Fixation (M) Flashcards
What is fixation?
It is the process of preserving or stabilizing, by inactivating enzymes and in turn halting autolysis
*What are the results of fixation?
1) It kills bacteria halting putrefaction
2) It makes tissue more receptive to dyes
3) It makes tissues able to retain its cell form
4) It alters the tissue by stabilizing the protein so that it is resistant to further changes
What must be done by fixation?
It must change the solute substances so that those substances are not lost during the subsequent processing steps
What is denaturation?
It causes the protein to unfold and the internal bonds to become disrupted
What is the 1st and most critical step in tissue fixation?
Fixation
Why is fixation considered the most critical step in tissue fixation?
Because once this process is wrong, the succeeding step will be affected because the tissue is not preserved correctly
What is the primary aim of fixation?
To preserve the tissue
What is the other aim of fixation?
Preventing the degeneration, decomposition, or distortion of the tissue after its removal in the body
Once the tissue is removed in the pt’s body, what should be done?
It must be put in a fixative as soon as it is removed in the body
What will happen to the tissue after it is removed in the body?
The cells starts to die, once they die, the chemical and structural changes takes place
What will happen to the tissue if it is left in the air for a long period of time?
There is a dehydration, hence, the tissue will dry out and cells become distorted
What is the 2ndary aim of fixation?
To harden and protect the tissue from trauma of further handling
What are basic mechanisms involved in fixation?
1) Additive fixation
2) Nonadditive fixatives
3) Coagulant
4) Noncoagulant
*What is the principle of additive fixation?
This disruption enables the protein to combine chemically w/ fixative molecules
What is the result of additive fixation?
Protein becomes insoluble
What are the exs of nonadditive fixatives?
1) Alcohol
2) Acetone
*What is the principle of nonadditive fixatives?
Denaturation causes protein to become less capable of maintaining an intimate relationship w/ H2O and to become more reactive, but the fixative molecule does not combine the protein
*What is the action of coagulant?
It establishes a network on tissue that allows solutions to easily penetrate
What are the characteristics of noncoagulant?
1) It is not easy to penetrate
2) It creates a gel called jello
*What are the types of fixatives?
1) Additive
2) Non-additive
*What are the types of additive?
1) Noncoagulants
2) Coagulants
What are the exs of additive noncoagulants?
1) Formaldehyde
2) Glutaraldehyde
3) Glyoxal
4) Osmium tetroxide / osmic acid
5) Potassium dichromate
What are the exs of additive coagulants?
1) Mercuric chloride
2) Chromic acid
3) Picric acid
4) Zinc salts
5) Cupric salts
6) Alcohols
7) Acetone
8) Acetic acid (texts differ)
What are the exs of non-additive?
1) Alcohols
2) Acetone
3) Acetic acid
What are the hallmarks of a good fixation?
1) No cell shrinkage or swelling
2) Crisp nuclear membrane
3) No nuclear bubbling, or smudginess
4) Cytoplasm should be intact and stain well
5) Visible chromatin patterns
6) No artifactual space between cells
What are the characteristics of a good fixative?
1) Must be cheap
2) Must be stable
3) Must be safe to handle
4) Must kill the cell quickly
5) Must inhibit bacterial growth
6) Must produce min. shrinkage
7) Must permit rapid and even penetration of tissues
8) Must permit subsequent application of many staining procedures
What is the importance of the characteristic of a good fixative w/c is that it must kill the cell quickly?
It should be the 1st one before the cell commit apoptosis for it, there will be changes in the components of the cell and they cannot produce similar tissue
Why should a good fixative must produce min. shrinkage?
Because shrinkage produce distortion
Why should a good fixative must permit subsequent application of many staining procedures?
Because some fixative is not compatible w/ some stains
A good fixative must permit application of many staining procedures, so far there is no single fixative has these qualities
What are the fxns of fixatives?
1) Penetration
2) Kill
3) Hardening
4) Help maintain the proper relationship between cells and extracellular substances
5) Brings out differences in refractive indexes
6) Increases the visibility or contrast between different tissue elements
What is the purpose of penetration (as a fxn of fixatives)?
