Intro Flashcards
(126 cards)
1
Q
Study of the tissues of the body and how these tissues are arranged to constitute organs
A
Histology
2
Q
Involves all aspect of tissue biology, with the focus on how cell’s structure and arrangement optimize functions specific to each organ.
A
Histology
3
Q
Histology
Two interacting components:
A
cells and extracellular matrix (ECM)
4
Q
Logos
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study
5
Q
is basically the environment where these cells thrive and the one that nurtures the cells that forms its tissues; one that cells interact with to perform its functions.
A
ECM
6
Q
consists of many kinds of macromolecules, most of which form complex structures, such as collagen fibrils and basement membranes
A
ECM
7
Q
ECM consists of many kinds of macromolecules, most of which form complex structures, such as
A
collagen fibrils and basement membranes
8
Q
supports the cells and the fluid that transports nutrients to the cells, and carries away their catabolites and secretory products
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ECM
9
Q
form a continuum that functions together and reacts to stimuli and inhibitors together.
A
cells and ecm
10
Q
most common procedure used in histologic research is the ______ that can be studied with the light microscope.
A
preparation of tissue sections or slices
11
Q
Because most tissues and organs are too thick for light to pass through them, they must be ____ to obtain ____
A
sliced, thin, translucent sections
12
Q
The ideal microscopic preparation is ___ so that the tissue on the slide has
the same structure and molecular composition as it had in the body.
A
preserved
13
Q
Cut into a small segments before adding fixatives ( a chemical that will preserve the tissue)
A
fixation
14
Q
usually involves immersion in solutions of stabilizing or crosslinking compounds called
A
fixatives
15
Q
initial treatment
A
fixation
16
Q
One fixative widely used for light microscopy
A
formalin
17
Q
a buffered isotonic solution of 37%
formaldehyde.
A
Formalin
18
Q
a fixative often used for electron microscopy,
react with the amine groups (NH2 ) of tissue proteins, preventing their degradation
A
Glutaraldehyde and Osmium Tetroxide
19
Q
The tissue is transferred through a series of ending in 100%, which removes all water.
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Dehydration
20
Q
Most common dehydrating agent:
A
Ethanol or Ethyl Alcohol
30%, 50%, 70%
21
Q
Alcohol is removed in toulene or other agents in which both alcohol and paraffin are miscible (well-mixed)
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clearing
22
Q
Alcohol is removed in toulene or other agents in which both ___ and ____ are miscible (well-mixed)
A
alcohol and paraffin
23
Q
substance used for clearing
A
Xylene
24
Q
The tissue is then placed in melted paraffin
until it becomes completely infiltrated with this
substance
A
infiltration
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The tissue is then placed in _____
until it becomes completely infiltrated with this
substance.
melted paraffin
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temperature required for infiltration
52-60 degrees celsius
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52-60 degrees celcius. At such temperatures the clearing solvent evaporates and the tissue is filled with ____. The impregnated tissue then hardens in a small container of paraffin at room temperature.
liquid paraffin
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The paraffin-infiltrated tissue is placed in a small mold with melted paraffin and allowed to harden
embedding
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avoids the higher temperatures needed for paraffin embedding, which helps avoid shrinkage and major distortion of the tissue.
plastic embedding
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Preparation of Tissues
Slice, Fixation, Dehydration, Clearing, Infiltration, Embedding
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Embedding materials include
paraffin and plastic resins
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is used routinely for light microscopy,
paraffin
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used for both light and electron microscopy
resins
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Embedded
Tissue=
Tissue block
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The resulting paraffin block is trimmed to expose the tissue for sectioning (slicing) on a microtome
trimming
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The resulting paraffin block is trimmed to expose the tissue for sectioning (slicing) on a
microtome
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similar steps are used in preparing tissues for ____ except special fixatives and dehydrating solutions are used with smaller tissue samples and embedding involves epoxy resins which become harder than paraffin to allow very thin sectioning
transmission electron microscopy (TEM)
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is used for sectioning paraffin-embedded tissues for light microscopy.
microtome
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For TEM, sections less than ___ thick are prepared from resin-embedded cells using an ____ with a glass or diamond knife.
1micrometre, ultramicrotome
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Paraffin sections are generally cut at ____ thickness
1-10 μm
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Principle: Opposite attracts
staining
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Acidic structures - basic dye =
basophilic (eg.
Nucleus)
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Basic structures- acidic dye =
acidophilic (eg.
