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Histology > Microscopy and Epithelia > Flashcards

Microscopy and Epithelia Flashcards

1
Q

How do light microscopes work

A

They use visible light which passes through the specimen and is collected by the image forming optics to reveal the structure of living cells and tissue

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2
Q

How do you estimate the size of a structure under a light microscope

A

Imagine how many can fit into the diameter of a field of view

Or

By comparing it to a structure of known size (eg human blood cell with a diameter of 7μm). This method is best when using EM

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3
Q

What is fixation

A

A procedure whereby a given cellular structure is preserved at the expense of other structures

Fixatives that coagulate protein preserve the cytoplasm and nucleus

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4
Q

Can fixatives and use chemical changes to the tissue

A

Yes

Alcohol and other organic solvent extract fat and cause a fat droplets to look empty

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5
Q

Name 2 commonly used fixatives

A

Formaldehyde

Glutaraldehyde

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6
Q

What are the two methods of tissue fixation

A

Immersion

Perfusion

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7
Q

Describe immersion fixation

A

Tissues are immersed in a fixative a solution for the fixing to diffuse into the tissue

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8
Q

For immersion how do you ensure rapid penetration of the fixative into every cell

What can happen

A

The tissue is rapidly chopped up with a razor blade into small fragments before immersion

Structures in the deeper parts of tissue may be less preserved and some crushing and mechanical damage to cells is unavoidable

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9
Q

Why are capillaries difficult to identify in immersion

A

Capillary space is squashed

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10
Q

True or false: blood cells are preserved when a tissue is fixed by immersion

A

True

Some blood vessels may appear full of blood cells

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11
Q

How does perfusion work

A

The fixative reaches tissues through the blood vessels as it is pumped in via the largest arterial supply

Every cell is fixed virtually instantaneously without cell stress other than induced by the fixative

The capillary blood volume is preserved as it is in vivo but most of the blood cells flushed out

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12
Q

When are distortions introduced

A

During slide preparation

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13
Q

What does basophilic mean

Eg?

A

Refers to cellular structures stained by basically dyes

Nucleus, rough ER, RNA, DNA, acid glycoproteins

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14
Q

What do you call structures stained by acidic dyes

A

Acidophilic

Eg Mitochondria, collagen, cytoplasmic proteins, Secretory granules

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15
Q

Which combination of two days is the most commonly used

Why is it good

A

H and E

(Haematoxylin and eosin)

H is a basic dye and E is an acidic dye

H dyes acidic structures blue
E dyes basic structures pink

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16
Q

What is good about Masson trichrome

A

Shows nucleus and cytoplasm very well as well as connective tissue including collagen

This helps to differentiate collagen from smooth muscle

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17
Q

What colours does Masson trichrome stain

A

Basophilic: blue

Cytoplasm, muscle, RBC, keratin: bright red

Collagen: green/ blue

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18
Q

What colours does Van Gieson stain

A

Collagen: red

Nuclei: blue

Cytoplasm and RBC: yellow

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19
Q

When is Alcuin blue used

A

To stain mucin and cartilage

Often combined with H and E or Van Gieson

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20
Q

When used with Van Gieson,, Alcian blue changes times what colour?

A

Green

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21
Q

What is PAS

A

Periodic acid-Schiff reaction

It is used to detect polysaccharides, glycoproteins, glycolipids and neutral mucins

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22
Q

What does the Schiff reagent comprise

A

Basic fuchsin , HCL and sodium metabisulphite

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23
Q

What is Giemsa

A

Commonly used for staying in blood cells

Nuclei: dark blue

Cytoplasm: pale blue

RBC: pale pink

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24
Q

What is osmium tetroxide

A

A good fixative and staying for lipids in membranous structures and vesicles

Commonly used to stain myelin with a brown-black colour

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25
Q

How is osmium tetroxide

A

In electron microscopy both as a fixative and a heavy metal stain

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26
Q

How thick is a plasma membrane

A

10nm

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27
Q

What is the endoplasmic reticulum

A

A system of intra cellular membranes which serves both two separate special areas from the cytosol and to provide a service to support organised arrays of enzymes

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28
Q

How does the name endoplasmic reticulum reflect its nature

A

Endoplasmic reflects its position in the cell and the fact that parts of it joined together to form a loose arrangement and it links to the etymology of reticulum (from the Latin meaning net)