It ensures fixation of the interior of the tissue as well as the few exterior cell layers
What is the purpose of kill (as a fxn of fixatives)?
To kill the tissue so that postmortem activities are prevented
What are the processes that can prevent postmortem activities as a result of killing the tissue (w/c is a fxn of fixative)?
1) Putrefaction
2) Autolysis
What is putrefaction?
It is the bacterial attack
What is autolysis?
It is the enzyme attack
In the fxn of fixative w/c is hardening, much of the section was done in what manner?
Done freehand
The fxn of fixative w/c is to help maintain the proper relationship between cells and extracellular substances, what are the cells affected?
Such as connective tissue fibers and amorphous ground substance
What are the exs of connective tissue fibers whereas its proper relationship is helped maintained by fixatives?
1) Collagen
2) Reticulin
3) Elastin
What are the actions of fixatives?
1) Enzymes are rendered inactive as a result of the protein-stabilizing action of fixatives, preventing further autolysis
2) Fixatives kill bacteria and molds w/c cause putrefaction
3) It makes the tissue more receptive to dyes and act as mordants, w/c serve to link the dye to the tissue
4) It modify tissue constituents for the max. retention of form through subsequent processing steps
What are enzymes?
These are proteins
What are the factors affecting fixation?
1) Temperature
2) Thickness of section
3) Volume ratio
4) Time / duration of fixation
5) Choice of fixative
6) Penetration
7) Tissue storage
8) Hydrogen Ion concentration / pH
9) Osmolality
Many labs use tissue processors that work at what temp for tissue processing?
40 DC
What is the ideal temp for electron microscopy (EM) and some histochemistry?
0 - 4 DC
Generally, fixation at what temp is sufficient to maintain excellent morphological detail?
Room temp
Some cells such as mast cells are best fixed at what temp even for EM?
Room temp
What is the fxn of refrigeration?
It is used to slow down decomposition if the tissue needs to be photographed and cannot be fixed immediately
True or False
Cells from different parts of the body decompose at different time intervals
True
What cells deteriorate very quickly?
Brain cells
True or False
Bone marrow continues to undergo mitosis (growth) up to 40 mins after death when refrigerated
False, because one marrow continues to undergo mitosis (growth) up to 30 mins after death when refrigerated
At any extent at room temp, does nucleic acids react w/ fixatives?
No
Chemical rxns including those involved in fixation are more rapid at what temp?
At higher temps
What is used for fixation in bacteriology and for blood films w/c is well-known?
Heat
Formalin heated to 60 DC is sometimes used for what?
For the rapid fixation of very urgent biopsy sxs
What is the bad effect of using formalin heated to 60 DC for the rapid fixation of very urgent biopsy sxs?
The risk of tissue distortion is increased
What is the result of increasing the temp, as w/ chemical rxns?
It will increase the speed of fixation, as long as you don’t “cook” the tissue
What is the result of an increase in temp?
It can increase the rate of fixation, but it can also increase the rate of autolysis
What are the effects / results of increasing the temp?
1) It increases the rate of fixation
2) But it also increases the rate of autolysis
3) It also increases the rate of diffusion of cellular elements
4) It will increase the rate of penetration
What should be the time duration that formalin should have for it to act before the remaining of the processing sched continues?
6 - 8 hrs
What does temp affects?
It affects morphology
*What is the temp whereas autolysis and diffusion of cell components occur in standard histology?
Room temp
*What is the temp whereas autolysis and diffusion of cell components occur in EM?
0 - 4 DC
What is the relationship between tissue being thick and penetration of the fixative?
If the tissue is thick, the penetration of the fixative is slower`
What is the relationship between tissue being thin and penetration of the fixative?
If the tissue is thin, the fixation will occur immediately
In EM, what is the optimal thickness of the tissue?
1 - 2 mm^2
In light microscopy (LM), what is the optimal thickness of the tissue?
2 cm^2
In LM, what is the optimal width of the tissue?
No 0.4 cm >
True or False
The volume of fixative is not impt
False, because the volume of the fixative is impt
What is the traditional amt of fixative that is used in relation or in coordination to the volume of tissue to be fixed?
