HISTOPATHOLOGY & MT LAWS

• The purpose of the fixative is to preserve tissue structure.
• Immunohistochemical techniques make use of antigen–antibody interactions, whereby the site of antigen binding is demonstrated by direct labeling of the antibody, or by means of a secondary labeling method.
• Animals used for the production of polyclonal antibodies include rabbit, goat, pig, sheep, horse, and guinea pig, with the rabbit being the most commonly used.
• Polyclonal antibodies are not identical to each other.
• Polyclonal antibodies are produced by immunizing an animal with a purified specific molecule (immunogen) that contains an antigen of interest and collecting immunoglobulin-rich serum.
• Immunohistochemical techniques are used to identify specific or highly selective cellular epitopes or antigens in frozen or paraffin-embedded tissues.
• Citric acid-citrate buffer does not produce cell or tissue distortion and is not recommended for fluids containing mineral acids such as nitric or hydrochloric acids.
• Formic acid sodium citrate (20% Aq Sodium citrate + 45% Formic acid) is recommended for autopsy materials, bone marrow, cartilage, and tissues studied for research purposes.
• Trichloroacetic acid (TCA) does not require washing out and is not recommended for urgent examinations due to its slow-acting nature.
• Sulfurous acid is the weakest decalcifying agent, suitable only for minute pieces of bone.
• Ion exchange resin, an ammonium form of polystyrene resin, hastens decalcification and removes calcium ions from formic acid, thereby increasing solubility from the tissue.
• Formic acid has a decalcification time of 2-7 days and is a moderate-acting decalcifying agent, recommended for routine decalcification of post-mortem research tissues.
• Electrical ionization (Electrophoresis) is a process where positively charged calcium ions are attracted to a negative electrode (cathode) and subsequently removed from the decalcifying solution.
• Solutions used in electrical ionization include 88% Formic acid, Concentrated HCl, and Distilled water.
• Hydrochloric acid, also known as muriatic acid, is inferior to nitric acid and is recommended for surface decalcification of tissue blocks if used in a 1% solution with 70% alcohol.
• The time required for decalcification is shortened due to the heat and electrolytic reaction produced in this process.
• Perenyi’s fluid is used for decalcification with a decalc time of 1-3 days and is recommended for urgent biopsies with good nuclear staining.
• Chromic acid is used as a fixative and decalcifying agent and should be handled with caution due to its environmental toxicity, corrosive nature, and carcinogenic properties.
• The principle applied in electrical ionization is similar to that of chelating agents with the main difference being that this process utilizes electricity and is dependent upon a supply of direct current to remove calcium deposits.
• Von Ebner’s Fluid, a mixture of 36% Saturated aqueous NaCl, concentrated HCl, and distilled water, is recommended for teeth and small pieces of bones.
• Detection of calcium in acid solutions occurs by precipitation of insoluble calcium hydroxide if ammonium hydroxide was used or calcium oxalate if ammonium oxalate was used.
• Solutions used in the Chemical Method of Measuring Extent of Decalcification include concentrated ammonium hydroxide and saturated aqueous ammonium oxalate.
• Physical/Mechanical Method of Measuring Extent of Decalcification involves touching or bending the tissue with the fingers to determine the consistency of tissues.
• Tissue softeners are used in cases of unduly hard tissues which are reliable to damage the microtome knives.
• Presence of cloudiness in the Chemical Method of Measuring Extent of Decalcification indicates the presence of calcium (incomplete decalcification).
• Chemical Method of Measuring Extent of Decalcification, also known as calcium oxalate test, is a simple, reliable, and convenient method recommended for routine purposes.
• X-ray/Radiological Method of Measuring Extent of Decalcification is very expensive, but it is the most ideal, most sensitive, and most reliable method.
• In the Chemical Method of Measuring Extent of Decalcification, decalcifying fluid is changed every 24–48 hours.
• Perenyi’s fluid acts both as a decalcifying agent and tissue softener.
• Molliflex (tissues may appear swollen and soapy) is another tissue softener.
• The X-ray/Radiological Method of Measuring Extent of Decalcification is not always convenient and is not recommended for mercuric chloride-fixed tissues because radio-opacity will interfere with the plate interpretation.
• 2% HCl is another tissue softener.
• 4% phenol (1–3 days) is another tissue softener.
• 1% HCl in 70% Alcohol is another tissue softener.
• Another method of Measuring Extent of Decalcification is by pricking the tissue with a fine needle/probe.
• Fixation/Preservation is the process by which the constituents of the cells, and therefore of the tissues are fixed in a physical, and partly also in a chemical state so that they will withstand subsequent treatment with various reagents with minimum loss or significant distortion or decomposition.
• The secondary goal of fixation is to harden and protect tissue from the trauma of further handling and for easy cutting during gross examination.
• The most important reaction in fixation is stabilization of proteins, achieved by forming cross-links between proteins.
• A single uninterrupted motion of pulling apart is applied.
• In non-additive fixation, the fixing agent is not incorporated into the tissue but alters the tissue composition and stabilizes it through water removal, examples include Alcoholic fixatives.