M4- STAINING OF NERVOUS TISSUE MICROORGANISMS AND PROTEIN AND NUCLEIC ACIDS Flashcards
(182 cards)
1
Q
Protective coats covering CNS
A
Dura mater+Arachnoid+Pia mater
2
Q
Components of gray matter
A
Neurons+Supporting cells
3
Q
Components of white matter
A
Myelinated axons+Oligodendrocytes
4
Q
Cells lining ventricles and central canal
A
Ependymal cells
5
Q
Main excitable cell of CNS
A
Neuron
6
Q
Neuron cell body function
A
Metabolic upkeep with various organelles
7
Q
Neuron nucleus function
A
Storage of genetic code in DNA
8
Q
Neuron dendrites function
A
Receive synaptic inputs from other neurons
9
Q
Neuron axon function
A
Transmit electrical impulses away from soma
10
Q
Neuron cytoskeleton composition
A
Microtubules+Neurofilaments+Microfilaments
11
Q
Neurofibril description
A
Network of fibrils in cytoplasm and processes
12
Q
Myelin sheath elaborated by
A
Oligodendrocytes
13
Q
Major neuroglial cell types
A
Astrocytes+Oligodendrocytes+Microglia
14
Q
Astrocyte functions
A
Support brain+Maintain extracellular ion and neurotransmitter balance+Repair and scarring+Part of blood-brain barrier
15
Q
Astrocyte injury response
A
Produce dense network of processes forming glial scar
16
Q
Major astrocyte cytoskeletal protein
A
Glial fibrillary acidic protein (GFAP)
17
Q
Oligodendrocyte features
A
Small rounded lymphocyte-like nuclei+Wrap axons to form myelin sheath+Smaller than astrocytes
18
Q
Microglia features
A
Smallest neuroglia cells+Macrophages of brain+Dense elongated nuclei+Located near neurons in gray matter
19
Q
Microglia origin
A
Derived from circulating phagocytic cells
20
Q
Microglia function
A
Tissue repair+Form foamy macrophages+Form microglial nodules in viral infections
21
Q
Preferred fixatives for CNS
A
Formol-saline+Buffered formalin+Lillie’s calcium acetate formalin (for myelin histochemistry)
22
Q
Fixation duration
A
At least 24 hours; whole brain ~7 days
23
Q
Common artifacts
A
Perivascular retraction spaces due to shrinkage or fixation
24
Q
Common CNS staining techniques
A
Hematoxylin and eosin+Immunohistochemistry+Silver impregnation+Basic aniline dyes (cresyl violet
25
Note on silver impregnation and basic aniline dyes
Not commonly used for CNS staining; immunohistochemistry preferred
26
Purpose of hematoxylin and eosin stain
CNS tumor demonstration
27
Bodian silver staining target
Neuronal cell bodies and nerve processes (dark brown)
28
Holzer’s crystal violet staining target
Astrocyte fibrous components and gliosis (purple)
29
Nissl stain (cresyl violet) target
Neuronal Nissl substance (ribosomal RNA) staining cytoplasm blue-purple
30
Luxol Fast Blue stain target
Myelin sheaths in white matter
31
Kluver–Barrera stain
Combination of Luxol Fast Blue and Cresyl Violet for myelin and Nissl substance
32
Immunohistochemistry use
Specific molecular target visualization using antibodies
33
General CNS tissue preparation
Perfusion fixation+Embedding in paraffin+Sectioning+Dewaxing and rehydration before staining
34
Blocking step purpose
Reduce nonspecific antibody binding
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Nissl bodies composition
Coarse granules of ribonucleic acid scattered in neuron cytoplasm
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Significance of absence of Nissl bodies
Suggests nerve cell degeneration
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Common clinical use of Nissl staining
Autopsy especially in Alzheimer’s disease
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Cellular structure stained by Nissl stain
Rough endoplasmic reticulum with ribosomes
39
Best dyes for Nissl staining
Basic dyes such as cresyl fast violet
40
Colors in Cresyl Fast Violet (Nissl) stain
Nissl bodies purple-dark blue+Neurons pale purple-blue+Cell nuclei purple-blue
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Definition of central chromatolysis
Absence of Nissl bodies in neurons indicating degeneration
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Stains used for astrocytes
Cajal’s Gold sublimate+PTAH+Holzer technique+Hortega Lithium carbonate+IHC for GFAP and other proteins
43
Cajal’s Gold sublimate method features
