Week 10 Flashcards
What are mechanisms
◦ Processes or techniques by which something works
◦ Sequential
◦ Has beginning and ending
what are the two types of mechanisms
- Physical mechanisms
o Natural phenomenon - Chemical mechanisms
o Artificial phenomenon
What plays a part in the physical M of S
- Adsorption
- Selective solubility
- Porosity
What is Adsorption
what is its capacity provided by
and what forces are responsible for it
◦Combination of tissue and dye is by latent valences of atoms on the surface
Dye (Adsorbed material) + Tissue (Absorbent) - dyes can be removed with water or alcohol- temporary
capacity provided by :
◦ Surface area- larger the area the more place for dye to be and less intense the stain , small area= more intense the stain
◦ Size of pores- dense material = intense stain
◦ Surface polarity - charges on dye , opposite sites attract = lots of binding sites
forces responsible - weaker forces
◦ Van der Waals
◦ Electrostatic forces
difference between Adsorption & Absorption
◦ ADSORPTION: accumulation of gas, liquids, or solutes on the surface of a solid or liquid - can be readily removed
◦ ABSORPTION: transfer of material from one phase to another to form a “mixture”
What is selective solubility
◦ Used by Lysochromes (e.g. Oil Red O) to stain neutral lipids
-dye prefers where it is more soluble and it is more soluble in its own lipid than working solution
◦ Lysochrome dissolves in fat but1 not in its solvent
◦ Cannot use paraffin wax sections; frozen section only
-dye molecules select and choose to be dissolved in lipid
-dye molecules in a working solution of Oil red are unstable because it has a lower alcohol content so they prefer to be in lipid than in the solution where they are soluble.
The molecules would be happier in the stock solution where there is 100% isoprop
What is porosity
◦ Fixed protein precipitates into meshwork that measures void spaces in material
Porosity depends on
◦ tissue densities - densest tissue is least porous (collagen least - RBC most)
◦ tissue porosities
◦ Dye charge
◦ Dye molecular weight
all when tissue is fixed
collagen - least dense and RBCs most dense but both have positive charge. So they have an affinity to negatively charged dyes. The denser the meshwork the more positive charges there are. Therefore dye will enter the collagen first because its less dense but also leaves quicker.
acidophilic tissues - Positive charge
small weight dye larges stay in RBCs longer because the small one stay in the small pores so large ones will only fit in large pore like in collage
What are the chemical M of S
- Non-ionic bonding
- Ionic bonding
- Leuco compounds
- Metachromasia
- Histochemical reactions
- Metallic impregnation
What is non ionic bonding
- no charges involved
-hydrogen bonds
-van der waals forces
collagen and elastin
What is ionic bonding
routine h&e
opposite attract
change of isoelectronic point
if the tissue is
ACIDOPHILIC = POSITIVE
E.g. CYTOPLASM: RBC, MUSCLE, COLLAGEN
ACID DYE- ANIONIC DYE = NEGATIVE
if tissue is
BASOPHILIC = NEGATIVE
E.g. NUCLEI, CHROMATIDS, DNA
BASIC DYE CATIONIC DYE = POSITIVE
Zwitterion form of amino acid when it is pretreated with acid
one has positive charge and other has neg therefore net charge is 0 - IEP
-Adding acid would make the net charge +1 net increase in positive charge
-when acid stain (-) put on this pretreated tissue there would be bright color because of increase in dye binding sites due to increase of positive binding sites on the tissue
-when a basic stain positive charge is put on this tissue there would be a dull color because there are less negative charges on the protein
Zwitterion form of amino acid when it is pretreated with akaline
addition of alkaline would make charge -1
-basic dyes like an alkaline solution
when acid stain (-) is put on the color would be dull because there are less positive charges on the tissue protein
When there is a basic stain (+) applied the color would be bright because there is an increase in number of dye binding sites due to more negative binding sites
What are leuco compounds in staining
◦ Colorless compounds obtained from or converted into colored dye
Dye chromaphore gets reduced
Sulphonation > Schiff reagent
Hydrogenation > carbinol
-Both lose ability to stain because their chromophores were destroyed- leuco compound
The compound loses color - get oxidized by aldehyde group in tissue-chromophoric structure is restored. substance has color again
What is Metachromasia
Chromotrope are tissue components capable of changing the colour of (metachromatic) dyes
META- changing CHROME- colour
tissue (chromotrope can change the color of metachromatic dye) + toluidine blue ->chromotrope stains purple red but most will stain blue orthochromatic
stains purple and not blue because the matric is a chromatrope
What is polymerization
metachromasia takes place when there is polymerization
Basic dye + chromotropes
◦ Chromotropes must be close together and spaced evenly
◦ Chromotropes are basophilic or anionic – negatively charged, metachromatic dyes are cationic – positively charged
◦ Use water as solvent (no alcohol)
◦Presence of water and where the dye molecules will allow Chromotropes to absorb different light from the dye a – while the rest of the tissue pick up the blue colour of the dye.