Cytoplasm, eosinophils)
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Of all staining methods, the simple combination of ______ is used most commonly.
hematoxylin and eosin (H&E)
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produces a dark blue or purple color
hematoxylin
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staining DNA in the cell nucleus and other acidic structures (such as RNA-rich portions of the cytoplasm and the matrix of cartilage)
Hematoxylin
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stains other cytoplasmic components and collagen pink
Eosin
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stained preparations are examined by means of ordinary light that passes through the specimen
Bright-Field Microscopy
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Reliant to resolving power
Bright-Field Microscopy
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critical factor in obtaining a crisp, detailed image with a light microscope
resolving power
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smallest distance between two particles at which they can be seen as separate objects
resolving power
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The maximal resolving power of the light microscope is approximately
0.2 μm
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Objects smaller or thinner than 0.2 μm (such as a _________) cannot be distinguished with Bright field microscopy
ribosome, a membrane, or a filament of actin
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The optical system has three sets of lenses:
condenser
objective lenses
eyepieces or oculars
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• The ___ collects and focuses a cone of light that illuminates the tissue slide on the stage.
condenser
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• _____ enlarge and project the illuminated image of the object toward the eyepiece. Interchangeable objectives with different magnifications routinely used in histology include X4 for observing a large area (field) of the tissue at low magnification; X10 for medium magnification of a smaller field; and X40 for high magnification of more detailed areas.
objective lenses
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The ______ magnify this image another X10 and project it to the viewer, yielding a total magnification of X40, X100, or X400.
two eyepieces or oculars
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tissue sections are usually irradiated with ultraviolet (UV) light and the emission is in the visible portion of the spectrum
fluorescence microscopy
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The fluorescent substances appear brilliant on a
dark background
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Phenomenon wherein certain cellular substances are irradiated by light of a proper wavelength, they emit light with a longer wavelength
fluorescence
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based on the principle that light changes its speed when passing through cellular and extracellular structures with different refractive indices
phase-contrast mocroscopy
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uses a lens system that produces visible images from transparent objects and, importantly, can be used with living, cultures cells
Phase-contrast microscopy
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Allows examination of unstained cells and tissues and is especially useful for living cells
Phase-contrast microscopy
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Used when you want to visualized actual cellular process (eg. Phagocytosis)
Phase-contrast microscopy
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Allows quantification of tissue mass
Interference
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Using Nomarski optics ( produces an image of living cells with a more apparent three- dimensional (3D) aspect )
Differential Interference Microscope
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Useful for assessing surface properties of cells and other biologic objects
Differential Interference Microscope
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Using Nomarski optics ( produces an image of living cells with a more apparent three- dimensional (3D) aspect )
Nomarski optics
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No direct light from the light source is gathered by the objective lens
Dark field microscopy
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Only light has been scattered or diffracted by structures in the specimen reaches the objective
Dark-Field Microscopy
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Equipped with a special condenser that illuminates the specimen with strong, oblique light
Dark-Field Microscopy
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Field of view appears as a dark background on which small particles in the specimen that reflect light into the objective appear bright.
Dark-Field Microscopy
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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
Confocal Microscopy
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are all optically conjugated or aligned to each other in the focal plane (confocal), and unfocused light does not pass through the pinhole
point light source, the focal point of the lens, and the detector’s pinpoint aperture
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Combines components of a light microscope with a scanning system to dissect a specimen optically
Confocal Microscopy
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Combines components of a light microscope with a scanning system to dissect a specimen optically
Confocal Microscopy
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Uses (1) a small point of high-intensity light, often from a laser, and (2) a plate with a pinhole aperture in front of the image detector
Confocal Microscopy
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allows the recognition of stained or unstained structures made of highly organized subunits
Polarizing microscopy
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Uses the fact that highly ordered molecules or arrays of molecules can rotate the angle of the plane of polarized light
Polarizing microscopy
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ability to rotate the direction of vibration of polarized light
Birefringence
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a feature of crystalline substances or substances containing highly oriented molecules, such as cellulose, collagen, microtubules, and actin filaments
birefringence
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When normal light passes through a polarizing
filter, it exits vibrating in only
one direction
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If a second filter is placed in the microscope above the first one, with its main axis perpendicular to the first filter,
no light passes through
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If, however, tissue structures containing oriented macromolecules are located between the two polarizing filters, their repetitive structure rotates the axis of the light emerging from the polarizer and they appear
bright background structures
against a dark background
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Types of Light Microscope
Bright Field Microscope
Fluorescence
Phase Contrast
Dark Field
Confocal
Polarizing
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based on the interaction of tissue components with beams of electrons
electron microscopy
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The wavelength in the electron beam is much shorter than that of light, allowing a _____ increase in resolution
1000-fold
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imaging system that permits resolution around 3nm
Transmission Electron Microscopy
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Transmission Electron Microscopy allows magnifications of up to _____ times to be viewed in detail
400,000
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The image of the specimen shows areas of __________ corresponding to areas through which electrons passed readily (appearing brughter or electron lucent)
white, black, and shades of gray
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areas where electrons were absorbed or
deflected
appearing darker or more electron dense
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normally requires very thin sections (40-90 nm); therefore tissue is embedded in a hard epoxy and sectioned with a glass or diamond knife.
