| Question | Answer |
| Artifact pigment | Formed as a result of chemical action -
fixation, processing,
usually on tissue, not w/i cell,
e.g. formalin, mercury, chrome pigments |
| Exogenous pigment | From outside the body,
carbon pigment, asbestos fibers,
tattoo pigments, metals - normal (iron) or pathologic (copper) |
| Carbon pigment | Seen in lung & lymph node,
resists bleaching & extration,
insol. in conc'd sulfuric acid - use to differentiate |
| Asbestos fibers | Birefringent, magnesium silicate,
in body fibers coated w/ iron-containing
protein, known as asbestos bodies,
lose birefringence demo w/ Prussian blue |
| Endogenous hematogenous pigments | From blood, e.g. hemoglobin, hemosiderin, biliverdin |
| Hemoglobin | Conjugated protein,
anionic, stains w/ acid dyes (eosin),
hemoglobin breaks into globin protein & heme,
heme splits into iron & biliverdin |
| Hemosiderin | Yellow to brown pigment,
stored iron (Fe2+),
usu. in marrow, pathologic elsewhere (liver),
differentiate from other yellow-brown pigments
w/ Prussian blue rxn |
| Hemochromatosis | Disease caused by excessive absorption of dietary iron,
excessive hemosiderin deposits in liver, pancreas,
damaging to tissue |
| Biliverdin | Greenish bile pigment,
prod'd from destruction of rbc,
liver red. to bilirubin - remove from circulation,
abnormal accumulation - jaundice, pigment in bile canaliculi, cytoplasm of Kupffer cells & hepatocytes |
| Hematoidin | Similar to bilirubin, ox. to bilirubin,
formed as result of hemorrhage & red'd O2 tension |
| Endogenous nonhematogenous pigments | Lipid - e.g. lipofucsin, ceroid,
nonlipidic - e.g. melanin |
| Melanin pigment | Formed by ox. tyrosine,
histochem. demo melanocytes,
bleach w/ H2O2, potassium permanganate & oxalic acid,
dissolved by strong alkali,
insol. weak acid, base, organic solvents,
argentaffin,
positive Schmorl |
| Lipofuscin pigment | "Wear & tear" pigment,
yellow-brown,
in more permanent cells (heart, liver, neurons) |
| Ceroid pigment | Brownish-yellow,
hepatocytes & macrophages of rats w/ cirrhosis,
rarely seen in humans |
| What stains for lipofuscin & ceroid pigments? | Oil red O, Sudan black B, & PAS,
differentiate - ceroid is acid-fast, lipofsucin is not |
| Engodenous deposits | Urate crytals (gouty tophi),
chronic gout may deposit in soft tissue too,
sol. in water, alcoholic fix.,
birefringent w/ polarizing,
demo w/ argentaffin rxn |
| Minerals | Special stains can demo: Ca2+, Fe2+/Fe3+, Cu2+,
PO43+, CO32+, silver, lead, copper, gold,
microincineration can be used to ID inorganic components |
| Cytoplasmic granules | Adrenal chromaffin granules, pancreatic endocrine cells,
GI enterochromaffin cells, "C" cells of thyroid,
some pituitary cells,
are APUD cells
demo w/ argyrophil or argentaffin rxns,
fixative v. important |
| APUD cells | Have high uptake of amine-precursors & ability to decarboxylate |
| Best fixative for chromaffin granules? | Primary chromate fixatives,
e.g. Orth |
| Argentaffin granules of GI are destroyed by? | Alcoholic fixatives |
| Paneth cell granules are destroyed by? | Acetic acid |
| Prussian blue stain purpose | Detect ferric (Fe3+) iron,
usu. small amount in marrow & spleen,
large deposits - hemochromatosis, hemosiderosis |
| Prussian blue stain facts | Fixative: alcohol or 10% NBF,
QC: section w/ Fe3+, don't want excess iron,
chemically clean glassware,
iron-free reagents |
| Prussian blue stain solutions | potassium ferrocyanide 2% (bind Fe3+)
HCl 2% (acidify)
nuclear-fast red |
| Prussian blue stain results | nuclei & hemofuchsin - bright red
hemosiderin (iron) - blue
background - pink |
| Nuclear-fast red solution | aluminum sulfate
nuclear-fast red
thymol (preservative)
dH2O |
| Fixation for iron demo: | Some acidic fixatives & decal. solns. dissolve iron,
can demo iron after Zenker w/ acetic acid,
