Week 5 Downing - Histology Methods, Epithelium, CT Proper Flashcards
Light Microscope
A.K.A. Bright Field
-routine laboratory microscope used for studying tissue sections
Transmission Electron Microscope (TEM)
Used to study cytology or internal structures of cells
(study of electron micrographs)
Scanning Electron Microscope (SEM)
Used to study the surface features of cells and tissues
(obtain a 3-dimensional picture of the tissue)
Polarizing Microscope
Permits one to determine whether biological materials have different refractive indices along different optical axes.
Phase Microscope
Used to study living tissue.
(works on a principal of different refractive indices of cellular and sub-cellular components)
Interference Microscope
A modification of the phase microscope used for the study of living tissue.
Fluorescence Microscope
Used to examine the presence of fluorescent material in tissue sections.
(Uses UV light as the light source)
Confocal Scanning Microscope
Used to optically section a cell and with the appropriate computer equipment can reconstruct a
3-D image of the cell.
(uses a layer energy beam)
What are the 6 steps in preparing tissues for bright field microscopy?
- Fixation (stabilize tissue structure)
- Dehydration (remove water, make miscible)
- Clearing (replace alcohol with agent miscible to paraffin)
- Infiltration & Embedding (replace clearing agent with embedding material)
- Sectioning (cut tissue into thin sections)
- Staining (add color)
How can frozen histological sections be useful?
- pathology examination
- localization of enzymes
- analyze tissue sample
What are 6 common artifact in tissue slides?
- Post-mortem degeneration
- Shrinkage
- Precipitates (bad formalin)
- Wrinkles & Folds
- Nick in the microtome knife (results in tearing or scraping of tissue when cutting)
- Mishandling (pinching of tissue)
What is an acidic dye?
Capable of forming a salt linkage with a positively charged tissue group.
(dye molecule is negatively charged - anionic)
What is a basic dye?
A positively charged (cationic) and hence forms a salt with a negatively charged tissue group.
What is an H stain?
Hematoxylin (blue → purple)
basic stain
What is an H stain used for?
To stain basophilic substances.
- DNA
- chroms
- heterochromatin
- RNA
- nucleolus
- cytoplasmic ribosomes
What are basophilic tissues?
Structures in the cell or tissue that love basic stains.
(e.g. nucleus)
What is an E stain?
Eosin (red → pink)
acid stain
What is an E stain used for?
Acidophilic substances (eosinophilic substances)
- Proteins
- Cations associated with side chains
What are acidophilic tissues?
Structures in the cell or tissue that love acid stains.
(a.k.a. eosinophilic substances)
When is Trichrome stain used?
Stain for connective tissue (collagen) rather than cells.
(e.g. Masson’s, Mallory’s)
When is Elastic stain used?
When staining for elastic fibers or elastic tissue in connective tissue.
(e.g. aldehyde fuchsin, orcein, resorcin-fuchsin)
When are Silver stains used?
When staining for reticular fibers in connective tissue.
Also for staining cells of the CNS.
(connective tissue fibers love silver and stain black - argyrophilic)
When are Fat stains used?
Oil red O - stains fats red
Sudan black - stains fats black
*soluble in both alcohol & fats
What is used to stain nucleic acid (DNA & RNA)?
Feulgen staining reaction for DNA
What is used for staining carbohydrates?
Periodic acid-Schiff reaction (PAS)
What does Immunocytochemistry allow?
Permits the cellular or intracellular localization of specific proteins.
Why are red blood cells useful in determining approximate magnifications of tissue images?
They are about 7um (micrometer) thick.
Can be used as a reference size.
What is the basic mechanism of the Periodic acid-Schiff reaction (PAS)?
- Oxidizes diols in sugars → creates two aldehydes
- aldehydes react with Schiff’s agent
*Used to detect carbs (polysaccharides, glycogen)
What is the basic mechanism of the Feulgen staining reaction?
- HCl removes purine bases from DNA
- unmasks free aldehyde groups
- free aldehyde groups react with Schiff’s reagent
- purple stained DNA!!!
(does not work on RNA)
What are the four major types of tissues in the body?
- Epithelium tissue
- Connective tissue
- Nervous tissue
- Muscle tissue
What are the four general characteristics of Epithelia?
