Fixation Flashcards
1
Q
This process is
- done at 0-4 degrees C
- fixatives containing glutaraldehyde followed by secondary fixative, osmium tetroxide
A
ELECTRON MICROSCOPY
2
Q
- maximal preservation of enzyme activity to where it
originates, while also preserving structural integrity - overnight fixation using 4% formaldehyde or formal-saline
A
ENZYME HISTOCHEMISTRY
3
Q
- fixed with acetone or formaldehyde for frozen sections
A
ENZYME HISTOCHEMISTRY
4
Q
- uses the principle of Ag-Ab reaction
- epitopes (sites where the Ab will attach to) of antigens must be preserved for antibodies to bind on
antigens to be detected - if prolonged fixation was performed, antigenicity of Ag will be lost because of the loss of epitope
A
IMMUNOFLUORESCENCE AND
IMMUNOHISTOCHEMISTRY
5
Q
- antigen retrieval can be done to bring back antigenicity using Heat-induced epitope retrieval
(HIFR)
A
IMMUNOFLUORESCENCE AND
IMMUNOHISTOCHEMISTRY
6
Q
- employing the advantage of heat to fi tissue, microwaves aid in the fixation process by either allowing heat to permeate and fix the tissue itself (microwave fixation) or heat using microwave assists the permeation of a chemical fixative (microwave assisted fixation)
A
MICROWAVE FIXATION
7
Q
- use of chemical reagents to preserve tissues
- chemical fixatives have their own different actions
o cross-links with proteins, providing additional rigidity to the cells
o denatures proteins
A
CHEMICAL FIXATION
8
Q
- Formaldehyde (product of the production of methanol) is sold as _______ w/v formaldehyde
solution
A
FORMALDEHYDE AND FORMALIN in 10% concentration
35-40%
9
Q
- white powder; formaldehyde polymers
- allows paraffin embedding and sectioning, and IHC
- effects are reversible by excess water
- avoids pigmentation produced by formalin
A
PARAFORMALDEHYDE
10
Q
- standard fixative for EM
- less dangerous than para/formaldehyde;
pleasing smell; less irritating
A
GLUTARALDEHYDE
11
Q
GLUTARALDEHYDE
- 2%-
- 2.5% -
- 4% -
A
GLUTARALDEHYDE
- 2%- EM
- 2.5% - small tissue biopsies and needle biopsies
- 4% - for larger tissues less than 4mm thick
12
Q
- 4% paraformaldehyde – 1% glutaraldehyde in 0.1M
Phosphate Buffer
A
KARNOVSKY’S FIXATIVE
13
Q
- suitable to use when tissues are to be embedded in
resin - suitable fixative for EM because of the presence of
glutaraldehyde
A
KARNOVSKY’S FIXATIVE
14
Q
- denatures CHONS
- not routinely used for tissues as it can produce severe distortion on tissue samples, thus only used in
slide samples which has a thickness around just a cell or 2 layers of cells - rarely used alone for fixing locks, unless studying nucleic acid
A
ALCOHOLIC FIXATIVES
15
Q
- good for cytologic smear fixation
- used as 70& to 100% concentration
- mixed with acetic acid, water, chloroforms to reduce distortions
A
ALCOHOLIC FIXATIVES
16
Q
– general fixative for Peripheral Blood Smear (PBS), dry and wet smears, and Bone Marrow
Aspirates (BMA) smears
A
Methanol
17
Q
– touch preps
A
95% Isopropanol
18
Q
– used as a fixative and a
dehydrating agents
A
Ethanol (70-100%)
19
Q
– preserves Nissl bodies, cytoplasmic granules,
nucleoproteins, nucleic acids; for fixing brain tissues for rabies dx
A
Carnoy’s Fluid (absolute alcohol in Chloroform)
20
Q
– fixation or post- fixation of large
fatty specimens
A
Alcoholic Formalin
21
Q
– can be used on frozen sections
A
Clarke’s solution
22
Q
– sometimes used for fixing
tissues for diagnostic cryostat
A
Formol – acetic Alcohol
23
Q
– enhances immunoperoxidase
studies; for glycogen and micro-incineration
A
Gendre’s Fluid
24
Q
– for fixing mucopolysaccharides
and nuclear proteins
A
Newcomer’s Fluid
25
- most common metallic fixative
- used as a 5-7% aqueous solution
- works by additive and coagulative means
* it can incorporate itself in cells and precipitate CHONS, thus denaturing them in process
MERCURIC CHLORIDE
26
– recommended for congested
specimens, and for trichrome staining and Phophotungstic acid Hematoxylin (PTAH) staining
Zenker’s solution
27
- excellent fixative
