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Stack #170785

Nucleic Acid Stains

Two types DNA AND RNA Nuclei Acid
Has 5 carbon sugars called Deoxyribose, Found in nucleus, its major constituent is nuclear chromatin DNA
Has 5 carbon sugars called Ribose, found in the nucleolus and ribosomes RNA
The sugars in DNA and RNA differ by a single hydroxyl group (OH-). the difference allows for selective demonstration of DNA and RNA by special staining technique DNA AND RNA
process by which a chemical compound decomposes by reacting with water Hydrolysis
demonstrates DNA, hydrolysis of DNA by hydrochloric acid Feulgen stain
With Feulgen DNA is hydrolyzed by 1N HCl which removes the purine (adenine and guanine)base but leaves the sugars and phosphates of DNA intact, this generates an aldehyde which can be demonstrated with Schiff's reagent Feulgen stain
time of hydrolysis must be carefully controled, after removal of purine bases and formation of aldehyde groups, there is a progressive removal of histones and apurinic acids, second reaction leads to false negative feulgen reaction Feulgen stain
DNA appreas Magenta/reddish purple Feulgen stain
cytoplasm stains only if a counter stain is applied (light green) Feulgen stain
Dont use BOUINS to fix tissues it hydrolyzes nuclei excessively, dont overcounterstain with light green Feulgen stain special consideration
masked feulgen overcounterstained with light green
NONE required all nuclei will stain Feulgen control tissue
purpose to differentiate between RNA and DNA Methyl green pyronin Y
differential staining is due to differing degrees of polymerization between DNA and RNA Methyl green pyronin Y
DNA high Polymer, RNA lower Polymer Methyl green pyronin Y
used to identify plasma cells and immunoblasts in tissues Methyl green pyronin Y
DNA binds w methyl green (green/blue color), RNA binds with Pyronin (red color), background pale pink to colorless, mucins, epilthelium and cartilage appear pink to red Methyl green pyronin Y
Carnoyed fixed best, 10%NBF works, B-5, Helly or Zenker, Methyl green pyronin Y fixatives
Control tissue, a section containing numerous plasma cells Methyl green pyronin Y Controls
acids and fixatives may depolymerize DNA resulting in loss of methyl green staining, While the RNA bind to pyronin. resulting in loss of differential staining. Methyl green pyronin Y special consideration
pyronin is not specific for RNA. cartilage, osteoid, keratin, eosinphil granules and mast cell granules will also stain, DNA will stain but pyronin cant compete with Methyl green for DNA Pyronin
process where a dye forms other dyes spontaneously Polychroming
A compound dye or dye mix that contains components of different colors Polychromatic stain example
Most common example of Romanowsky stain May-Grunwald Giemsa
stains for hematopoietic tissues another group of nuclear and cytoplasmic special staining technique May-Grunwald Giemsa
where red and white blood cells are formed hematopoietic tissues
stains used to identify different cell lineages found in hematopoietic tissues, like spleen,bone marrow and blood May-Grunwald Giemsa
basophilic or basic dye, methylene blue is combined w/eosinphilic or acid dye, to create "neutral dyes" that demonstrate a wide variety of colors when used to stain hematopoietic cell nuclei and platelets. May-Grunwald Giemsa principle (chemistry)
differentiate cells present in hematopoietic tissue. demonstrates presence of microorganisms. hitological dx for malaria, spirochete and protozoan blood parasites May-Grunwald Giemsa
Nuclei stains blue, cytoplasm leukocytes pink, gray or blue, bacteria stains blue May-Grunwald Giemsa
Zenker or B-5 preferred, 10%NBF ok to use May-Grunwald Giemsa fixative
working solutions not stable, prepare solutions before use and then discard. May-Grunwald Giemsa
if staining is poor, the effect of the pH adjustment should be investigated. pH should be between 6.4-6.9 May-Grunwald Giemsa
Control spleen May-Grunwald Giemsa
made of 1-2% carbohydrate (chondroitin,heparin,and dermatan)and acid mucopolysaccharides Amyloid