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29
Q

What is the cytosol

A

The colloid solution of substances in which the organelles are set

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30
Q

What is the rER

A

The rough endoplasmic reticulum

The region of the ER with ribosomes attached to the cytosol surface

Membranes are arranged as large flattened sacs called cisternae

These regions are specialised for synthesis and segregation of proteins destined to be stored or secreted by the cell

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31
Q

What makes the rER rough

A

The ribosomes which are attached to the membrane

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32
Q

What is the sER

A

The regions of the ER with a smooth appearance that are not associated with ribosomes

These regions are tubular and form anastomosing network which may join onto bicycles or sister night of the rER

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33
Q

What does the sER provide

A

The space for the synthesis of lipids,lipoproteins and steroid substances; The formation of glycogen and for the segregation of some ions within the cytoplasm

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34
Q

Which ions in particular are segregated in the smooth endoplasmic reticulum

A

Calcium

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35
Q

Describe ribosomes

A

Rounded structures about 15 mm in diameter particularly abundant in cells including large amount of protein

A ribosome is an enzyme complex composed of RNA molecules and protein

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36
Q

Where can free ribosomes be found

A

In the cytosol

They are often linked by mRNA to form a polyribosome

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37
Q

Describe the Golgi apparatus

How does it appear

A

A large organelle highly developed in secretory cells

As a series of flattened, smooth membrane sacs stopped upon one another, surrounded by a number of membrane icicles and support proteins to and from the apparatus

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38
Q

Name some functions of the Golgi

A

Condensation of proteins to form secretion droplets or granules,

the recycling and alteration of cytoplasmic membrane is,

the addition of the carbohydrate to glycoproteins, glycoproteins and mucins

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39
Q

What are lysosomes

A

Spherical organelles which contain a large amount of hydrolytic enzymes involved in the intracellular digestion of macro molecules, membranes and organelles

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40
Q

What are lysosomes known as initially

A

Primary lysosomes

These are spherical organelles which appear homogenous in content and do not contain particles or membranes

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41
Q

What do primary lysosomes do

A

Fuse with material taken into the cell by phagocytosis or pinocytosis

They also give rise to secondary lysosomes (which have a diverse morphology and contain particles undergoing enzymatic digestion)

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42
Q

What are peroxisomes

A

AKA micro bodies

Small spherical organelles which contain enzymes involved oxidative reactions, possibly forming hydrogen peroxide

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43
Q

How is the concentration of hydrogen peroxide from peroxisomes regulated

A

By the enzyme catalase (present in peroxisomes in high concentrations)

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44
Q

Which 2 organelles Were thought to be the same for many years

Why

A

Lysosomes and peroxisomes

Due to the morphological similarity

45
Q

Name 2 fuel reserves in cells

A

Glycogen granules

Lipid droplets

46
Q

What does glycogen appear as

A

Rosettes which are larger and darker than ribosomes in an EM

47
Q

Why do lipid droplets aggregate

A

Lipid cannot mix well with the aqueous materials in the cytoplasm

48
Q

What is resolution

A

The minimum distance that must exist between two small structures before they can be distinguished a separate entities

49
Q

What is embedding

What does it facilitate

A

The procedure to infiltrate the tissue with a stiff flexible material that can be cut

Thin section of the tissue with a microtome

50
Q

What is often used for embedding

A

Paraffin

51
Q

What was to happen before a tissue is embedded

A

Because you must be dehydrated by passing it through a series of graded alcohol is ending in 98% alcohol

Tissue is then immersed into melted paraffin wax in the oven at about 6°C. This causes the spaces within the tissues to become impregnated with paraffin

When the tissue is taken out of the oven the impregnated paraffin solidifies

52
Q

What is sectioning normally done with

How thin should it be

A

A microtome

Thin enough to transmit light (~7μm)

53
Q

Which dyes are generally used to dye each biochemical component

A

Alcian blue- mucopolysacchrides

Eosin - acidophilic components

Haematoxylin - basophilic components

Ponceau S - elastin

Osmium tetroxide- lipid

54
Q

Give five important differences between transmission electron microscopy and LM

A

A beam of electrons rather than a beam of light is used

Prevent the destruction of electrons, the electron beam operates in a vacuum

As a vacuum is employed, specimen must always be dead. Specimens must be fixed in plastic. The vacuum and the electron beam can be damaging to the specimen

A high resolution and higher magnification can be achieved

55
Q

What resolution and magnification can be achieved with transmission light microscopy

A

Resolution: 0.2nm

Magnification: x500,000

56
Q

What does SEM provide

A

Scanning electron microscopy provides 3D high resolution images of cells and tissue

