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Fixation
Fixation in the histology lab.
| Question | Answer |
|---|---|
| Fixative | Alters tissue by stabilizing proteins so that they are resistant to further changes. |
| With additive fixation... | The disruption enables the protein to combine chemically with a fixative molecule; making the protein insoluble/ |
| With non additive fixation...(Alcohol, acetone) | Denaturation causes the protein to become less capable of maintaining an intimate relationship with water and to become more reactive, but the fixative molecule does not combine with protein. |
| Putrefaction | Bacterial attack |
| Autolysis | Enzyme attack |
| In tissue that fails to stain, what is one possible reason for it? | Autolysis |
| Tissues that are rich in enzymes: (Prone to autolysis) | Liver, pancreas, and brain. |
| Fixatives kill _ & _ | Bacteria & molds |
| _ can act as a link between stains and tissue. | Fixatives |
| Microwaves are a form of | nonionizing radiation |
| Additive reagents | Mercuric Chloride, Chromium Trioxide, Picric Acid, Formaldehyde, Glutaraldehyde, Glyoxal, Osmium Tetroxide, and Zinc Sulfate or Chloride. |
| Nonadditive fixatives | Acetone and the alcohols. |
| Heavy Metal Cations (+) | Chromium, Osmium, Mercury |
| Anionic | Negative charge |
| Cationic | Positive charge |
| Coagulation | Mesh ball (penetrates easily) |
| Non Coagulent | Jello, doesn't allow anything to penetrate it easily. |
| Coagulent Fixatives | Zinc salts, mercuric chloride, cupric sulfate, ethyl alcohol, methyl alcohol, acetone, and picric acid. |
| Non coagulent Fixatives | Formaldehyde, glutaraldehyde, glyoxal, osmium tetroxide, potassium dichromate, and acetic acid. |
| After using a non coagulant fixative this type of embedding medium should be avoided due to its lack of ability to penetrate. | Paraffin (Plastics is an example of something that could be used) |
| Fixation Temp for specimens used for Electron Microscopy | 0-4 degrees Celsius |
| Size of specimens should be less than or equal to _mm thick. | 3 |
| Fixative should be _-_ times greater than the tissue volume. | 15-20 |
| Formalin should have at least _to_ hours to fix before completely being processed. | 6-8 |
| Specimens to be fixed for HER2 testing should be fixed in 10%NBF for how long? | Minimum of 6 hours and a maximum of 72 |
| Prolonged sating of HER2 testing could have what effect? | False negative results. |
| HER2 testing is used to help identify... | Invasive breast cancer |
| Tissue must be removed from what fixatives? | Gultaraldehyde, Helly, Zenker, and Bouins. (If tissue stays in these fixative then the tissue will over harden and staining will be impaired.) |
| When is no fixation preferred? | When performing immunoflourescence or an enzyme profile. Then the tissue is frozen. |
| When staining muscle cross-striations are to be stained with PTAH (phosphotungstic acid-hematoxyllin) what fixative should be used? | Zenker or Bouin |
| What Fixative should be used when performing a Trichrome stain? | Bouins |
| Mordant used in the Trichrome stain | Bouins |
| Chromaffin granules are found where in the body? | Adrenal gland |
| Chromaffin granules are helpful in identifying what? | Pheochromocytomas |
| Chromaffin granules should be fixed in what? | Orth or another primary dichromate fixative |
| Urate crystals should be fixed in what? | Absolute Alcohol |
| Formalin penetrates _ but continues to cross link proteins for a long time after the penetration is complete. | Fast |
| Fixative ingredients in order of decreasing penetration | Formaldehyde, Acetic Acid, Mercuric Chloride, Methyl Alcohol, Osmium Tetroxide, and Picric Acid. |