Traditionally, the amt of fixative used has been 10 - 20 times the volume of tissue to be fixed
What happens as the fixative molecules bind to the tissue?
The amt / volume of fixative (/ fixative molecules) are even depleted
What are the often results of the depletion of fixative molecules because these binds to the tissue?
1) It often results to poor fixation
2) It can result in staining artifacts
What is the resolution for the depletion of fixative molecules because these bind to the tissue?
One way to partially solve this is to change the solution at intervals to avoid exhaustion of the fixative
What can be done to enhance the fixation of the sx?
Agitation
What is the most common error in histotechnology?
Insufficient ratio of tissue volume to fixative volume
What should be the fixation volume?
It should be at least 15 - 20 times greater than the tissue volume
What is the dilution of fixation volume being 15 - 20 times greater than the tissue volume?
20:1
What should be the thickness of the tissue?
It should be not 3 - 5 mm thick >
What should be observed in the tissue cassette containing the tissue sx?
The tissue should never touch the top and bottom of the cassette
What is the result of poor fixation?
Poor staining
After surgical removal, what is ideal to be done to tissue?
It should be placed in fixative immediately
What is preferred to be done immediately after death?
Autopsies
True or False
Tissue that is not well-fixed can still be processed and stained well
False, because tissue that is not well-fixed does not process well and will not stain well
Incomplete fixation is often indicated by what?
By the appearance of smudgy nuclei
The appearance of smudgy nuclei is present or can be observed where?
In H&E stained sections
Incomplete fixation before going into the alcohols used in processing will result in what?
In nuclear bubbling-nuclei
What is the characteristic of the nuclear bubbling-nuclei present (w/c is a result of incomplete fixation before going into the alcohols used in processing)?
It has a soap-sud appearance
What is the result if the tissue is present in fixative for too long?
The tissue may become over-hardened
True or False
Most fixatives will penetrate tissue to the depth of approximately 1 mm in one hour
True
*What are the 3 results that can be observed in relation to the time / duration of fixation?
1) Artifacts-free-sections
2) Proper fixation
3) Incomplete fixation
What is the min. time duration whereas the artifacts-free-sections could be produced?
Artifacts-free-sections could be produced only after a min. of 30 - 40 hrs of fixation
*What should be used as a fixative whereas artifacts-free-sections are only produced after a min. of 30 - 40 hrs?
NBF (Dapson)
What is the result of proper fixation?
The tissue is almost immune to artifacts
What is the result of incomplete fixation?
The sx is vulnerable to the effects of any subsequent denaturing agent
A fixative may be chosen to ensure what?
To ensure optimal demonstration of a particular tissue element
Provide an ex of the application of choosing a fixative
Zenker solution when muscle cross-striations are to be stained w/ PTAH or Bouin solution when the tissues are to be stained w/ a trichrome technique
What must be the type of fixative and the fixative itself used for urate crystals?
Nonaqueous fixative: 100% alcohol
When are urate crystals present?
If the pt has gout
*What are the fixatives used for trichome staining?
1) Mercuric chloride
2) Potassium dichromate
3) Sodium sulfate
4) Distilled H2O
5) Glacial acetic acid
What affects penetration (w/c is a factor that affects fixation)?
Heat
Does the concentration of fixative affect penetration (w/c is 1 of the factors affecting fixation)?
No
What are the fixatives (arranged) in order of speed of penetration?
1) Formaldehyde
2) Acetic acid
3) Mercuric chloride
4) Methyl alcohol / ethyl alcohol
5) Osmium tetroxide
6) Picric acid
True or False
Additional studies may still be possible if the tissue has not been fixed and stored properly
False, because additional studies may be impossible if the tissue has not been fixed and stored properly
For immunohistochemistry (IHC) stains, what should be done to the tissue?
It should be transferred from formalin to 70% alcohol
Why should the tissue (for IHC stains) be transferred from formalin to 70% alcohol?
To stop cross-linking
*What is the optimal pH of fixatives?
Between 6 - 8
True or False
In fixatives, the more neutral the pH, much better
True
What occurs in formalin fixation if the pH is acidic?
A pigment
What is the factor that is very impt for EM?