for bone marrow, liver (to demonstrate extramedullary hematopoiesis), spleen, pituitary
glands, and cardiac muscle
Zenker-Formol (Helly’s) Solution
28
– for fixing hematopoietic BM
biopsies, and lymphoid tissues
Lillie’s B-5 fixative
29
– for tumor biopsies of the skin;
permits easier sectioning of large blocks of fibrous connective tissue
Heidenhain’s Susa
30
- causes complete denaturation of CHONS
- traditionally used in EM (fixative and stain); now, it act as secondary fixative
OSMIUM TETROXIDE; OSMIC ACID
31
- good fixative and stain for lipids
- preserves cytoplasmic structures
- penetrates at a slower rate vs. metallic fixatives
OSMIUM TETROXIDE; OSMIC ACID
32
- may give off a black stain
- extremely volatile; prolonged exposure to vapors may cause conjunctivitis
OSMIUM TETROXIDE; OSMIC ACID
33
OSMIUM-BASED FIXATIVES
1. _________________ – most common; permanent fixative for fat
2. _________________________________ – recommended for demonstrating mitochondria
OSMIUM-BASED FIXATIVES
1. Flemming’s Fluid – most common; permanent fixative for fat
2. Flemming’s w/out acetic acid – recommended for demonstrating mitochondria
34
– good in preserving
mitochondria at pH 4.5-5.2
Chromic acid (1-2%)
35
– recommended for demonstrating
mitochondria, mitotic figures, golgi bodies, RBC and colloid- containing tissues
Regaud’s fluid
36
– recommended for studying early
degenerative processes and tissue necrosis; also advisable for demonstrating rickettsiae and other
bacteria
Orth’s Fluid
37
- performs almost as well as mercuric fixative; doesn’t harden as much
- reacts with histones and basic proteins; precipitates al, proteins
PICRATE FIXATIVES
38
- good fixative for connective tissues, preserves glycogen well, extracts lipids
- May impart a yellow pigments
o removed using lithium carbonate or wash in
alcohol in changes of 50-70%
PICRATE FIXATIVES
39
- recommended for fixing embryos and pituitary biopsies, also glycogen
Bouin's Fluid
40
- recommended for gastrointestinal tract specimen and endocrine tissues; has some decalcifying properties
Hollande's Solution
41
- highly recommended for glycogen
and carbohydrate preservation; produces minimal distortion of micro-anatomical structures.
Gendre's Fluid
42
- better and less messy than Bouin's; excellent for preserving
glycogen.
Brasil's Alcoholic Picroformol Fixative
43
- Anhydrous
- Freezes at 16°C
GLACIAL ACETIC ACID
44
- Enables cell swelling Precipitates and fixes nucleoproteins
- Compounded with other fixatives to counteract the swelling effect
GLACIAL ACETIC ACID
45
- metallic fixative because of the presence of lead
- Recommended for fixing acid mucopolysaccharides, mucin
- May produce lead carbonate upon standing.
o white ppt at the bottom; filtered out
LEAD FIXATIVES
46
- Fixative and decalcifying agent
TRICHLOROACETIC ACID (TCA)
47
- Precipitates proteins and nucleic acids
- Used only on small pieces; penetrates poorly
TRICHLOROACETIC ACID (TCA)
48
- non-alcoholic
- Not recommended as a morphological fixative
because it is known to distort tissue
ACETONE
49
- Used in fixing cryostat sections and for tissues
requiring enzyme preservation.
- Used to fix tissues at cold temperatures (0-4°C)
ACETONE
50
- Recommended for fixing brain tissues to diagnose rabies
- Preserves glycogen poorly; dissolves fats.
ACETONE
51
- Not a fixative
- Stable medium for transporting fresh, unfixed tissues that will undergo frozen sectioning or IF studies.
- Specimens can be kept in the solution for _ days until it can be delivered to a reference laboratory.
- Not suitable for keeping tissues for ____.
- Tissues in such solution must be kept at an ambient temperature of ______
MICHEL'S SOLUTION
- Not a fixative
- Stable medium for transporting fresh, unfixed tissues that will undergo frozen sectioning or IF studies.
- Specimens can be kept in the solution for 5 days until
it can be delivered to a reference laboratory.
- Not suitable for keeping tissues for FISH.
- Tissues in such solution must be kept at an ambient temperature of 4-22°C
