Introduction Flashcards
Study Deck for Introductory
Hematoxylin
Basic dye that stains (purple):
1) DNA - heterochromatin and nucleoli
2) Cytoplasmic RNA/ribosomes
3) extracellular complex carbohydrates (ex; cartilage)
Fixation
- Typically Formaldehyde
- Embedding a paraffin and thinly sliced on a microtome
- Hematoxylin & Eosin (H&E) staining: most common for histological preps
Basophilia
ability of anionic groups to react with a basic dye
Eosin
Acidic dye that stains (pink):
1) general membranous cytoplasmic organelles
2) intracellular fibers (ex; muscle cells)
3) extra cellular fibers (ex; connective tissue)
Acidophilia
Ability of cationic groups groups to react with acidic dye
Electron microscopy
higher magnification than light microscopy
Scanning electron microscopy (SEM)
surface
Transmission electron microscopy
inside
immunoflourescence
- visualization of proteins in fixed tissues
- direct: antibodies directly label with fluorescent marker
- indirect: primary and secondary antibodies
Direct immunofluorescence
antigen to antibody
Indirect immunofluorescence
antigen to primary antibody to fluorescent secondary antibody
more cost efficient
polycolonal and monoclonal antibody production
1) Inject antigen into the animal
2) bleed them out
3) collect antibodies
monoclonal antibody production
1) isolate B cell (antibody-making WBC) on cell culture
2) find one cell that makes desired antibody
3) fuse to a myeloma, keep growing
Immunogold labeling
Antibody conjugated to a gold particle (electron-dense secondary antibody due to gold)
Used under TEM to find proteins proteins (ex; proteins found in secretory vesicles)
In situ hybridization
- visualizing mRNA in tissues or living organisms
- DNA or RNA probe conjugated to fluorescent probes, biotin or digoxigenin
- Uses DNA/RNA probe instead of antibody
ex) fruit fly embryos and fluorescence in trisomy 21
Regular In situ hybridization shows up purple and fluorescent shows up fluorescent
Hematoxylin and eosin staining steps:
1) Hematoxylin (water soluble)
2) series of alcohol treatments to dehydrate the sample
3) eosin (alcohol soluble)
4) cleared with decreasing concentration of alcohol (xylol/toluol)
5) permanently embedded in a non-aqueous medium, cover-slip, and sealed
Cell Structure
Cell function is defined by shape, cytoplasmic organelle composition, cytoplasmic protein composition, and surface marker identity
visualizing microvilli
H&E staining, appearing hazy outside cells
function: increase SA
Found in: placenta, small intestines
Small Intestines: associated with a glycocalyx - mixture of carbohydrates and glycoproteins
Intestines: terminal digestive enzymes (for proteins and carbs)
Visualizing membrane protein
Existence of plasma proteins confirmed by freeze-fracture
Causes membrane to split:
- E-face: backed by extracellular space
- P-face: backed by cytoplasm
Membrane trafficking
1) endocytosis
- pinocytosis
- phagocytosis
- receptor-mediated endocytosis
2) exocytosis
Pinocytosis
a form of endocytosis, every cell does this as a part of natural homeostasis (equilibrium-healthy)
Visualized by light and TEM - large vesicles under light and small under EM
Phagocytosis
Specific endocytosis to neutrophils
- need pseudopodia to engulf
visualized by EM
Exocytosis - secretory cells
ex) endocrine/exocrine cells, neurons
Signal results in flux of Ca2+ and vesicle release
Protein making organelles
Rough ER and golgi apparatus
