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Dynamic Genome > Experiment 4: Microscopy > Flashcards

Experiment 4: Microscopy Flashcards

1
Q

What is the purpose of this lab?

A

To observe Arabidopsis leaves using fluorescence and the dissecting scope.

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2
Q

Explain important thing about the Binocular Eyepiece Tube

A

This assembly holds the ocular lenses; it is designed to allow adjustment of the distance between the oculars and the focus of one of them. The tube is held together by a set screw WHICH SHOULD REMAIN TIGHTENED AT ALL TIMES.

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3
Q

Explain important things about the Revolving Nose Piece

A

The lenses are threaded into this structure and the nosepiece allows objectives of different power to be clicked into position of the optical path.

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4
Q

Should you PUSH or PULL the objective in the revolving nose piece?

A

No

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5
Q

Explain important things about the Arm and Base

A

They support the optical system. The light source is built into the base and off to the side of the base are the combined on and ooff switch and rheostat, which controls brightness.

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6
Q

What is the function of the Rheostat?

A

Controlling the brightness.

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7
Q

What is the Stage?

A

The plane surface on which slides are placed for viewing.

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8
Q

What is the function of the Focus Controls?

A

They raise and lower the stage and thus vary the distance between the specimens and the objective lens (this distance is referred to as the working distance). The larger knob that moves the stage greatly when rotated is called the COARSE FOCUS and the smaller knob that moves the stage slightly is called the FINE FOCUS.

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9
Q

The coarse focus should only be used when working with _____ objectives.

A

Low Power

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10
Q

Should the stage be moved upwards or downwards when utilizing coarse focus?

A

No, this could result isn damaging the object and/or slide.

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11
Q

What is the function of the Condenser Mounting Bracket?

A

The bracket holds the assembly and can be moved up and down with the condenser focus control.

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12
Q

What is the function of the Mechanical Stage Assembly?

A

To move the slide front and back. MF

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13
Q

What is the Mechanical Stage Assembly consisted of?

A

Movable Slide Clamp (this device is spring-loaded and grasps the slide by its EDGE)- Stage Motion Controls (Two knobs that project downward from the stage and function in moving the slide side to side and back and forth)

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14
Q

What are the 4 Optical Components of a microscope?

A

Light Source-Condenser Lens-Objective Lens-Ocular Lens. L-COO

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15
Q

Define the Light Source

A

Light from the sub-stage bulb is diffused by a frosted glass surface located above the bulb.

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16
Q

What will excessive brightness from the light source result in?

A

Loss of image contrast

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17
Q

What is the function of the condenser?

A

The function of the condenser lens is to focus the maximum amount of light from the light source upon the specimen.

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18
Q

What is the relationship between the condenser diaphragm and the contrast of the specimen?

A

As the condenser diaphragm is CLOSED, the specimen contrast INCREASES. (INVERSE Relationship)

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19
Q

What is an example of specimens hat have a low inherent contrast?

A

Living Cells

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20
Q

What is the function of the objective lenses?

A

Accomplish the first stage of magnification in the compound microscope.

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21
Q

How should lens cleaning be done?

A

It should exclusively be done with LENS PAPER.

22
Q

Define Numerical Aperture (NA)

A

The measure of the light gathering capacity of the objetive lens.

23
Q

What is the relationship between the value of NA and the resolving power of the lens?

A

The greater the value of NA, the greater the resolving power of the lens.

24
Q

Define the Resolving Power

A

The minimum distance that two points can be spatially separated and still be seen as two individual parts.

25
Q

Define Depth of Field

A

A term that relates to the thickness of the microscopic field that is in usable focus at a particular time.

26
Q

What is the relationship between Depth of Field and Magnification?

A

Depth of field decreases with the increase of magnification.

27
Q

When should lenses with a thicker depth of field be used?

A

When locating smaller specimens or when studying the spatial relationships of the larger structures where high resolution is not important.

28
Q

When should lenses with a thinner depth of field be used?

A

To study the relationships of structures where high resolution is more important.

29
Q

Define Working Distance

A

Working distance for a focused objective is the distance between the tip of that objective and the specimens being viewed.

30
Q

What is the relationship between the working distance and the power of the objective?

A

Working distance DECREASES with an INCREASE in the power of the objective.

31
Q

How is the problem of potential contact of objectives and slides averted?

A

Observations on a new slide should begin with LOWER power objectives.

32
Q

What is a benefit of the long working distance of a low power objective?

A

It eliminates the need for lowering the stage when a new slide is selected.

33
Q

What is the function of ocular lenses?

A

To use the real image formed by the objective lens as its object and produce a virtual image that the viewer sees.

34
Q

Define Contrast

A

Relates to the ability of the viewer to distinguish an object from its background.

35
Q

What techniques will be used in this experiment to increase the contrast in this lab?

A

Closure of the Condenser Diaphragm-Differential Staining. CD

36
Q

Define Differential Staining

A

Involves the use of stains (dyes) which bind to various components of the specimen to increase contrast. Stains usually bind specifically to particular substances in cells or tissues with a unique chemical makeup.

37
Q

What are the two stains that are going to be used in this experiment?

A

Aceto-Orcein; 12KI

38
Q

Explain everything about Aceto-Orcein

A

It’s a natural stain that comes from an insect. It binds to chromatin and stains it red; so, it is useful for identifying nuclei.

39
Q

Explain everything about 12KI

A

It is a mix of iodine and potassium iodide. It works through a chemical reaction between the iodine and the starch granules of green plants. The resulting complex has a purple/blue color.

40
Q

When utilizing coarse focus, what are some things to note; if not done properly, what are some possible consequences?

A

It should only be used when working with low power objectives. The stage should NEVER be moved upward when looking through the microscope- doing so could damage the objective and/or slide. Such damage can be prevented by watching, from the side of the stage to see that the slide will not touch the objective when the stage is raised with the coarse focus.

41
Q

For our microscopes, how/when is the condenser properly focused?

A

When the condenser assembly is near the uppermost range of adjustment.

42
Q

How can condensers cause harm to a live specimen? How is this problem averted?

A

The small spot of illumination created by the condenser is also hot in addition to being bright. To avoid this problem, make the observations quickly, or lower the light intensity with the brightness control knob (DO NOT LOWER THE CONDENSER).

43
Q

What does the condenser diaphragm do?

A

Regulates the diameter of the light beam passing through the condenser.

44
Q

How should the rotating nosepiece be used? If it is not used in the proper manner, what happens?

A

It should be done with the use of the “grooved” surface on the edge of the revolving nosepiece and NOT BY PUSHING OR TUGGING ON THE OBJECTIVES THEMSELVES. Doing so could loosen the objectives and eliminate the parfocality of the microscope- this could result in scope or slide damage.

45
Q

How much does the ocular lens magnify an image?

A

10X

46
Q

What is an advantage of the binocular viewing system?

A

The microscopist has a greater depth perception of the selected field of view.

47
Q

What is a disadvantage of the binocular viewing system?

A

The oculars need to be adjusted to accommodate different users.

48
Q

What are some possible problems associated with people that have difficulty using binocular microscopes?

A

The microscope has been damaged by a previous user; the viewer is holding his/her eyes too close to the oculars. The solution is to back your eyes off slightly form the oculars and redo the two adjustments.

49
Q

What can be seen in the light microscope is limited by the _____.

A

Size of the Object.

50
Q

What are two useful microscopy techniques?

A

Making of Wet Mounts- Perfusion of Stains. WP

Dynamic Genome (9 decks)

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