Decalcification Flashcards
What is decalcification
The process of removing minerals from bone or other calcified tissues such as teeth. This is carried out without distorting cells and connective tissue, and without leaving any harmful effects on staining reactions
Importance of decalcification
Softening of the tissue so that it can be sectioned in the gross room and using a microtome. It ensures good quality paraffin sections while preserving all the essential microscopic elements.
Results of over decalcification and under decalcification
under - difficulty sectioninng the tissue
over - lack of nuclear staining, stains very poorly
what is calcified tissue made of
Mad eup of the strong and hard mineral hydroxyapatite. This is an inorganic material
What should be carried out before decalcification
- full fixation
- skin and surrounding tissue removed and separately fixed
- section cut prior using fine thoothed hacksaw
When are undecalcified bone sections examined?
for the diagnosis of metabolic bone disease
What 4 factors influence which decalcifier is used
- urgency of the case
- degree of mineralization
- extent of investigation required
- staining reactions to be carried out
What 2 methods can be used for decalcification
Acid solutions ; nitric acid, HCl or formic acid
or chelating agents such as ethylenediaminetetraacetic acid
How do acids decalcify tissue
they lower the pH causing excess hydrogen ions to combine with hydroxyl ions to give water and calcium ions, which will form salts
Talk about acid decalcification method and how different acids work
They are commonly used at solutions of 5-10%, the stronger the acidity the more maceration of the tissue (swelling, loss of staining ability and poorer morphology)
- HCl and HNO3 - act rapidly. Nitric acid can lead to serious deterioration of tissue. HCl can form a carcinogen (bis chloromethyl ether) if previous formaldehyde is not well washed out.
- formic acid is slower acting.
What 2 methods can be implemented with acids to make decalcification more efficient?
Ion exchange resin - makes use of formic acid over a layer of an ammoniated salt of a sulphonated resin. The calcium is rapdily removed from the solution into the resin.
Electrolytic method - mixture of formic acid and HCl are placed in a glass jar. The specimen is suspended by a platinum wire anode in the jar with temp kept between 30-45 degrees
advantages of the ion exchange resin
well preserved cell detail, fast process, elimination of daily solution change and resin can be reused
How are chelating agents used
they have the property of binding to certain metals such as EDTA which binds to calcium ions for removal. pH is kept between 5 and 7.2
Advantages and disadvantages of chelating agents
adv - can take place at neutral pH, no damage to the tissue and preservation is excellent, allows for staining with most staining techniques.
dis - very slow acting
How can the end point of decalcification be determined
Mechanical, chemical or radiographic methods
