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Fixation Decalci

Fixation and Decalcification ICLS

Is a process used to kill, harden and preserve tissue materials for microscopic study Fixation
The structure, shape, intracellular relationships and chemical constituents of cells are preserved by preventing degeneration, putrefaction, autolysis, decomposition and distortion of tissues after death, after removal from the body. Fixation
T or F Fixation hardens soft and friable tissues and make the handling and cutting of sections easier. This is usually accelerated by the action of alcohol during the dehydration process T
CHARACTERISTICS of a Good Fixative: Inexpensive, Stable. Safe to handle, It must fix the cell quickly, Inhibit bacterial decomposition and autolysis, minimum shrinkage, must permit rapid and even penetration of tissue, Harden tissue thereby making the cutting easy, It must be isotonic
permit the general microscopic study of tissue structures without altering the structural pattern and normal intercellular relationship of the tissues in question Microanatomical
those which preserve specific parts and particular microscopic elements of the cell itself Cytological
those that preserve the chemical constituents of cells and tissues Histochemical
It is recommended for preservation and storage of surgical, post-mortem and research specimens Neutral Buffered Formalin (10%)
Reagent Preparation buffer for fixatives sodium phosphate
include heating, microwaving and cryo-preservation (freeze drying) Physical Method
is usually achieved by immersing the specimen in the fixative solution (immersion fixation) or, in the case of small animals or some whole organs such as a lung, by perfusing or injecting the vascular system with fixative (perfusion fixation) Chemical Fixation
FACTORS AFFECTING FIXATION DELAYED BY: 1. Size and thickness of tissue 2. Time 3. Presence of Fat, Blood, Mucus 4. pH 5. Cold Temperature
Thickness of Section Tissue blocks should be small ( e.g., 1 to 2 mm) for electron microscopy and 2 cm wide for light microscopy) and thin (not more than 0.4 cm for light microscopy).
Fixation Time 4 - 6 hours
recommended size of the tissue 2 cm, and no more than 4 mm thick.
The best results are usually obtained using slightly hypertonic solutions (mOsm) 400-450 mOsm
T or F Concentration of fixative should be adjusted down to the highest level possible. F
Decalcifying agent ratio 20:1
T or F More concentrated acid solutions decalcify more rapidly and will not destroy tissue. F
TYPES OF DECALCIFYING AGENTS 1. Acids 2. Chelating Agents 3. Ion Exchange Resins 4. Electrophoresis
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