but mercury - bad,
best to use another fixative & decal briefly in acetic or formic acid |
| Turnball blue stain purpose | Detect ferrous (Fe2+) iron,
ferrous iron v. toxic, not normally stored,
easily absorbed by intestine but converted to ferric,
used w/ Schmorl technique |
| Turnball blue stain facts | Fixative: alcohol or 10% NBF
QC: section w/ Fe2+ |
| Turnball blue stain solutions | potassium ferricyanide staining soln. (bind Fe2+)
acetic acid 1% (post-stain rinse)
nuclear-fast red |
| Turnball blue stain results | ferrous iron - blue
background - pink-red |
| Potassium ferricyanide staining solution | potassium ferricyanide
HCl 0.06N |
| Schmorl technique purpose | Indicate reducing substances - will red. ferric ions,
stains melanin, argentaffin granules, formalin pigment,
mostly replaced by IHC |
| Schmorl technique facts | Fixative: 10% NBF
QC: section w/ melanin or argentaffin granules |
| Schmorl technique solutions | ferric chloride-potassium ferricyanide working soln. pH 2.4
Mayer mucicarmine soln. or metanil yellow |
| Schmorl technique results | reducing/argentaffin substances - blue-green
goblet cells, mucin - rose
background - yellow-green (less better) |
| Mayer mucicarmine solution | carmine
alum lake
aluminum hydroxide
anhy. aluminum chloride
50% ethanol
25% ethanol |
| Fontana-Masson & microwave Fontana-Masson stains purpose | Demo argentaffin substances, e.g. melanin,
argentaffin granules of carcinoid tumors,
some neurosecretory granules,
demos other red'ing substances (formalin pigment) |
| Fontana-Masson & microwave Fontana-Masson stains facts | Fixative: 10% NBF, avoid alcohol
QC: skin for melanin,
small intestine or appendix for argentaffin granules,
Melanoma, tumor w/ argentaffin granules,
chemically clean glassware, nonmetallic instruments |
| Fontana-Masson stain solutions | Fontana silver nitrate w/ NH4OH
gold chloride
sodium thiosulfate
nuclear-fast red |
| Fontana-Masson stain results | melanin, argentaffin granules - black
nuclei - pink
overstaining gives gray background & loss of contrast |
| Microwave Fontana-Masson stain solutions | Fontana silver nitrate w/ NH4OH
gold chloride
sodium thiosulfate
nuclear-fast red |
| Microwave Fontana-Masson stain results | argentaffin cell granules, chromaffin granules, melanin,
other argentaffin substances - black
nuclei - pink |
| Grimelius stain purpose | Demo argyrophil granules in neurosecretory tumors,
also stains argentaffin substances |
| Grimelius stain facts | QC: argyrophil-positive carcinoid tumor preferred,
can use small intestine,
chemically clean glassware, nonmetallic forceps |
| Grimelius stain solutions | working silver solution
reducing solution
nuclear-fast red |
| Grimelius stain results | argentaffin & argyrophil granules - dark brown to black
nuclei - red
background - pale yellow-brown |
| Grimelius working silver solution | acetic acid-acetate buffer pH 5.6
silver nitrate 1%
dH2O |
| Grimelius, Churukian-Schenk, microwave Churukian-Schenk reducing solution | hydroquinone
sodium sulfite
dH2O |
| How to differentiate between argentaffin & argyophil substances? | Stain w/ both Grimelius (argyophil) & Fontana-Masson (argentaffin) & compare |
| Churukian-Schenk & microwave Churukian-Schenk methods purpose | Demo argyrophil granules in neurosecretory tumors |
| Churukian-Schenk & microwave Churukian-Schenk methods facts | QC: argyrophil-positive carcinoid tumor preferred,
can use small intestine,
chemically clean glassware, nonmetallic forceps |
| Churukian-Schenk method solutions | acidified water pH 4.0 to 4.2
silver nitrate 0.5%
reducing solution
nuclear-fast red |
| Churukian-Schenk method results | argyophil granules, argentaffin substances - black
nuclei - red
background - yellow-brown |
| Microwave Churukian-Schenk method solutions | citric acid-glycine soln.