57
Q

How is the diameter of SEM determined

A

By the diameter of the electron beam (2.5-7.5nm)

58
Q

How are specimens for SEM prepared

A

Fixed and dehydrated but not embedded

The surface has to be coated with heavy metal before examination

59
Q

How thin are specimens for EM

How is sectioning done

How is staining done

A

(50-80nm)

Using ultramicrotomes

By exposing the sections to have a metal salts which render the various cellular constituents electron opaque
Different constituents are revealed with Darius degrees of contrast according to that degree of impregnation with these salts

60
Q

What is freeze fracture etch

A

Specimens are quickly frozen prior to a embedding (below -150° C) then fractured

At this temperature breakage tends to occur along the natural lines of weakness, between the phospholipid bilayer of the cell membrane

Action is done by exposing the material in a vacuum

Water sublimes and the relief of the surface is enhanced

A heavy metal replica is made to preserve the surface which is then examined by TEM

61
Q

What is the depth of field of a typical electron microscope

A

5μm

62
Q

When do membranes form sharp images in an electron micrograph

A

When they are running vertically through the section i.e. parallel to the electron beam

63
Q

What is immunocytochemistry

A

A technique used to detect the presence of specific proteins or antigens in cells sampled by use of antibodies but bind to get a target with high specificity and affinity

64
Q

How does immunocytochemistry work

What can it be used to do

A

Antibodies which point we specific protein can be generated by me reactions. Such antibodies when isolated and provide contact with radioactive isotopes etc.

To quantify the amount of protein present But must be done in cells that have had the extracellular matrix removed

65
Q

What is immunohistochemistry

A

The same as immunocytochemistry but the original architecture of the tissue surrounding the cell is maintained

66
Q

What in situ hybridisation

A

When shot nuclear segment of a particular sequence or synthesise chemically and tagged

They bind to coiled complementary DNA in the nucleus or mRNA in the cytosol

This allows gene expression to be determined

67
Q

What is usually used to localise the probe in in situ hybridisation

A

Auto radiography

68
Q

What is autoradiography

A

When the tissue is fixed, embedded and section and the sections covered in the dark with a photographic emulsion

Molecules in the section with Incorporated radioactivity will activate the silver greens in the emulsion lying in above them

The emotion can then be developed photographically to demonstrate the activated silver grains and the section stained to identify the exact position of the label

69
Q

What do all epithelia rest on

A

A basement membrane known as the basal lamina

70
Q

Describe the basal lamina

A

I specialise extracellular matrix that marks the boundary between the epithelial cells and the underlying supporting tissue

It anchors down the epithelium to its connective tissue underneath

71
Q

How is the basal lamina connected to the underlying tissue

A

Proteins in the cell membrane is attached to proteins in the basement membrane which in turn is linked to the extracellular matrix of connective tissue

72
Q

Is there blood supply between epithelial cells

A

No because blood vessels do not cross the basement membrane

Instead epithelial cells rely on diffusion of oxygen and metabolite from capillaries in the underlying supporting tissue

73
Q

Name one important component of the basement membrane

A

Type IV collagen

74
Q

What does it mean to say epithelial cells show polarity

A

They have two different surfaces: apical and basolateral, each with specific functions and membrane constituents

75
Q

What is a main common function in of epithelia

A

To serve as a selective permeable membrane separating compartments that have different chemical compositions and regulating the substances that move between them

76
Q

Why do most epithelium have stem cells

A

To replenish the tissue on a continuous basis

77
Q

Where is simple squamous epithelium found

A

Lining surfaces at which diffusion occurs

78
Q

Describe cuboidal epithelium

A

Round with a centrally located nucleus, often more polygonal than cuboidal

79
Q

Where is simple cuboidal epithelium found

A

Lining tubules and ducts that may be involved in secretary, excretory and absorptive processes

80
Q

Describe simple columnar epithelium

Where is it found

A

Tall columnar cells with a size that varies depending on location and elongated nuclei that often lies at the base of the cell (may also be at centre or apex)

Lining the large duct of exocrine glands and maybe ciliated at the luminal surface of certain locations e.g. in the Fallopian tube

81
Q

Describe pseudostratified epithelium

A

In some simple epithelia, cells can be much taller than others and their nuclei are disposed at different levels, hence there appear to be two or more distinct layers

82
Q

True or false

Most of the cells of pseudostratified epithelium are ciliated

A

True

They line the larger airways of the respiratory system

83
Q

What are cilia

A

Motile cell projections which develop from basal bodies in the apical portion of the cytoplasm