Transmission Electron Microscopy
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Transmission Electron Microscopy normally requires very thin sections (______); therefore tissue is embedded in a _____ and sectioned with a glass or diamond knife.
40-90 nm, hard epoxy
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provides a highresolution view of the surfaces of cells, tissues, and organs
Scanning Electron Microscopy
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Scanning Electron Microscopy: The electron beam does ____ through the specimen but is ____ across its surface
not pass, scanned
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Scanning Electron Microscopy: the surface of the specimen is first dried and
spray-coated with a very thin layer of ____ through which electrons do not pass readily
metal (often gold)
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When the beam is scanned from point to point across the specimen, it interacts with the ____ and produces ______ emitted from the metal
metal atoms, reflected electrons or secondary electrons
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are large instruments generally house in a specialized EM facility
Electron microscope
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Types of Electron Microscope
Transmission Electron Microscopy
Scanning Electron Microscopy
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a method for localizing cellular structures using a specific enzymatic activity present in those structures
Enzyme Histochemistry (Cytochemistry)
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Enzyme Histochemistry (Cytochemistry): To preserve these enzymes, histochemical procedures usually use _____ tissue, which is sectioned on a ____ to avoid adverse effects of ____ and _____ on enzymatic activity.
unfixed or mildly fixed, cryostat, heat, organic solvents
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Examples of enzymes that can be detected histochemically include the following:
Phosphatases
Dehydrogenase
Peroxidase
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split the bond between a phosphate group and phosphorylated molecules
Phosphatases
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remove hydrogen ions from one substrate and transfer them to another
Dehydrogenases
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promotes the oxidation of substrates with the transfer of hydrogen ions to hydrogen peroxide, forming water molecules
Peroxidase
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A highly specific interaction between molecules is that between an antigen and its antibody
Immunohistochemistry
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very widely used to detect specific proteins (or other molecules) of interest in cells and tissues
Immunohistochemistry
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Immunohistochemistry requires an ___ against the protein that is to be detected, which means that the protein must have been previously purified using _____ or _____ so that antibodies against it can be produced
antibody, biochemical, molecular approaches
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The body’s immune cells interact with and produce antibodies against other macromolecules—called — that are recognized as “foreign,” not a normal part of the organism, and potentially dangerous.
antigens
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Antibodies belong to the ____ family of glycoproteins, produced by _____.
immunoglobulin, lymphocytes
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In immunohistochemistry a tissue section (or cells in culture) that one believes contains the protein of interest is ____ in a solution containing _____ to this protein.
incubated, labeled antibody
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The antibody binds specifically to the
protein
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Antibodies are commonly tagged with ___, with ____ for histochemical detection, or with _____ for TEM.
fluorescent compounds, peroxidase or alkaline phosphatase, electron-dense gold particles
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Immunocytochemistry (or immunohistochemistry) can be
direct or indirect
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uses an antibody made against the tissue protein of interest and tagged directly with a label such as a fluorescent compound or peroxidase. When placed with the tissue section on a slide, these labeled antibodies bind specifically to the protein (anti-gen) against which they were produced and can be visualized by the appropriate method.
Direct Immunocytochemistry
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uses first a primary antibody made against the protein (antigen) of interest and applied to the tissue section to bind
Indirect immunocytochemistry (right)
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method of localizing messenger RNA (mRNA) or DNA by hybridizing the sequence of interest to a complementary strand of a nucleotide probe
hybridization techniques
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Binding of the nucleotide probe to the DNA or RNA sequence of interest is performed within cells or tissues, such as cultured cells or whole embryos
In Situ Hybridization
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This technique is ideal for
(1) determining if a cell has a specific sequence of DNA, such as a gene or part of a gene
In situ hybridization
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identifying the cells containing specific messenger RNAs (mRNAs) (in which the corresponding gene is being transcribed)
hybridization
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determining the localization of a gene in a specific chromosome.
hybridization
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Certain steps in this procedure may distort the tissues slightly, producing minor structural abnormalities called ___ not present in the living tissue.
artifacts
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can create artificial spaces between cells and other tissue components. Such spaces can also result from the loss of lipids, glycogen, or lowmolecular-weight substances not preserved by the fixative or removed by the dehydrating and clearing fluids.
Shrinkage
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One such distortion is ____ of cells or tissue regions produced by the fixative, by the ethanol, or by the heat needed for paraffin embedding.
minor shrinkage
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in sections also appear as large spaces in the tissues
Slight cracks
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when a structure’s three-dimensional volume is cut into very thin sections, the sections appear microscopically to have only two dimensions:
length and width
