Lab 1: Brightfield Microscopy Flashcards

60 practice question-and-answer flashcards based on Lab 1: Brightfield Microscopy to prepare for lab examinations.

What is a brightfield microscope?

A microscope that allows light rays to pass directly to the eye; it is the most common type of microscope used by students in beginning biology courses.

What are the three lens systems of a compound microscope?

The oculars (eyepieces), the objective lenses, and the condenser.

What is the ocular (eyepiece)?

The lens system at the top of the microscope. It usually has a magnification of $10\times$.

What are the objective lenses?

The lenses on the nosepiece that provide the primary magnification of the specimen.

What objective magnifications are commonly found on laboratory microscopes?

$10\times$ (low-power), $40\times$ (high-dry), and $100\times$ (oil immersion). Some microscopes also have a $4\times$ objective for rapid scanning.

What is the total magnification of a compound microscope?

Ocular lens magnification \times objective lens magnification.

What is the total magnification with a $10\times$ ocular and $10\times$ objective?

$100\times$.

What is the total magnification with a $10\times$ ocular and $40\times$ objective?

$400\times$.

What is the total magnification with a $10\times$ ocular and $100\times$ oil-immersion objective?

$1000\times$.

What is the condenser?

The lens system located under the stage that collects and directs light from the light source to the specimen.

Does the condenser affect magnification?

No. Unlike the ocular and objective lenses, the condenser does not affect the magnifying power of the microscope.

What does the diaphragm do?

It regulates the amount of light passing through the condenser to the slide.

What happens when the diaphragm is closed somewhat?

It reduces the light intensity and can improve image contrast.

What happens if the diaphragm is closed too much?

Contrast may improve, but numerical aperture is reduced, which reduces resolving power.

Where should the condenser be positioned for optimal viewing?

At its highest position, allowing the maximum amount of light to enter the objective lens.

What is resolving power?

The ability of a lens to completely separate two objects in the microscope field.

What determines the limit of resolution?

The wavelength of the light used and the numerical aperture of the lens.

What is numerical aperture (NA)?

A mathematical expression describing how the condenser concentrates and focuses light rays and how the objective lens accepts them.

What is the approximate limit of resolution for a typical light microscope?

About $0.2\,\mu\text{m}$.

What does a $0.2\,\mu\text{m}$ limit of resolution mean?

Two objects closer than $0.2\,\mu\text{m}$ apart would not be seen as two distinct objects.

How does increasing magnification relate to resolution?

Increasing magnification alone does not produce better resolution; magnification must be accompanied by sufficient resolving power.

What is the relationship between working distance and magnification?

As objective magnification increases, the working distance decreases.

What is the coarse adjustment knob used for?

Bringing objects into focus by moving the stage a considerable distance.

What is the fine adjustment knob used for?

Making small focusing adjustments to sharpen the image.

Which focusing knob should be used with the high-dry objective?

Only the fine adjustment knob.

Which focusing knob should be used with the oil-immersion objective?

Only the fine adjustment knob.

Why should the coarse adjustment knob not be used with high-dry or oil immersion?

Using it with those objectives can cause the objective lens and/or slide to break.

Why should you begin viewing a slide with the low-power objective?

It allows you to explore the slide and locate the object of interest before proceeding to higher magnifications.

What should you do before changing to a higher-magnification objective?

Center the object of interest in the field of view.

What is a parfocal microscope?

A microscope that maintains focus when the objective magnification is increased.

What is a binocular microscope?

A microscope with an ocular (eyepiece) for each eye.

What are diopter adjustments used for?

Adjusting the oculars so that each eye can obtain a sharp image when using a binocular microscope.

How are diopter adjustments made according to the manual?

Focus first with the right eye only. Then, without touching the focusing knobs, adjust the diopter ring for the left eye until a sharp image is seen.

Why is immersion oil used with the $100\times$ objective?

The oil has the same refractive index as glass. It forms a continuous lens system that limits loss of light due to refraction and improves resolution.

Where is immersion oil placed?

Between the specimen/slide and the $100\times$ oil-immersion objective.

Can the oil-immersion objective be used without oil?

No. An oil-immersion lens should never be used without oil.

What should you do before rotating the oil-immersion objective into position?

Bring the specimen into focus at a lower magnification, center the object of interest in the field of view, and place a drop of immersion oil on the slide.

What should you never do after placing immersion oil on the slide?

Do not rotate the high-dry ($40\times$) objective back into place after oil has been placed on the slide. Clean the oil completely before returning to a lower magnification.

Why is more light needed when using the oil-immersion lens?

Higher magnification requires more light; the iris diaphragm should therefore be opened wider.

What should be done with the diaphragm when moving to oil immersion?

Open the diaphragm wider to provide adequate light.

What four things can maximize resolving power when using the oil-immersion lens?

Use the appropriate blue filter to shorten the wavelength; keep the condenser at its highest position; do not close the diaphragm too much; and use immersion oil between the slide and $100\times$ objective.

How should a microscope be carried?

Hold it firmly with both hands, as illustrated in the manual. Do not attempt to carry more than one microscope at a time.

What should be done with the microscope's electrical cord?

Avoid snagging it or allowing it to hang off the workbench. When storing, wrap it and secure it with a rubber band or cord wrap.

What should be used to clean microscope lenses?

Only lint-free, optically safe lens tissues and the type of cleaning liquid recommended by the instructor.

How can you tell whether an ocular is dirty?

Rotate the eyepiece while looking through the microscope. If objects appear to move with the rotation, the dirt/artifacts are associated with the ocular.

How should objective lenses be cleaned?

Use a piece of lens tissue moistened with an approved solvent or water. At the end of the lab, wipe immersion oil off the oil-immersion lens.

Why must microscope lenses be kept clean?

Dust, oil, and other contamination prevent the lenses from achieving their intended degree of resolution.

What should be done with the microscope stage after use?

Remove the slide, clean or disinfect the stage according to the instructor's directions, and adjust the mechanical stage so it is centered and as low as possible.

What is the correct order for increasing magnification?

Start with low power, then move to high-dry, and then to oil immersion when needed.

What are common student errors that can result in a dirty or broken microscope?

• Using the wrong focusing knob with high-dry/oil objectives

• Getting immersion oil on the 40× objective

• Not cleaning oil from the oil-immersion lens

• Using the wrong materials/solvents to clean lenses

• Carrying the microscope improperly

• Letting the electrical cord snag or pull the microscope off the bench

What is the best adjustment for increasing image contrast in low-power magnification?

Closing down the diaphragm somewhat.

A microscope that maintains focus when magnification is increased is called what?

Parfocal.

Which objective provides the highest magnification?

The oil-immersion objective ($100\times$).

Which objective provides the second-highest magnification?

The high-dry objective ($40\times$).

Which objective provides the lowest magnification among the standard three?

The low-power objective ($10\times$).

Which objective has the shortest working distance?

The oil-immersion objective.

Which microscope part collects and focuses light onto the specimen?

The condenser.

Which microscope part regulates the amount of light passing through the lens system?

The diaphragm.

What is the purpose of the light source?

It supplies the light used to illuminate the specimen.

What are the two adjustments that can be made to the condenser?

Its position can be adjusted up and down, and the diaphragm can be adjusted to regulate the amount of light.