| When the specimen is stored in 10%NBF and additional IHC stains are known to be needed in the future then how should the tissue be stored? | 70% Alcohol, to prevent crosslinking |
| Osmolarity | The number of particles in a solution. |
| Isotonic | Equal state nothing happens with the cell no exchange occurs |
| Hypotonic Solution | Cell Swells; solution drawn into the cell |
| Hypertonic Solution | Cell shrinks; solution is drawn out of the cell |
| Kidney biopsies that need to be sent out for testing are placed in what fixative? | Michels |
| What fixatives are preferred for nucleic acids? | Acetic alcohol and Carnoy |
| Lipids are fixed by? | Osmium Tetroxide and Chromic Acid |
| Penetrates fast and leaves tissue soft | Acetic Acid |
| Fixes nuclei and precipitates DNA | Acetic Acid |
| Makes tissue swell | Acetic Acid |
| Lyses red blood cells | Acetic Acid |
| Formaldehyde is what kind of fixative? | Non coagulant and additive |
| Formaldehyde fixes_ but penetrates _ | Slowly but penetrates fast |
| Formalin pigment can be seen by using polarization; what color does it appear as? | Red |
| Types of Formalin fixation | 1. microcrystalline dark brown pigment (Formed when acid aqueous solution of formaldehyde acts on tissues rich in blood) 2. Black acid hematin (forms when pH drops below 6) |
| Formalin pigment is removed by.... | Alcoholic picric acid or alkaline alcohol |
| Formaldehyde solutions are _ but the formaldehyde molecule is not osmatically active . | Hypertonic |
| 10% Aqueous Formalin | Hypotonic |
| 10% Formalin Saline | Isotonic |
| Formalin Ammonium Bromide | Recommended for Cajal Staining, lyses red blood cells, and allows nuclei to give a direct positive Feulgen reaction due to hydrolysis. |
| Modified Millonig Formalin | This is an isotonic dual purpose fixative. Allows tissue to be stored to be used later for electron microscopy. Harder to cut paraffin embedded tissue. |
| This fixative should not be used when performing a PAS stain. | Glutaraldehyde |
| If tissue for a PAS stain was fixed in Glutaraldehyde the results will be... | A false positive |
| Fixes at the rate it penetrates | Glutaraldehyde |
| Typically used to fix specimens for Electron Microscopy | Glutaraldehyde |
| Over hardens tissue and has poor penetration | Glutaraldehyde |
| Can harden tissue excessively and produce shrinkage | Mercuric Chloride |
| Mercury pigment is removed by | Iodine followed by sodium sulfate |
| Mercury pigment is | Bifringent |
| Preserves lipids | Osmium Tetroxide |
| Coagulent but leaves DNA soluble | Picric Acid |
| Causes extreme shrinkage | Picric Acid |
| Extremely explosive when dry | Picric Acid |
| _ must be completely removed or else all cellular structures will eventually be obliterated. | Picric Acid |
| All reagents should be tightly capped because why? | Many of them will absorb water or lose water through evaporation |
| Tissue is soft | Potassium Dichromate |
| Chromate pigment can be removed by | 1% hydrochloric acid and 70% alcohol for 30 minutes. |
| Typically used for lymph node and bone marrow tissues | B-5 |
| Mercuric Chloride Sodium Acetate Formaldehyde | B-5 |
| Used for hematopoetic and lymphoreticular tissue | B-5 |
| Must be treated for the removal of mercury | B-5 |
| Bouins | Picric Acid Acetic Acid Formaldehyde |
| Lyses RBCs | Bouins |
| Iron and calcium deposits are typically dissolved | Bouins |
| Best fixative for the Trichrome stain | Bouins |
| Yellow color from the Bouin fixative must be removed with | 50-70% alcohol, or 70% alcohol saturated with lithium carbonate. |
| Gendre | 95% alcohol with picric acid Acetic Acid Formaldehyde |
| Excellent for the preservation of some carbohydrates, especially glycogen. | Gendre |