Silver nitrate treated tissue+Photographic developers+Frozen sections+Astrocytes bluish black+Nerve cells red
44
Modified PTAH stain use
Detect reactive astrocytes in formalin-fixed paraffin sections
45
Modified Holzer’s method features
Formalin or Bouin’s fixation+Crystal violet stain+Glial fibrils blue+Nuclei pale blue
46
Immunohistochemical markers for oligodendrocytes
Antibodies to galactocerebroside
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Markers for microglial cells
CD68 positive macrophage markers+Faint CD45 positivity
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Composition of myelin sheath
Lipids including cerebrosides
49
Common myelin stains
Baker’s chromatin-acid hematein+Osmium tetroxide+Sudan dyes+Iron hematoxylin+Marchi’s reaction+Luxol fast blue
50
Weigert-Pal technique for myelin
Prolonged chromation with chromium fluoride and copper acetate+Oxidized hematoxylin staining blue-black deposits
51
Kluver & Barrera stain
Combination of Luxol Fast Blue and Cresyl Violet for myelin and Nissl substance
52
Luxol Fast Blue-PAS-Hematoxylin staining results
Myelin blue+Fungi and PAS-positive elements rose to red+Nuclei dark blue+Cytoplasmic nucleoproteins bluish purple+Capillaries red
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Weil’s method features
Ferric ammonium sulfate and oxidized hematoxylin+Myelin black+Background yellow
54
Baker’s chromate acid hematin stain
Calcium dichromate binds choline lipids forming colored complex with hematein+Myelin sheaths mitochondria erythrocytes dark blue
55
Degenerating myelin stain in Swank & Davenport’s Marchi method
Black
56
Background color in Marchi method
Colorless to pale brown
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Technique using silver ammonium hydroxide reduction by formalin
Microwave modification of Bielschowsky’s technique
58
Color of axons in Bielschowsky’s technique
Brown to black
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Color of cytoplasmic neurofibrils in Bielschowsky’s technique
Brown to black
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Color of neurofibrillary tangles and plaques in Bielschowsky’s technique
Dark brown or black
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Color of neuromelanin in Bielschowsky’s technique
Black
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Color of lipofuscin in Bielschowsky’s technique
Brown or black
63
Stain using Protargol-S and copper metal with gold chloride toning
Bodian stain
64
Bodian stain application
Neuritic plaques and neurofibrillary tangles in Alzheimer’s disease
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Silver impregnation with ammoniacal silver solution similar to Bielschowsky
Sevier-Munger technique
66
Color of peripheral neurites in Sevier-Munger technique
Black
67
Color of axons in Sevier-Munger technique
Black
68
Color of myelin sheath in Sevier-Munger technique
Light brown
69
Color of neuritic plaques and tangles in Sevier-Munger technique
Black
70
Color of argentaffin granules in Sevier-Munger technique
Black
71
Impregnation with potassium dichromate and silver nitrate
Golgi’s silver staining technique
72
Appearance of nerve cells and neuroglia in Golgi’s technique
Black deposits on colorless background
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Modification involving vacuum for reagent penetration
Modified Golgi method
74
Intermediate filament protein expressed by astrocytes and ependymal cells
Glial fibrillary acidic protein (GFAP)
75
Marker for astrocytomas and ependymomas by IHC
GFAP antibody
76
GFAP antibody staining pattern
Cytoplasmic in astrocytes and ependymal cells
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Neuronal marker staining nuclei primarily
NeuN antibody
78
Major constituent of myelin sheath stained by MBP antibody
Myelin basic protein
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Components of peripheral nervous system
Ganglion cells+Nerve fibers plexuses nerves nerve roots
80
Fixative for peripheral nerve electron microscopy
2.5–3% glutaraldehyde
81
Fixative for peripheral nerve immunohistochemistry
1–4% paraformaldehyde
82
