MICROSCOPE

Question 1

Conventional bright-field microscopy and more specialized applications like fluorescence, phase-contrast, confocal, and polarizing microscopy are all based on the interaction of ______ with ______ and are used to reveal and study tissue
features.

Answer 1

light, tissue components

Question 2

Types of Light Microscopy:

Answer 2

Bright-Field Microscopy
Fluorescence Microscopy
Phase-Contrast Microscopy
Confocal Microscopy
Polarizing Microscopy

Question 3

With the ______, stained tissue is examined with ordinary light passing through the preparation.

Answer 3

bright-field microscope

Question 4

Bright-Field Microscopy

The microscope includes an ______ and mechanisms to move and focus the specimen.

Answer 4

optical system

Question 5

Bright-Field Microscopy

The optical components are the ______ focusing light on the object to be studied; the ______ enlarging and projecting the image of the object toward the observer; and the ______ (or ______) further magnifying this image and projecting it onto the viewer’s retina or a charge-coupled device (CCD) highly sensitive to low light levels with a camera and monitor.

Answer 5

condenser, objective lens, eyepiece, ocular lens

Question 6

Bright-Field Microscopy

The total magnification is obtained by multiplying the magnifying power of the ______ and ______.

Answer 6

objective, ocular lenses

Question 7

Bright-Field Microscopy

The critical factor in obtaining a crisp, detailed image with a light microscope is its ______, defined as the smallest distance between two structures at which they can be seen as separate objects.

Answer 7

resolving power

Question 8

Bright-Field Microscopy

The maximal resolving power of the light microscope is approximately ______ μm, which can permit clear images magnified ______-______ times.

Answer 8

0.2, 1000, 1500

Question 9

Bright-Field Microscopy

Objects smaller or thinner than 0.2 μm (such as a single ______ or cytoplasmic ______) cannot be distinguished with this instrument.

Answer 9

ribosome, microfilament

Question 10

Bright-Field Microscopy

Likewise, two structures such as mitochondria will be seen as only one object if they are separated by less than ______ μm.

Answer 10

0.2

Question 11

Bright-Field Microscopy

The microscope’s ______ determines the quality of the image, its clarity and richness of detail, and depends mainly on the quality of its ______. ______ is of value only when accompanied by high resolution.

Answer 11

resolving power, objective lens, Magnification

Question 12

Bright-Field Microscopy

Objective lenses providing higher magnification are designed to also have higher ______.

Answer 12

resolving power

Question 13

Bright-Field Microscopy

The eyepiece lens only enlarges the image obtained by the ______ and does not improve ______.

Answer 13

objective, resolution

Question 14

Bright-Field Microscopy

______, typically used for study of bright-field microscopic preparations, involves the conversion of a stained tissue preparation to high-resolution digital images and permits study of tissues using a computer or other digital device, without an actual stained slide or a microscope.

Answer 14

Virtual microscopy

Question 15

Bright-Field Microscopy

In this technique (______) regions of a glass-mounted specimen are captured digitally in a grid-like pattern at multiple magnifications using a specialized slide-scanning microscope and saved as thousands of consecutive image files.

Software then converts this dataset for storage on a server using a format that allows access, visualization, and navigation of the original slide with common web browsers or other devices. With advantages in cost and ease of use, it is rapidly replacing light microscopes and collections of glass slides in histology laboratories for students.

Answer 15

virtual microscopy

Question 16

Bright-Field Microscopy

The optical system has three sets of lenses:

Answer 16

condenser
Objective lenses
eyepieces or oculars

Question 17

Bright-Field Microscopy

The optical system has three sets of lenses:

The ______ collects and focuses a cone of light that illuminates the tissue slide on the stage.

Answer 17

condenser

Question 18

Bright-Field Microscopy

The optical system has three sets of lenses:

______ enlarge and project the illuminated image of the object toward the eyepiece. ______ with different magnifications routinely used in histology include ______ for observing a large area (field) of the tissue at low magnification; ______ for medium magnification of a smaller field; and ______ for high magnification of more detailed areas.

Answer 18

Objective lenses, Interchangeable objectives, X4, X10, X40

Question 19

Bright-Field Microscopy

The optical system has three sets of lenses:

The two ______ or ______ magnify this image another ______ and project it to the viewer, yielding a total magnification of ______, ______, or ______.

Answer 19

eyepieces, oculars, X10, X40, X100, X400

Question 20

When certain cellular substances are irradiated by light of a proper wavelength, they emit light with a longer wavelength—a phenomenon called ______.