Hydrogen ion concentration / pH
*What should be the pH of fixative for EM?
7.2 - 7.4
What happens to the structure of the tissue if the pH of fixative used is between 4 and 9?
It’s barely altered
What is osmolality?
It is the # of particles in solution and is not as impt in light microscopic studies as in ultrastructural studies
What is the osmolality of body fluids?
About 340 mOsm or 0.3 Osm
What happens if hypotonic fixative is used (/ what is the action of hypotonic fixative)?
H2O will be drawn into the cell in an effort to equalize the tonicity
What are the effects of the action of hypotonic fixative?
1) It will cause swelling
2) It will also cause possible rupture of the cell membrane
What happens if hypertonic fixative is used (/ what is the action of hypotonic fixative)?
H2O will be drawn from the cell in an attempt to equalize the tonicity
What is the effect of the action of hypertonic fixative?
It will cause shrinkage of the cell
What happens if isotonic fixative is used (/ what is the action of isotonic fixative)?
The cell will neither shrink nor swell
What are the types of fixation?
1) Heat fixation
2) Microwave fixation
3) Freeze-drying and freeze substitution
What is the characteristic of heat fixation?
It is the simplest form of fixation
What is done in heat fixation?
Picking up a frozen section on a warm slide- partially fixed by heat and dehydration
What are the results of microwave fixation?
1) It speeds fixation
2) It reduces time from 12 hrs > to < 20 mins
What are the types of fixative accdg to composition?
1) Simple
2) Compound
What is simple fixative?
It is made up of 1 active ingredient only
What are the exs of simple fixative?
1) Formaldehyde
2) Glutaraldehyde
3) Mercuric chloride
4) Acetone
5) Alcohol
What is compound fixative?
It is the combination of 2 simple fixatives
What are the exs of compound fixative?
1) Gendre’s solution
2) Formol-corrosive
What are the types of fixative accdg to action?
1) Microanatomical fixatives
2) Cytological fixatives (Nuclear)
3) Histochemical
What is the action of microanatomical fixatives?
It permits the general microscopic study of the tissue structure
What is the action of cytological fixatives (/ cytological)?
It covers cells w/ a tough, soluble film that protects cell morphology for microscopic examination
What is the action of nuclear (/ nuclear fixative)?
It preserves the nucleus
What is the action of histochemical (/ histochemical fixative)?
It preserves the chemical constituents of the cells
What are the chemical constituents of the cells that are preserved by histochemical fixative?
1) Fats
2) Carbs
What are the types of fixative accdg to active component substance?
1) Aldehydes (glutaraldehyde and formaldehyde)
2) Metallic fixatives (lead, chromate, and mercuric chloride)
3) Glacial acetic acid
4) Acetone
5) Alcohol
6) Picric acid
7) Osmium tetroxide
8) Trichloroacetic acid
What are the types of aldehyde?
1) Formaldehyde
2) Glutaraldehyde
What are the types of fixatives that contain formaldehyde?
1) 10% Neutral buffered formalin (NBF)
2) 10% Formol saline
3) Formol-corrosive
4) Gendre’s solution
What is the most widely used type of fixative?
Formaldehyde
What is the optimal concentration of formaldehyde?
10% formalin
True or False
Formaldehyde is not delivered in the lab as 10%
True
What is the concentration of pure stock solution of formaldehyde?
40% concentration
What is the effect of 40% concentration of pure stock solution of formaldehyde?
It over hardens the tissue
What is the characteristic of formaldehyde?
It is known as tolerant fixative
Why is formaldehyde known as tolerant fixative?
Because it does not overharden tissues, even w/ prolonged periods of time
What is the use of formaldehyde?
It is used for mailing for it can leave fixative for a long period of time
What are the advantages of using formaldehyde?
1) It is cheap
2) It is easy to prepare
3) It is relatively stable
4) It is compatible w/ many staining procedures
5) It does not overharden tissues, even w/ prolonged periods of time
6) It penetrates the tissues well
7) It preserves but not precipitate proteins, thereby making enzyme studies possible
8) It preserves glycogen, fat, and mucin
What are the disadvantages of using formaldehyde?