84
Q

What is the structure of cilia

A

A central core of microtubules called the axoneme Which shows a characteristic arrangement of a central pair of microtubules surrounded by nine concentrically arranged microtubular pairs

85
Q

What do micro villi contain

A

A core of cross-linked microfilaments

86
Q

Compare the sizes of microvilli and cilia

A

Microvilli are 10x smaller and cannot be resolved in LM

87
Q

The cells of the basal layer of stratified epithelium are generally which shape

How are stratified epithelium classified

A

Cuboidal

On the basis of the shape of the cells in the surface layers

88
Q

Describe the stratified squamous epithelium

A

Have a flat surface layer

The Bazel cells are undifferentiated and divide continuously. The new cells push upwards to replace the dead cells at the top layer which ultimately shed

The cells covering the basal layer of differentiating cells which become highly differentiated as they reach the top layer

As the cells migrate there is a progressive change in cell shape (from cuboidal to squamous)

89
Q

Why have we adapted stratified squamous epithelium

How

Where is it found

A

To withstand abrasion so it’s main function is protection

The loss of the surface cells does not compromise the underlying tissue

Found lining the skin, oesophagus, vagina etc

90
Q

What is the epithelium of the skin covered by

A

A layer of fibrous protein known as keratin

It is therefore known as stratified squamous keratinised epithelium

91
Q

What is urothelium?

A

Transitional epithelium

Specialised epithelium to allow large degrees of stretch and to withstand the toxicity of urine

92
Q

Why is it known are transitional epithelium

A

Its appearance changes dramatically between stretched and unstretched states

Unstretched it appears to be four or five cell layers thick but stretched it appears as only two or three cell layers thick and the intermediate and surface layers are flattened

93
Q

What shape are the base and intermediate cells of transitional epithelium

What about the surface cells?

A

Cuboidal or polygonal but this shape and size changes with degrees of distension

They are umbrella cells which are large and domed shaped, often overlapping two or more of the underlying cells. They may contain multiple nuclei

94
Q

What do the surface cells of transitional epithelium contain

A

Plaques of urine resistant proteins

These are added to the surface to allow the increase in area when the cells change from an unstretched to a flattened stretched shape

95
Q

How are epithelial cells attached to each other

A

Laterally by specialised intercellular junctions.

96
Q

What are the main types of junction present in mammalian epithelia

What is the combination of these known as

A

Tight junction
Intermediate junction
Spot desmosome

A junctional complex

97
Q

How are tight junctions and intermediate junctions usually present

What about spot desmosomes?

A

one under the other as belt structures surrounding the cell

Spot desmosomes occupy discrete limited areas

98
Q

What are the two other names of a tight junction

A

Occluding junction

Zonula occludens

99
Q

What are the two other names of an intermediate junction

A

Zonula adherens

Belt desmosome

100
Q

What are the two other names of a spot desmosome

A

Desmosome

Macula adherens

101
Q

What do intercellular junctions allow

A

Intercellular mechanical attachment,

prevention of passage of macromolecules through the intercellular space

allow metabolic cell coupling

102
Q

What is a tight junction

A

A belt like zone that runs around the intercellular region immediately below cell’s luminar surface

Holds cells together,

functions as a permeability barrier between the two compartments separated by the epithelium,

and prevents the diffusion of membrane proteins along the plasma membrane

103
Q

What is an intermediate junction

A

Located below the T-junction at forms an anchoring belt around the cells

104
Q

Describe a desmosome

A

A circular spot of strong mechanical attachment

It is located deeper, below the intermediate junction

There can be a variable number of desmosomes binding 2 adjacent cells in epithelium

105
Q

What is a hemidesmosome?

A

A variant of desmosome that provides point attachment to the basement membrane

106
Q

Which stain allows junctional complexes to be seen

A

A protein stain eg iron Haematoxylin

They appear as short dark lines between cells

107
Q

What are gap junctions

What is their primary role

A

Minute intercellular channels that allow the passage of small molecules and ions directly between cells

To coordinate the activities of adjacent cells

108
Q

What is the terminal web

What is the function

A

A specialisation of the cytoskeleton found in certain epithelial cells with microvilli such as intestinal epithelium cells

To allow limited movement of the cell apex and microvilli

109
Q

Describe a terminal web

A

At the Junctional complex, fibrous elements of the cytoskeleton are attached at the same membrane. These fibres, mostly actin microfilaments, run across the apex of the epithelial cells, up into any microvilli and attach to the intermediate junction

Histology (9 decks)

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