silver nitrate 0.5%
reducing solution
nuclear-fast red |
| Microwave Churukian-Schenk method results | argyrophil & argentaffin cells - black
nuclei - orange to red
background - light yellow-orange |
| Gomori methenamine-silver method purpose | Demo urates,
may demo large calcium deposits |
| Gomori methenamine-silver method facts | Fixative: abs. alcohol
QC: section w/ urates |
| Gomori methenamine-silver method solutions | working methenamine-silver nitrate
sodium thiosulfate
light green w/ acetic acid |
| Gomori methenamine-silver method results | urates - black
background - green |
| Bile stain purpose | Detect bilirubin in tissue,
aka Hall stain,
aka Fouchet rxn |
| Bile stain solutions | Fouchet reagent
van Gieson |
| Bile stain results | bile, bilirubin - emerald green to olive drab
background - yellow |
| Fouchet reagent solution | trichloracetic acid
ferric chloride
dH2O |
| van Gieson solution | acid fuchsin
picric acid |
| von Kossa stain purpose | ID calcium in tissue,
detects anions bound to calcium, not the calcium itself |
| von Kossa stain facts | Fixative: alcohol preferred, 10% NBF ok |
| von Kossa stain solutions | silver nitrate
sunlight
sodium thiosulfate
nuclear-fast red |
| von Kossa stain results | calcium salts - black
background - red |
| Alizarin Red S stain purpose | ID calcium in tissue,
reaction product is birefringent |
| Alizarin Red S stain facts | Fixative: alcoholic formalin or 10% NBF |
| Alizarin Red S staining solution | Alizarin red S (chelate w/ calcium)
dH2O
NH4OH to pH 4.1 to 4.3 - critical |
| Alizarin Red S stain results | calcium deposits - orange-red |
| Rhodanine & microwave Rhodanine methods purpose | Detect copper in tissue,
esp. liver - Wilson disease,
aka Lindquist rhodanine |
| Rhodanine & microwave Rhondanine methods facts | Fixative: 10% NBF
Sections: paraffin 6-8 µm
QC: section w/ copper (fetal liver) |
| Rhodanine method solutions | working rhodanine soln.
Mayer hematoxylin w/ dH2O
borax (blue Mayer) |
| Rhodanine method results | copper - bright red to red yellow
nuclei - light blue
If copper conc. low, stain may fade after coverslipping |
| Microwave Rhodanine method solutions | working Rhodanine w/ sodium acetate-formalin
Mayer hematoxylin w/ dH2O
borax |
| Microwave Rhodanine method results | copper - bright red or rust-red intracytoplasmic granules
nuclei - blue |
| Working Rhodanine solution | 5-(p-dimethylamino-benyzlidine) rhodanine
abs. ethanol
dH2O
filter |
| Pigment | A substance that has enough color to be visible w/o any further staining |
| What are some diseases associated w/ melanin? | Melanomas,
Addison's disease |
| Prussian blue vs. Turnball blue | Prussian - ferrocyanide for ferric ions
Turnball - ferricyanide for ferrous ions |
| Fontana Masson w/ melanin bleach | To compare,
deparaffinize & hydrate 2 slides,
leave 1 in dH2O,
1 in potassium permanganate, wash w/ tap & dH2O,
in oxalic acid, wash w/ tap & dH2O,
stain both w/ Fontana Masson protocol |
| Argentaffin stains | Gomori methenamine-silver,
Grocott methenamine-silver,
Fontana Masson,
von Kossa |
| Argyrophil stains | Reticulin, Holmes,
Bielschowsky, Warthin-Starry,
Dieterle, Steiner & Steiner,
Grimelius, Churukian-Schenk |
| Formalin pigment | Is a reducing substance,
may give positive rxn to Schmorl & Fontana-Masson |
| Chloroacetate Esterase (Leder) stain purpose | Detection of neutrophils |
| Stained starch granules show: | A characteristic Maltese cross configuration when polarized,
may be introduced from talcum powder of gloves |
| Ferrous ions | Fe2+, toxic, not normally stored,
Turnbull blue |
| Ferric ions | Fe3+, normally present in tissue,
usu. bound to protein (hemosiderin),
in marrow, spleen, decaying, hemorrhagic material,
excess - hemachromatosis,
Prussian blue |