disease charactarized by amorphorous, eosinphilic, extracellular deposit, that slowly replace cellular elements of vital organs and causes progressive loss of function and eventually death amyloidosis
varies between patients and between organs in the same patient composition of amyloid
occurs spontaneous in the absence of any predisposing disease primary amyloidosis
organs affected are muscle, heart, skin, tongue primary amyloidosis
associated with a predisposing disease, chronic inflamatory disease rheumatoid arthritis, ankylosing spondylitis, infections such as tuberculosis and osteomyelitis secondary amyloidosis
this type of amyloid most frequently deposited in kidneys, liver, spleen and adrenal gland secondary amyloidosis
associated with disease of the immunological system, resembles primary amyloid Myeloma-associated amyloid
found w/many tumors APUD Tumor associated amyloid
group of cells of embryonic origin that secrete most of the bodys hormones APUD celss
compromise both specialized neurons and endocrine cells.synthesize structually related polypeptides and biogenic amines APUD cells
amine precursor uptake and decarboxylation APUD
insulin, ACTH, glucagon,antidiuretic hormes peptide hormone examples
dopamine, norepinephrine, serotonin and histamine amine hormone examples
oncology tumor that produces small peptide hormones of the APUD system Apudoma
small-oat cell carcinoma of lung, carcinoids of lung, thymus, GI tract and prostate, medullary CA of thyroid, pancreatic islet cell tumor, malignant melanoma, Ganglioneuroma Apudoma Examples
linked to uptake of precursor amino acid and its decarboxyaltion in the cell to produce an amine polypeptide production
immunoglobulin derived and includes primary amyloid and myeoloma associated amyloid AL amyloid
Unknown origin and includes secondary amyloid AA amyloid
heredofamilial amyloid (familial mediterranean fever) AF amyloid
polypeptide-hormone derived,associated with tumors of the APUD system APUD
demonstrates amyloid in tissues, most specific technique to demonstrate amyloid, Alkaline congo red
pretreatment with alkali aids in the release of native internal hydrogen bonds between adjacent protein chains creating mores sites for the dye to bind alkaline congo red
amyloid stains deep pink to red with light microscope congo red
amyloid stains bright green apple with polorazing microscope Congo red
elastic tissues stain pale pink congo red
nuclei stains blue congo red
cut section at 8-10 microns congo red
any tissue w/amyloid, staining intensity decreased w/time, dont cut too many control slides ahead of time congo red special considerations
need polorazing scope to view green apple birefringence congo red
rapid screening method to demonstrate amyloid crystal violet
not as specific as congo red crystal violet
addition of HCl in the stain prevents the cytoplasmic component from overstatining crystal violet
amyloid stains violet, other tissue elements stain blue crystal violet
cut sections 10-12 microns crystal violet
any tissue with amyloid crystal violet control
cover slip w/apathy mounting media or aqueous based mounting media or air dry slide completely then dip in xylene and use resinous media crystal violet
purpose demonstrate presence of amyloid. not as good as congo red w/polarizing light, background nuclear fluoresece is quench w/ aluminum hematoxylin. no differentiation is required Thioflavin T fluorescent Method
Amyloid fluoresces yellow/yellow green Thioflavin T fluorescent Method
Sections at 6-10 microns Thioflavin T fluorescent Method
Thioflavin T at acidic concentration increases the selectivity of the dye for amyloid, pH 1.4 is recommended Thioflavin T fluorescent Method special considerations
lipid granules, mast cells and juxtaglomerular granules may give yellow fluoresence Thioflavin T fluorescent Method
pH of staining solution critical. More acid levels give more selective chromatin staining and less cytobasophilia, less acid pH levels give denser nuclei and increased cytoplasmic basophilia May-Grunwald Giemsa
Created by: nperez
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