| Excess picric acid can be removed with... | 80% alcohol |
| Hollande | Copper Acetate Picric Acid Formaldehyde Acetic Acid |
| Will decalcify small specimens of bone. | Hollande |
| Cupric acetate stabilizes _ membranes and the _ of esinophils and endocrine cells. (Lysis is less severe) | RBC membranes and the granules of esinophils and endocrine cells |
| This fixative must be washed out before the specimen is placed in a phosphate buffered formalin solution of the tissue processor, because if present the solution will form an insoluble phosphate precipitate. | Hollande |
| Formaldehyde Glutaraldehyde | Monobasic sodium phosphate Sodium hydroxide Formaldehyde Glutaraldehyde |
| Store at 4 degrees Celsius | Formaldehyde Glutaraldehyde |
| Stable for 3 months when stored at 4 degrees Celsius | Formaldehyde Glutaraldehyde |
| Zenker | Mercuric chloride Potassium dichromate acetic acid |
| Helly | Mercuric chloride Potassium dichromate Formaldehyde |
| If formaldehyde is not added immediately before use, the solution will darken and become turbid on standing. | Helly |
| Used to fix and decalcify needle biopsy specimens of bone marrow, but can dissolve iron. | Zenker |
| Fixative recommended for PTAH with the exception of silver staining | Zenker |
| Orth | Potassium dichromate Sodium sulfate Formaldehyde |
| Preferred fixative for demonstration of chromatin granules in the cytoplasm of adrenal cells | Orth |
| Demonstration of granules are important in the diagnosis of... | Pheochromocytoma |
| Granules are typically colored _ to _ with chromate | Orange to brown |
| Zamboni | Paraformaldehyde Picric Acid NaH2PO4 Na2HPO4 |
| Stable fixative | Zamboni |
| Zinc Formalin | Could potentially replace NBF |
| Antigenicity is enhanced and nuclear detail improved. | Alcoholic zinc chloride formalin |
| Which is preferred Zinc Chloride or Zinc Sulfate | Zinc sulfate because it is considered a moderate health risk. |
| Acetone, Alcohol, Carnoy, Clarke | Nonaqueous fixatives |
| Used as a fixative for brain when a rabies diagnosis is suspected. | Acetone |
| OSHA TWA of 1,000ppm and a National Institute for Occupational Safety and Health TWA of 250ppm | Acetone |
| Used as a fixative for touch preps and blood smears | Methyl alcohol |
| Used to preserve water soluble tissue components | Ethyl alchol |
| Water soluble tissue components | Glycogen and urate crystals |
| Preserves pigment, dissolves fat, and over hardens and shrinks tissue | Ethyl alcohol |
| TWA is 1,000ppm for _ alcohol and 200ppm for _ alcohol | ethyl and methyl |
| Carnoy | Ethyl Alcohol Chloroform Acetic Acid |
| Clarke | Absolute Alcohol Acetic Acid |
| Great fixative for paraffin | Clarke |
| Michel medium is recommended for _ biopsies but not _ biopsies. | Kidney but not for muscle. |
| Michel transport medium | Anhydrous citric acid Ammonium Sulfate (MOST) N-ethylmalemide Magnesium sulfate |
| Discard after 6 months | Michel |
| PBS | Potassium phosphate dibasic Sodium phosphate, monobasic Sodium chloride |
| Primary fixatives for ultrastructural studies include | Osmium tetroxide (formaldehyde and glutaraldehyde), and buffered PAF (Zamboni) solutions. |
| Specimen must be removed within 2-4 hours, preferably less | Osmium tetroxide |
| Specimens can be left in this type of fixative indefinately | primary buffered PAF |
| Fixative that can be used for both light and electron microscopy | Primary buffered PAF |
| How to remove formalin pigment | Absolute alcohol with picric acid or 70% alcohol containing ammonium hydroxide |
| How to remove mercury pigment | Treat slides with iodine and then wash with 5% sodium thiosulfate |
Created by:
Ziek98
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