1) It produces fumes w/c are irritating to the nose, eyes, and skin
2) It may produce considerable tissue shrinkage
3) It is a soft fixative
4) If unbuffered, it may form abundant brown pigment granules on blood containing tissues
True or False
Formaldehyde is not buffered
True
If formaldehyde is not buffered, what should be done for it to be buffered and what is its effect / result?
For it to be able to be buffered, the solution must be phosphate buffer w/c is added into the formalin to maintain the pH of 7
10% NBF is recommended for what?
Preservation and storage of surgical, post-mortem, and research sxs
What are the advantages of using 10% NBF?
Its advantages are same w/ formol-saline w/ the addition of it being the best fixative for containing iron pigments and for elastic fibers
Also, it requires no post-treatment after fixation
What are the disadvantages of using 10% NBF?
1) It takes longer to prepare
2) Mucin is not well demonstrated
3) It may produce gradual loss in basophilic staining of cells
What is 10% formol saline and what is its composition?
It is a simple microanatomical fixative w/c is made up of saturated formaldehyde diluted w/ 10% sodium chloride
The use of 10% formol saline is recommended for what?
For fixation of central nervous tissues and general post-mortem tissues for histochemical fixation
What are the advantages of using 10% formol saline?
1) It preserves microanatomic and cytologic details w/ min. shrinkage
2) It penetrates the tissues well
3) It demonstrates fats, mucins, enzymes, and nucleoproteins
4) It is ideal for most staining techniques, including silver impregnation
5) It does not overharden the tissue
What is the composition of formol-corrosive?
Saturated formaldehyde + Saturated aqueous mercuric chloride
Formol-corrosive is recommended for what?
For post-mortem tissues
What are the advantages of using formol-corrosive?
1) It penetrates small tissues rapidly
* 2) The duration for it to penetrate small tissues is 3 - 24 hrs
3) It is excellent for many staining procedures including silver reticulum method
4) Cytological structures and blood cells are well preserved
5) There is no need for washing out
6) It fixes lipids, especially neutral fats and phospholipids
What is the disadvantage of using formol-corrosive?
If normal tissue are being cut thick (1 cm) the fixative it slow penetrate
What is the disadvantage of using formol-corrosive?
If normal tissue are being cut thick (1 cm) the fixative will slowly penetrate it
What is the composition of gendre’s solution?
95% ethyl alcohol + Formaldehyde + Glacial acetic acid
What are the actions of gendre’s solution?
It can fix and dehydrated at the same time
What are the advantages of using gendre’s solution?
1) It is used to fix sputum since it coagulates mucus
2) It can be used for rapid dx
3) It is good for preserving glycogen and for micro-incineration technique
What are the disadvantages of using gendre’s solution?
1) It causes partial lysis of RBCs
2) Its preservation of iron-containing pigments is poor
What is the composition of glutaraldehyde?
It is made up of 2 formaldehyde residues, linked by 3 carbon chains
What are the advantages of using glutaraldehyde?
1) It has more stable effect on tissues, especially when fixing central nervous tissues
2) It preserves plasma proteins better
3) It produces less tissue shrinkage
4) It preserves cellular structures better
5) It is recommended for enzyme histochemistry and EM
6) It does not cause dermatitis
7) It is less irritating to nose and eyes
What is the disadvantage of using glutaraldehyde?
It is more expensive than formaldehyde
What are the types of metallic fixative?
1) Mercuric chloride
2) Chromate
3) Lead
*What are the types of metallic fixatives that contains mercuric chloride?
1) Zenker’s fluid
2) Zenker-formol (Helly’s)
3) Heidenhain’s susa
4) B5
*What are the types of metallic fixatives w/c contains chromate?
1) Chromic acid
2) Potassium dichromate
3) Regaud’s (Moller’s)
4) Orth’s
What are the advantages of using mercuric chloride?
1) It penetrates and hardens tissues well
2) The nuclear components are shown in fine detail
3) It precipitates all proteins
4) It has great affinity to acid dyes
5) It is preferred in lieu of formaldehyde for cytoplasmic staining
6) Trichrome staining is excellent
7) It is recommended for tissue photography
8) It is recommended for renal tissues, fibrin, connective tissues, and muscles
What are the disadvantages of using mercuric chloride?
