type of microscopy.

1. A slide stained withcrystal violet simple stain to observe the arrangement of cells
2. Theswimming motility of living microalgae cell
3. An unstained samplethat cannot be visualized with bright field because microorganism absorbs very little light
4. The localization of a proteinof interest under experimental conditions in a cell
5. Builda 3-dimensional image of a cell

A. Bright field
B. Phase contrast
C. Dark field
D. Florescence
E. Confocal

Answer 23

1A
2B
3C
4D
5E