PS1_Histo_Lect2_part 2_Power Point Presentation.ppt

Standard Methods of Fixation … Standard method of fixation Histological examination for routine processing: Neutral Buffered Formalin Cytological examination: Alcohol fixatives Formaldehyde      Pungent toxic gas soluble in water Very reactive chemical Undergoes changes in storage Sold...

Standard Methods of Fixation … Standard method of fixation Histological examination for routine processing: Neutral Buffered Formalin Cytological examination: Alcohol fixatives Formaldehyde      Pungent toxic gas soluble in water Very reactive chemical Undergoes changes in storage Sold as 37-40% solution of formaldehyde in water = can be often called formalin (originally a company brand) 4% solution of formaldehyde is used for routine fixation: 1 part of formaldehyde to 9 parts of water (1 in 10 dilution) = 10% formalin (also called formalin!) Formaldehyde     Most of the reactions of formaldehyde with amino acids and proteins In aqueous solution, formaldehyde forms methylene hydrate, a methylene glycol as the first step in fixation Methylene hydrate reacts with several side chains of proteins to form reactive hydroxymethyl side groups (CH2-OH) – This is the primary reaction Formaldehyde reacts with nuclear proteins and nucleic acids – It penetrates between nucleic acids and proteins and stabilizes the nucleic acid-protein shell, also modifying nucleotides by reacting with free amino groups as it does with proteins Formaldehyde Formaldehyde reacts with C+C and –SH bonds in unsaturated lipids but does not interact with carbohydrates  Formic Acid formed by oxidation of formaldehyde by the oxygen in the air Or by Cannizzaro reaction, also methanol will form  Polymerizes to form paraformaldehyde  In solution in hydrated form of methylene glycol = acts as a penetrant and fixative  Formaldehyde Slow acting fixative  Adequate fixation in 12-24h (depends on surface area and volume of the fixative)  Fixative in Histology: Formaldehyde Formaldehyde (HCHO) vs formalin: Formalin: formaldehyde+H2O+10-15% methylic alcohol (methanol) + buffers Why? To prevent polymerization of formaldehyde and to inhibit Canizzaro Reaction Is formaldehyde toxic? Yes highly! Absorbed by inhalation Cause damage to respiratory tract Irritation of skin Causes dizziness or suffocation Contact with formaldehyde solution may cause severe burns to the eyes and skin Formaldehyde-releasing preservatives … Many feminine care products contained formaldehyde-releasing preservatives. Formaldehyde is an expected human carcinogen. Women’s Voices for the Earth http://www.womensvoices.org/feminine-care-products/chem-fatale-report/ Always read the labels and try to avoid the nasty chemicals Who was the world’s 14th richest person in 2017? Hint: Daughter of Eugène Schueller (a chemist), inherited his company, died age 94, whose net worth estimated £29bn Who was world’s 14th richest person? Liliane Bettencourt https://www.bbc.co.uk/news/uk-41358593 Formalin ideal anti-bacterial treatment? Formalin: ideal anti-bacterial treatment Most anti-bacterial agents poison the bacteria  Formalin kills by dehydrating the tissue and bacteria cells  Replaces the normal fluid in the cells with a gel-like rigid compound (coagulation properties)  Tissue and bacterium are made of protoplasm Introduction of formaldehyde into the tissue dries out the protoplasm and destroys the cell  Formalin: ideal anti-bacterial treatment A perfect process: not only disinfects, but replaces the water inside tissue with a rigid gel Allows the embalmed tissue to maintain its shape, contour The “new” cell structure will resist further bacterial attacks, as its composition now contains a formaldehyde-based compound Owing to the presence of formalin, histopathology specimens are not normally considered infectious, but are often collected with other samples that may be infectious. So, histology fixation the overall aim … Histology Fixation and processing Overall aim: accurate diagnosis/interpretation  Must have a representative sample  Must have sufficient sample  Must have cells/tissues in life-like condition  All processes should enable sectioning/staining  Sections on slides must still be reactive with stains Other methods of preservation? Gunther von Hagen - Plastination Steps from specimen to report            1. Fixation 2. Specimen collection, transportation and receipt 3. Tissue selection and description 4. Tissue processing 5. Tissue embedding 6. Microtomy 7. Staining and mounting 8. Quality assurance 9. Reporting 10. Specimen disposal 11. Block & Slide archiving Tissue selection and description … Female Reproductive System Whole Organ – resection (surgical specimen) A small representative piece – biopsy specimen Opening of fresh specimen of a uterus for fixation and gross examination video Fixation timeline Fresh Formalin 18h Fixation timeline Fresh Formalin 18h