1) It causes considerable lysis of RBCs
2) It is inert to fats and lipids
3) It leads to the formation of black granular deposits
* 4) Remedy using alcoholic iodine
5) It is extremely corrosive to metals
What is the composition of zenker’s fluid?
It is made up of mercuric chloride w/ glacial acetic acid
Zenker’s fluid is recommended for what?
Trichome staining
The use of zenker’s fluid causes what?
Lysis of RBCs
Zenker’s fluid may form what?
Mercuric pigment deposits
Zenker’s fluid is used in what?
Small pcs of:
1) Liver
2) Spleen
3) Connective tissues
*What is the other name of zenker-formol?
Helly’s
What is the composition of zenker-formol?
Mercuric chloride + Potassium dichromate + Formaldehyde
Zenker-formol is recommended for fixing what?
1) Pituitary gland
2) Bone marrow
3) Blood containing organs
Heidenhain’s susa is recommended for what?
Tumor biopsies
What is the characteristic of Heidenhain’s susa?
It is an excellent cytologic fixative
Heidenhain’s susa permits what?
It permits easier sectioning of large blocks of fibrous connective tissues
Heidenhain’s susa is used for what?
Skin biopsies
B5 is recommended for what?
Bone marrow biopsies
What is the active component of chromate?
Potassium dichromate
What is the characteristic of chromic acid?
It is a strong oxidizing agent
What is the action of chromic acid?
It precipitates all proteins
True or False
Chromic acid is not used as simple fixative
True
If chromic acid is used, what is the concentration of the solution that is used?
1 - 2% solution
What are the actions of potassium dichromate?
1) It preserves lipids well
2) It preserves mitochondria
*What is the other name for Regaud’s?
Moller’s
Regaud’s is recommended to be used for what?
For the demonstration of mitochondria, mitotic figures, golgi bodies, RBC, and colloid containing tissues
Orth’s is recommended to be used for what?
For the study of early degenerative processes and tissue necrosis
What are the actions of Orth’s?
1) It demonstrates rickettsiae and other bacteria
2) It preserves myelin better than buffered formalin
Lead is recommended to be used for what?
For the demonstration of acid mucopolysaccharides
What is the action of lead?
It fixes connective tissue mucin
*What is the concentration of the solution of lead that is used?
4% solution
What are the actions of picric acid?
1) It may also dye the tissues yellow
2) It removes the yellow stain by treatment to another dye w/c is lithium carbonate
3) It removes the yellow stain by 70% ethyl alcohol, 5% sodium thiosulfate, and running H2O
4) It preserves glycogen well but cause considerable damage
5) It precipitates all proteins
6) It causes RBC lysis
7) It is not suitable for frozen tissue sections
8) It must never be washed w/ H2O before dehydration
9) It is explosive when dry
What are the types of picric acid fixative?
1) Bouin’s solution
2) Brasil’s solution
3) Hollande’s solution
The use of Bouin’s solution is recommended for what?
For embryos and pituitary biopsies
What is the composition of Bouin’s solution?
Picric acid + Formaldehyde + Glacial acetic acid
What are the advantages of using Bouin’s solution?
1) It is excellent for preserving soft and delicate tissues
2) It causes less shrinkage
3) It does not need washing out
What are the disadvantages of using Bouin’s solution?
1) It is not suitable for fixing kidney structures, lipid, and mucus
2) It lyses RBCs
3) It penetrates large tissues poorly
What is the composition of Brasil’s solution?
Picric acid + Formaldehyde + Alcohol
What are the advantages of using Brasil’s solution?
1) It is better and less messy than Bouin’s solution
2) It is an excellent fixative for glycogen
The use of Hollande’s solution is recommended for what?
For fixing GIT biopsies and endocrine tissues
Glacial acetic acid is normally used in what?
In conjunction w/ other fixatives to form a compound solution
What are the advantages of using glacial acetic acid?
1) It fixes and precipitates nucleoproteins
2) It precipitates chromosomes and chromatin materials
What are the disadvantages of using glacial acetic acid?
