Staining Flashcards
_____________ refers to the procedures in which tissue sections act as the medium in which biochemical reactions are carried out by the addition of substrates, inhibitors, or other chemicals.
Histochemistry
A ___________ is a colouring agent.
Chromogen
The __________ is the component that is stained.
Substrate
___________ substances bond to acidic groups or dyes.
Acidophilic
______________ substances bind to basic groups or dyes.
Basophilic
Give an example of a dichrome stain.
Haematoxylin and eosin
Masson’s trichrome has ____ component dyes.
Three
Staining reactions have both _________ and chemical characteristics.
Physical
Dye may actually be dissolved in the stained substrate; most ____ staining is achieved by this method.
Fat
A dye may be absorbed on the surface of a structure, or may be __________ within the structure.
Precipitated
Give two examples of how union between a dye and stained substance can be brought about.
Salt linkages and hydrogen bonds
True or false: staining is generally predictable, but can vary.
True
State two features which may affect staining efficacy.
Ionic strength of solutions used and temperature
Staining by direct bonding exploits the affinity for _______ found in a cellular component, due to binding and bonding.
Chromogen
Staining by adsorption involves _______ ____ staining.
Metallic ion
What method of staining is used to visualise fungi?
Adsorption
Staining by solubility accounts for ____ staining.
Lipid
____ ____ __ is used to visualise lipids.
Oil Red O
Chemical reactions include insoluble precipitate formation, due to a substrate creating a chromogen at the site of reactivity. What stain can achieve the above?
Perl’s Prussian Blue
Schmorl’s stain uses reducing compounds to attract and reduce ___________ to ferrocyanide.
Ferricyanide
Indirect methods, through antibody- or oligonucleotide probe-binding to the substrate, link to ___________.
Chromosomes
Perl’s Prussian Blue is used for ______.
Iron
Periodic Acid Schiff (PAS) stain is used for __________.
Carbohydrates
List three natural dyes.
Henna, carmine, and haematoxylin
List three synthetic dyes.
Eosin, methyl blue, and neutral red
__________ ________ numbers identify specific dyes.
Colour index
Dyes generally have an _______ structure.
Aromatic
_______________ structure enable the visibility of dyes.
Chromophoric
The chromophore is the portion of a molecule’s structure which absorbs visible light _________, causing the molecule to have colour.
Photons
Nitro dyes, such as picric acid, have a __________ group as the chromophore.
Nitro
_____ dyes, such as Orange G and xanthene, have a -N=N group as the chromophore.
Azo
Acidic dyes bind to basic tissue elements, and exist as _________ in solution.
Anions
_______ dyes bind to acidic tissue elements, and exist as cations in solution.
Basic
________ dyes stain through solubility.
Neutral
Any substance that is stained by basic dye is _________, carrying acids groups which bind the basic dye through salt linkages.
Basophilic
A substance that is stained by an ______ dye is referred to as acidophilic, carrying basic groups to bind acid.
Acid
Wright’s stain is an example of a ________ stain.
Neutral
Neutral stains are ___________ of acid and basic dye.
Compounds
Neutral stains are ___________ of acid and basic dye.
Compounds
The resultant precipitates of neutral dyes are usually insoluble in water, but soluble in _________.
Alcohol
____________ dyes stain one component.
Progressive
____________ dyes overstain all components, and excess must be removed.
Regressive
Differentiation is the microscopic control of dye removal, breaking bonds of _______-bound dye.
Weakly
An accelerator increases staining power, without acting as a __________.
Mordant
Fluorescent dyes emit light under ____.
UV
_____________ stains give different colours to the dye solution.
Metachromatic
Lyoschrome dyes demonstrate lipids, and are _______________ stains that rely on solubility.
Hydrophobic
True or false: microanatomical dyes are used for demonstrating general relationships between cellular components.
True
___________ dyes emphasise minute structure.
Cytological
What is a mordant?
Chemical that serves as a link between the dye and the substrate, resulting in an insoluble compound that helps adhere the dye to the cells
Haematein (from haematoxylin) has poor ________, and must be combined with a mordant, before use as a nuclear stain.
Affinity
Mordants confer a _________ charge, enabling binding to anionic tissue.
Positive
True or false: alum, iron, and tungsten are not examples of mordants.
False
A ‘haematoxylin lake’ results from the combination of a mordant and what substance?
Haematein
Application of silver or gold solution produces black, insoluble _________ deposits, on selected reactive sites.
Metal
Reduction of metal salts to the metallic state by _________ ________ occurs during adsorption staining.
Tissue aggregates
State the principle of Perl’s Prussian Blue stain.
The stain causes production of coloured, chemical compound, through interaction of a series of chemical solutions, with reactive tissue component. Tissue is treated with 2% hydrochloric acid, to liberate ferric iron from protein complexes, and then with potassium ferrocyanide, to allow ferrocyanide ion complexes to interact with ferric ion, to form a blue compound. It is useful in demonstrating haemochromatosis
What is the principle of the Periodic Acid Schiff (PAS) stain?
Firstly, tissue is treated with 1% periodic stain, whereby groups in hexose molecules are oxidised to dialgehyde, then treated with Schiff reagent, to bind dialdehydes, and recolour basic fuchsin, to turn magenta pink
What two modifications must be made to haematoxylin before it is used?
Oxidation, and addition of metal ion
Outline four mordants used with haematoxylin, and why they are used.
Aluminium stains nuclei red initially, which turns to blue-black after washing, and can be used to stain regressively, where the section is overstained, and the tissue is differentiated, then ‘bluing up’ is performed. Iron demonstrates a greater range of structures than aluminium. Tungsten uses 1% phosphotungstic acid as the mordant, staining blue-purple. Lead is useful for demonstration of granules in GI tract tissues
Although primarily used to stain nuclei, ___________ will also stain rough ER, ribosomes, collagen, myelin, elastic fibres, and acid mucins.
Haematoxylin
Haematoxylin may be used progressively, in which the length of time the tissue remains in contact with the staining solution is used to control the amount of colouration, or regressively, in which the tissue is over-stained, and excess stain is removed by _____________.
Differentiation
________ is a fluorescent, xanthene, synthetic dye, which binds salts with eosinophilic compounds containing positive charges.
Eosin
True or false: eosin is negatively-charged.
True
What is eosin’s primary staining target?
Cytoplasm
Most staining is performed in an __________ stainer.
Automated
Participation in external quality programmes, such as _________ , is important.
NEQAS
In Masson’s trichrome, what is Solution A?
Acid fuchsin in distilled water
In Masson’s trichrome, what is Solution B?
Phosphomolybdic acid in distilled water
What is Solution C usually, when found as part of Masson’s trichrome?
Methyl blue
Phosphomolybdic acid acts as a differentiator, forcing _________ of the acid dye out of collagen, while retaining it in the cytoplasm (collagen is less permeable than cytoplasm).
Diffusion
______________ ______facilitates the binding of aniline blue (or other dye) to the decolourised collagen.
Phosphomolybdic acid
What is the order of application of the stains of Masson’s trichrome?
Haematoxylin, Solution A, Solution B, and Solution C
Masson’s technique discerns _________ from muscle fibres.
Collagen
RBC are least permeable, and are coloured by dyes with _________ molecular size in trichrome staining.
Smallest
_______ increases the rate of penetration in trichrome staining.
Heat
Trichrome specimens must be prepared in a _____ pH, to achieve adequate and even staining of connective tissue.
Low
