| Question | Answer |
| pH of the staining sollution | cells have specific affinity for stains/dyes with specific pH ranges |
| hypertonic solution will | cause tissue to shrink (ie. picric acid) |
| Nine factors which influence tissue fixation: | Temperature, Size, Tissue/Fixative Volume ratio, Time, Choice of Fixative, Penetration Capability, Tissue storage, pH, Osmolality |
| Fixation changes the weak hydrogen bonds into | stable bonded complexes |
| Putrefaction | Natural & Artificial |
| show positive PAS (Rose Red) staining | Glycogen.
Neutral mucins.
Some epithelial mucins.
Basement membranes.
Fungal walls. |
| glutaraldehyde is s dialdehyde and gives false result for PAS becuase | its 2nd NH2 group is left free to react w/Schiff (which is used to detect aldehydes). |
| T or F: Lipids in the cell are easily fixed. | False |
| a good fixative will... | penetrate tissue quickly + harden the tissue retain micro anatomical structure; maintain histological relationships so the cells can be studied |
| Coagulation/Precipitation render the ________ more resilient. | nucleus.Giving it a sharper more intact appearance. |
| Some _________ are usually LOST during fixation. | carbohydrates |
| Osmolality is defined as... | the # of particles in a solution |
| Kidney biopsy specimens for immunofluorescence frequently are held, or even mailed in ________. | Michel transport solution. |
| hypotonic solution will | cause tissue to swell (ie. acetic acid) |
| T or F: Fixation brings out the differences in refractive indexes of tissues. | True |
| The rate of penetration is NOT affected by _______. | the concentration of the fixative |
| BASIC stains love ACID tissue elements (nuclei & other basophilics) | ACID stains love BASIC tissue elements (cytoplasm & other acidophilic) |
| Alcian Blue solution with a pH of 2.5 | stains CARBOLYLATED mucins in connective tissues and cartilage. |
| Biopsy specimen which cannot be immediately placed in fixative should be. | placed in a saline dampened gauze, put in plastic container and put on ice for the short holding time. |
| Artificial dyes are | from chemical reactions (greatly outnumber natural dyes) |
| Fixatives in order of decreasing speed of penetration: | formaldehyde-acetic acid-mercuric chloride-methy alcohol-osmium tetroxide-picric acid |
| Only 2 chemicals can fix lipids so they are not lost in subsequent processing. | (1) Osmium tetroxide (2) Chromic Acid |
| in the case of staining, FIXATION | alters and reorganizes certain molecular structures in tissue samples so that they have an increased permeability and are more receptive to staining. |
| Alcian Blue solution with a pH of 1.0 | stains most SULFATED acid mucins. These mucins can be found in cartilage, large intestine goblet cells and bronchial serous glands. |
| pH scale: 1 - 14 | 1-6.9 ACIDIC.
7-7.4 NEUTRAL.
7.5-14 BASIC. |
| Let's look at Stains...momentarily | stabilization of proteins in tissue |
| Acetic acid warnings: | add TO water; use under a hood; causes severe burns; exposure limit of 10ppm |
| Natural dyes are | from natural resources (ie. HEMATOXYLIN) |
| Stabilization of proteins is done in order to prevent | tissue destruction during subsequent steps in processing |
| Mucicarmine | rose red dye in the mucicarmine staining solution gets its color from the aluminum-carminic acid complex carmine |
| The most important step in the histology laboratory | fixation |
| Cross links makes tissue suitable for microtomy by... | strengthening and hardening the tissue |
| Mucicarmine results | Nuclei are stained black with Weigert's Iron Hematoxylin and the remaining background tissue elements are stained yellow with Metanil Yellow or Tartrazine. |
| Dyes are substances... | capable of imparting color chemically or physically binding with materials |
| T or F: Unfixed tissue elements have LIMITED binding sites for dyes. | TRUE |
| The cell is composed of | proteins, carbohydrates and lipids held loosely together by hydrogen bonds |
| Tissue FIXATION | |
| PAS counterstain with _______ hematoxylin | HARRIS hematoxylin with Acetic Acid |
| stabilization is accomplished by | denaturing of proteins |
| Fixation is the... | breakdown of tissue by bacterial action from within. also called postmortem decomposition |
| Formaldehyde (manufacturer label) = Formalin (37%-40% in water) | 37%-40% formalin |
| BUFFERS | prevent fluctuations in pH |
| NON-coagulant fixatives produce a gel that makes penetration difficult.Examples are... | formaldehyde, glutaraldehyde, acetic acid, potassium dichromate, osmium tetroxide |
| Mucicarmine is also used to identify | ADENOCARCINOMAS & Cryptococcus neoformans |
| Fixation causes ________ linkage of protein molecules. | cross linkage |
| TEMPERATURE of the dye | increased temperature = increased rate of diffusion throughout the tissue |
| Dyes impart color due to | the presence of color bearing chemical groups called CHROMOGENS |
| normal osmolality | isotonic |
| Alcian Blue | has an affinity for acidic tissue elements like mucin. The dye gets color from the copper in the molecule |
| 2 categories of dyes... | fixation alters and reorganizes certain molecular structures in tissue samples so that they have an increased permeability and are more receptive to staining. |
| Biological Stain Commission | agency responsible for ensuring the quality of biological stains and -PROMOTE COOPERATION AMONG VENDORS
-EDUCATE USERS OF STAINS
PUBLISH INFO CONCERNING NEW OR IMPROVED STAINS |
| CONCENTRATION of the dye | the greater the concentration, the more the dye is bound to tissue components |
| Acetic acid lyses _________, and thus their preservation is poor in in any fixative containing it. | red blood cells |
| Formaldehyde is colorless ________. | gas |
| Coagulant fixatives establish a network in tissue allowing penetration. Examples are... | Mercuric chloride; zinc salts; chromium trioxide; picric acid; and POTASSIUM DICHROMATE AT PH > 3.5 |
| PAS with Diastase | used in conjunction with the PAS staining procedure to specifically identify glycogen granules in tissue samples. Amylase enzymes in a malt extract are used to digest the glycogen, which is then washed out of the tissue sample. |
| OXIDATION = removal of electrons from molecules | REDUCTION = adding electrons |
| Acetic Acid is a non-coagulant and... | Does NOT fix or destroy carbohydrates. Does NOT fix lipids. YES penetrates slowly. YES preserves nucleoproteins. |
| Intent of fixation is: | preserve tissue in life-like state;prevent breakdown of cellular components; coagulate/precipitate proteins (insoluble);keep from losing proteins during processing |
| Glutaraldehyde fixes ____ and binds ________. | fixes slowly and binds slowly |
| ABsorption | a PHYSICAL process by which the dye DISSOLVES DIRECTLY INTO elements int he sample. |
| Autolysis | dissolving of cells by enzyme action |
| The rate of penetration is affected by _______. | heat |
| A ________ - storage form of _________ is retained during fixation. | A glycogen-storage form of glucose is retained during fixation. This is due to the entrapment by fixed proteins. |
| ADsorption | which the dye BINDS TO ELEMENTS IN THE SAMPLE that have an affinity for the dye. |
| Biological stain commission DOES NOT | manufacture biological stains |
| RNA and DNA are entrapped in the nucleus by the process of ... | fixation |
| Alcian Blue used with PAS | both acid and neutral mucins can be demonstrated. Alcian Blue will stain acidic mucins blue. PAS will stain neutral mucins rose red. This technique is particularlyuseful in diagnosing diseases of the gastrointestinal (GI) tract. |