1) When combined w/ potassium dichromate, the lipid-fixing property is destroyed
2) It destroys cytoplasmic structures
Alcoholic fixative may be used in what concentrations?
Ranging from 70% - 100%
What are the advantages of using alcoholic fixative?
1) It is ideal for small tissue fragments
2) It may be used as both fixative and dehydrating agent
3) It is excellent for glycogen preservation
What are the disadvantages of using alcoholic fixative?
- 1) It lower concentrations w/out the cause of RBC lysis
2) It dissolves fats and lipids
3) Tissue left in alcohol for too long will shrink
What are the types of alcoholic fixative?
1) 100% methyl alcohol
2) Ethyl alcohol
3) Carnoy’s fluid
4) Newcomers
5) Clarke’s solution
6) Methacarn
7) Rossman’s
What are the actions of 100% methyl alcohol?
1) It is excellent for fixing dry and wet smears, blood smears, and bone marrow tissues
2) It fixes and dehydrates at the same time
3) It is used for fixing touch preparations
True or False
The penetration of 100% methyl alcohol is fast
False, because the penetration of 100% methyl alcohol is slow
If the tissue is left for 48 hrs > in 100% methyl alcohol, what will happen to the tissue?
It will overharden
True or False
Ethyl alcohol is usually incorporated w/ other fixatives to produce better result
True
The use of Carnoy’s fluid is recommended for what?
For fixing:
1) Chromosomes
2) Lymph glands
3) Urgent biopsies
Carnoy’s fluid is used for fixing what?
Brain tissues
What is the purpose of using Carnoy’s fluid for fixing brain tissues?
For the dx of rabies
The use of Newcomers is recommended for fixing what?
1) Mucopolysaccharides
2) Nuclear proteins
Clarke’s solution produces good results w/ what?
H&E staining
What is the characteristic of Methacarn?
It causes less shrinkage > Carnoy’s
Rossman’s is good for what?
1) Connective tissue mucins
2) Umbilical cord
Osmium tetroxide is used for what?
1) For museum preparation
2) For neurological tissues
*What is the principle of usage of osmium tetroxide?
Yellow powder that dissolved in H2O to form strong oxidizing solution
What are the advantages of using osmium tetroxide?
1) It fixes fats and lipids
2) It preserves cytoplasmic structures well
3) It fixes myelin and peripheral nerves well
4) It permits ultrathin sectioning in EM
What are the disadvantages of using osmium tetroxide?
1) It is very expensive
2) It is a poor penetrating agent
3) It can irritate the eye upon prolonged exposure
What are the types of osmium tetroxide fixative?
1) Flemming’s solution
2) Flemming’s solution w/out acetic acid
What is the composition of Flemming’s solution?
Chrome + osmium + acetic acid
Due to the composition of Flemming’s solution, what is the characteristic of this fixative?
It is the most common chrome- osmium- acetic acid fixative
The use of Flemming’s solution is recommended for what?
For nuclear preparation
What is the action of Flemming’s solution?
It permanently fixes fat
What is the composition of Flemming’s solution w/out acetic acid?
Only chromic acid and osmic acid
The use of Flemming’s solution w/out acetic acid is recommended for what?
For cytoplasmic structures (particularly the mitochondria)
What is done to trichloroacetic acid?
It is usually incorporated w/ other fixatives
Why is tricholoroacetic acid usually incorporated w/ other fixatives?
To form a compound fixative
What is the action of trichloroacetic acid?
It can fix and decalcify the tissue at the same time
What are the advantages of using trichloroacetic acid?
1) It precipitates proteins
2) It has softening effect on dense fibrous tissues
What is the disadvantage of using trichloroacetic acid?
It is a poor penetrating agent
What are the uses of acetone?
1) It is used at ice cold temp ranging from -5 to 4 DC
2) It is used in fixing brain tissues for the dx of rabies
What are the advantages of using acetone?
1) It is recommended for the study of H2O diffusible enzymes especially phosphatases and lipases
2) It can fix and dehydrate at the same time
What are the disadvantages of using acetone?
1) It produces considerable shrinkage and distortion
2) It poorly demonstrate fat and glycogen and evaporates rapidly