Answer 20

fluorescence

Question 21

In ______, tissue sections are usually irradiated with ultraviolet (UV) light and the emission is in the visible portion of the spectrum.

Answer 21

fluorescence microscopy

Question 22

Fluorescence Microscopy

The fluorescent substances appear ______ on a ______ background. For fluorescent microscopy, the instrument has a source of ______ or other ______ and filters that select rays of different wavelengths emitted by the substances to be visualized.

Answer 22

bright, dark, UV, light

Question 23

Fluorescence Microscopy

Fluorescent compounds with affinity for specific cell macromolecules may be used as ______. ______, which binds both DNA and RNA, is an example. When observed in the fluorescence microscope, these ______ emit slightly different fluorescence, allowing them to be localized separately in cells.

Answer 23

fluorescent stains, Acridine orange, nucleic acids

Question 24

Fluorescence Microscopy

Other compounds, such as ______ and ______, stain specifically bind DNA and are used to stain cell nuclei, emitting a characteristic blue fluorescence under UV.

Answer 24

DAPI, Hoechst

Question 25

Fluorescence Microscopy Another important application of fluorescence microscopy is achieved by coupling compounds such as ______ to molecules that will specifically bind to certain cellular components and thus allow the identification of these structures under the microscope. Antibodies labeled with fluorescent compounds are extremely important in ______.

Answer 25

fluorescein, immunohistologic staining

Question 26

Phase-Contrast Microscopy ______ cells and tissue sections, which are usually ______ and ______, can be studied with these modified light microscopes.

Answer 26

Unstained, transparent, colorless

Question 27

Cellular detail is normally difficult to see in unstained tissues because all parts of the specimen have roughly similar optical densities. ______, however, uses a lens system that produces visible images from transparent objects and, importantly, can be used with living, cultured cells

Answer 27

Phase-contrast microscopy

Question 28

______ is based on the principle that light changes its speed when passing through cellular and extracellular structures with different refractive indices. These changes are used by the ______ to cause the structures to appear lighter or darker in relation to each other.

Answer 28

Phase-contrast microscopy, phase-contrast system

Question 29

Because they allow the examination of cells without fixation or staining, ______ are prominent tools in all cell culture laboratories.

Answer 29

phase-contrast microscopes

Question 30

A modification of phase-contrast microscopy is ______ with ______, which produces an image of living cells with a more apparent three-dimensional (3D) aspect.

Answer 30

differential interference contrast microscopy, Nomarski optics

Question 31

With a regular ______, the beam of light is relatively large and fills the specimen. Stray (excess) light reduces contrast within the image and compromises the resolving power of the objective lens. ______ avoids these problems and achieves high resolution and sharp focus by using (1) a small point of ______, often from a ______ and (2) a ______ with a ______ in front of the image detector.

Answer 31

bright-field microscope, Confocal microscopy, high-intensity light, laser, plate, pinhole aperture

Question 32

Confocal Microscopy The ______, the ______, and the ______ are all optically conjugated or aligned to each other in the focal plane (______), and unfocused light does not pass through the pinhole. This greatly improves resolution of the object in focus and allows the localization of specimen components with much greater precision than with the bright-field microscope.

Answer 32

point light source, focal point of the lens, detector’s pinpoint aperture, confocal

Question 33

Confocal Microscopy Confocal microscopes include a computer-driven mirror system (the ______) to move the point of illumination across the specimen automatically and rapidly. Digital images captured at many individual spots in a very thin plane of focus are used to produce an “______” of that plane. Creating such optical sections at a series of focal planes through the specimen allows them to be digitally reconstructed into a ______.

Answer 33

beam splitter, optical section, 3D image

Question 34

______ allows the recognition of stained or unstained structures made of highly organized subunits.

Answer 34

Polarizing microscopy

Question 35

Polarizing Microscopy When normal light passes through a ______ filter, it exits vibrating in only one direction. If a second filter is placed in the microscope above the first one, with its main axis ______ to the first filter, no light passes through.

Answer 35

polarizing, perpendicular

Question 36

Polarizing Microscopy If, however, tissue structures containing oriented macromolecules are located between the two ______ filters, their repetitive structure rotates the axis of the light emerging from the polarizer and they appear as ______ structures against a ______ background.

Answer 36

polarizing, bright, dark

Question 37

Polarizing Microscopy The ability to rotate the direction of vibration of polarized light is called ______ and is a feature of crystalline substances or substances containing highly oriented molecules, such as cellulose, collagen, microtubules, and actin filaments.