- Sheets or layers of cells in close apposition
- Cover surfaces/line cavities
- Very little intercellular material (epithelium = highly cellular tissue)
- Always in contact with a basal lamina
What germ layers give rise to Epithelia?
ALL THREE!
(ectodoerm, mesoderm, & endoderm)
What are tight junctions?
- Zonula occludens
- apical domain
- cadherin
- freeze fracture technique: pattern of ridges
What are integrins? What is one example?
Mediate cell-extracellular matrix interactions.
ex. Hemidesmosomes
What are cadherins? What are some examples?
Mediate cell-cell interaction.
ex. Zonula occludens, Zonula adherens, Macula adherens
What are belt desmosomes?
- Zonula adherens
- belt of adhesion
- cadherin
- not as close as tight junctions
- cytoplasmic domains interact with actin microfilaments via linker proteins
What are spot desmosomes?
- Macula adherens (desmosomes)
- form spot-like “welds” between adjacent epithelial cells
- cadherin
- transmembrane proteins
- desmogleins & desmocollins
- desmoplakins
- dense plaque
- interacts with intermediat filaments
What are hemidesmosomes?
- looks like half of a desmosome
- forms between cell membrane of a basal epithelial cell and the adjacent basal lamina
- integrin
- cytoplasmic domains interact with intermediate filaments via linker proteins
- extracellular domains interact with laminin found in the basal lamina
What are gap junctions?
- Nexus
- 6 transmembrane protein subunits form connexin
- line up with opposite and identical structure in an adjacent cell
- create channel pore that connects the cytoplasm of the two adjacent cells
What are the two components of the basement membrane?
- Basal lamina
- lamina lucida
- lamina densa
- lamina reticularis
- Hemidesmosomes
What does the first part of the basal lamina, the lamina lucida, do?
- electron lucent portion of basement membrane
- rich in glycoproteins, laminin, and entactin
- binds to integrins
What does the second part of the basal lamina, the lamina densa, do?
- dense meshwork of type IV collagen
- sandwiched by the proteoglycan perlacan containing heparan sulfate side chains
- tightly anchored to reticular lamina
- by fibronectin
What does the third part of the basal lamina, the lamina reticularis, do?
- forms interface between the basal lamina and underlying connective tissue
What is the structure and function of microvilli?
- microscopic projections above free surface
- covered with cell membrane
- core of microfilaments
- form “brush border”
- increase surface area of cell
What is the structure and function of stereocilia?
- unusually long microvilli appearing
- apical ends twist together
- non-motile
- increase absorptive surface of cells
What is the structure and function of cilia?
- motile processes that extend from the apical surface of the cell
- shaft covered with plasma membrane
- arise from basal bodies
- axoneme: internal/core structure
- two single, central microtubules
- nine peripheral pares of fused double microtubules
- A-tubule (complete) + B-tubule (incomplete)
- doublets connected by nexin link
- dynein walks toward minus (-) end of doublet
What is the structure and function of flagella?
- same axial structure as cilia
- much longer than cilia
- beat is more random, whip-like motion
Epithelioid
term for an epithelium-like appearance of non-epithelial cells
Metaplasia
An abnormal change from one distinctive kind of cell to another at a definite site after development is over.
Neoplasia
An abnormal change from on distinctive kind of cell to a tumor cell.
ex. carcinoma: a malignant neoplasm of epithelial origin
- benign
- metastatic
Hyperplasia
An increase in the number of cells in a tissue or organ, excluding tumor formation.
Increases the bulk of the part or organ.
Hypertrophy
Overgrowth or general increase in bulk of a part or organ.
Anaplasia
Loss of structural differentiation.
Particularly prevalent in malignant neoplasms.
What is the embryological origin of connective tissue?
Mesoderm!
What are the physical characteristics of collagen?
Microfibril x3 → Fibril x3 → Fiber → Aggregate to form Bundle
- flexible
- non-extensible (inelastic)
- forms gelatin when placed in boiling water
What is the chemical composition of collagen fibers?
- polymerization of tropocollagen molecules into fibrils
- three chains of polypeptides (280nm long)
- triple helix
What is the chemical composition of amorphous intercellular substance (ground substance)?
- water
- glycosaminoglycans
- proteoglycans
- structural multi-adhesive glycoproteins (fibronectin, laminin)
What is the function of ground substance?
- efficient space filler
- permits passages of gases, ions, and small particles
- helps to immobilize larger objects (e.g. bacteria)