Answer 37

birefringence

Question 38

Polarizing Microscopy The utility of all light microscopic methods is greatly extended through the use of ______. Many features of digitized histologic images can be analyzed quantitatively using appropriate software. Such images can also be enhanced to allow objects not directly visible through the eyepieces to be examined on a monitor.

Answer 38

digital cameras

Question 39

Transmission and scanning electron microscopes are based on the interaction of ______ with beams of ______.

Answer 39

tissue components, electrons

Question 40

The wavelength in an electron beam is much ______ than that of light, allowing a ______-fold increase in resolution.

Answer 40

shorter, 1000

Question 41

The ______ is an imaging system that permits resolution around 3 nm. This high resolution allows isolated particles magnified as much as ______ times to be viewed in detail.

Answer 41

transmission electron microscope (TEM), 400,000

Question 42

Very thin (40-90 nm), resin-embedded tissue sections are typically studied by ______ at magnifications up to approximately ______ times.

Answer 42

TEM, 120,000

Question 43

Figure 1–8a indicates the components of a TEM and the basic principles of its operation: a beam of electrons focused using ______ “lenses” passes through the tissue section to produce an image with black, white, and intermediate shades of gray regions. These regions of an electron micrograph correspond to tissue areas through which electrons ______ readily (appearing ______ or ______) and areas where electrons were ______ or ______ (appearing ______ or more ______).

Answer 43

electromagnetic, passed, brighter, electron-lucent, absorbed, deflected, darker, electron-dense

Question 44

To improve contrast and resolution in TEM, compounds with ______ are often added to the fixative or dehydrating solutions used for tissue preparation. These include osmium tetroxide, lead citrate, and uranyl compounds, which bind cellular macromolecules, increasing their electron ______ and ______.

Answer 44

heavy metal ions, density, visibility

Question 45

______ and ______ are techniques that allow TEM study of cells without fixation or embedding and have been particularly useful in the study of membrane structure. In these methods, very small tissue specimens are rapidly frozen in liquid nitrogen and then cut or fractured with a knife. A replica of the frozen exposed surface is produced in a vacuum by applying thin coats of vaporized platinum or other metal atoms. After removal of the organic material, the replica of the cut surface can be examined by TEM. With membranes the random fracture planes often split the lipid bilayers, exposing protein components whose size, shape, and distribution are difficult to study by other methods.

Answer 45

Cryofracture, freeze etching

Question 46

______ provides a high-resolution view of the surfaces of cells, tissues, and organs.

Answer 46

Scanning electron microscopy (SEM)

Question 47

Scanning Electron Microscopy Like the TEM, this microscope produces and focuses a very narrow beam of electrons, but in this instrument the beam does not ______ through the specimen. Instead, the surface of the specimen is first dried and spray-coated with a very thin layer of ______ (often ______) that reflects electrons in a beam scanning the specimen. The reflected electrons are captured by a detector, producing signals that are processed to produce a black-and-white image.

Answer 47

pass, heavy metal, gold

Question 48

SEM images are usually easy to interpret because they present a ______ view that appears to be illuminated in the same way that large objects are seen with highlights and shadows caused by light.

Answer 48

three-dimensional

Question 49

______ is a method of localizing newly synthesized macromolecules in cells or tissue sections.

Answer 49

Microscopic autoradiography

Question 50

AUTORADIOGRAPHY Radioactively labeled metabolites (nucleotides, amino acids, sugars) provided to the living cells are incorporated into specific macromolecules (DNA, RNA, protein, glycoproteins, and polysaccharides) and emit weak ______ that is restricted to those regions where the molecules are located.

Answer 50

radiation

Question 51

AUTORADIOGRAPHY Slides with radiolabeled cells or tissue sections are coated in a darkroom with ______ in which silver bromide crystals act as ______ of the radiation in the same way that they respond to light in photographic film. After an adequate exposure time in lightproof boxes, the slides are developed photographically. Silver bromide crystals reduced by the radiation produce small black grains of ______, which under either the light microscope or TEM indicate the locations of radiolabeled macromolecules in the tissue.

Answer 51

photographic emulsion, microdetectors, metallic silver

Question 52

AUTORADIOGRAPHY Much ______ information becomes available by autoradiography. If a radioactive precursor of DNA (such as tritium-labeled thymidine) is used, it is possible to know which cells in a tissue (and how many) are replicating DNA and preparing to ______. Dynamic events may also be analyzed. For example, if one wishes to know where in the cell protein is produced, if it is secreted, and its path in the cell before being secreted, several animals are injected with a radioactive amino acid and tissues collected at different times after the injections. Autoradiography of the tissues from the sequential times will indicate the migration of the ______.

Answer 52

histological, divide, radioactive proteins