Post-fixation for peripheral nerves
1% osmium tetroxide for 2 hours at room temperature
83
Embedding and sectioning method for peripheral nerves
Epoxy resin embedding+1-μm methylene blue stained sections
84
Appearance of myelin sheaths in H&E paraffin sections
Empty zones surrounding eosinophilic axons encircled by Schwann cell cytoplasm and endoneurium
85
Myelin staining techniques in peripheral nerves
Dye metal complexes of iron+Water insoluble anionic dyes
86
Myelin color with methylene blue-azure II-basic fuchsin stain
Blue
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Collagen color with methylene blue-azure II-basic fuchsin stain
Pink or red
88
Elastin color with methylene blue-azure II-basic fuchsin stain
Red
89
Function of osmium tetroxide
Fixative producing insoluble black deposits at sites of unsaturation+Primary or post-fixation after formaldehyde or glutaraldehyde
90
Type of dye used in negative staining for microorganisms
Acidic anionic dye such as nigrosin or India ink
91
Simple staining dye for bacteria
Methylene blue
92
Gram positive bacterial cell wall component
Peptidoglycan
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Gram negative bacterial cell wall component
Lipopolysaccharide
94
Key reagents in Gram staining
Crystal violet+Iodine+Acetone or ethyl alcohol+Safranin or carbol fuchsin
95
Color of Gram positive bacteria
Blue or purple
96
Color of Gram negative bacteria
Red or pink
97
Fixative used in Modified Brown-Brenn stain
Formalin
98
Color of Gram positive organisms in Modified Brown-Brenn stain
Blue
99
Color of Gram negative organisms in Modified Brown-Brenn stain
Red
100
Color of nuclei in Modified Brown-Brenn stain
Red
101
Color of other tissue elements in Modified Brown-Brenn stain
Yellow
102
Color of Gram positive organisms in Gram-Twort stain
Blue-black
103
Color of Gram negative organisms in Gram-Twort stain
Pink-red
104
Color of nuclei in Gram-Twort stain
Red
105
Color of RBCs and cytoplasmic structures in Gram-Twort stain
Green
106
Color of elastic fibers in Gram-Twort stain
Black
107
Acid-fast bacteria characteristic
Resist decolorization with acid alcohol due to mycolic acid
108
Fixative for Ziehl-Neelsen stain
Formalin or other than Carnoy’s
109
Color of mycobacteria in Ziehl-Neelsen stain
Red
110
Background color in Ziehl-Neelsen stain
Blue
111
Fixative for Fite stain
10% neutral buffered formalin
112
Color of acid-fast bacteria in Fite stain
Red
113
Background color in Fite stain
Light blue
114
Fluorescent stain for mycobacteria
Auramine O-Rhodamine B
115
Color of acid-fast organisms in Auramine-Rhodamine stain
Reddish-yellow fluorescence
116
Background color in Auramine-Rhodamine stain
Black
117
Stain for Helicobacter pylori
Toluidine blue (pH 6.8)
118
Color of Helicobacter in toluidine blue stain
Dark blue
119
Color of nuclei in toluidine blue stain
Variable blue shades
120
Color of Helicobacter and nuclei in cresyl violet acetate stain
Blue violet
121
Background color in cresyl violet acetate stain
Shades of blue-violet
122
Legionella pneumophila Gram stain characteristic
Gram-negative faintly staining coccobacillus
123
Legionella pneumophila morphology
Small pleomorphic coccobacillus 0.5–1 μm wide and 2–50 μm long
124
Preferred culture medium for Legionella
Buffered charcoal yeast extract (BCYE) agar
125
Legionella pneumophila growth on blood agar
Poor or no growth
126
Legionella pneumophila colony morphology
Gray-white with textured cut-glass or opal-like appearance
127
Legionella pneumophila special staining methods
Dieterle silver stain+Warthin-Starry silver stain+Steiner stain
128
Color of Legionella pneumophila in Dieterle stain
Brown to black
129
Background color in Dieterle stain
Pale yellow or tan
130
Color of Legionella pneumophila in Warthin-Starry stain
Black
131
Background color in Warthin-Starry stain
Golden yellow
132
Steiner and Steiner microwave procedure fixative
10% neutral buffered formalin (avoid mercurial and chromate fixatives)
133
Color of Legionella pneumophila in Steiner and Steiner stain
Dark brown to black
134
Other bacteria stained by Steiner and Steiner
Helicobacter pylori and other non-filamentous bacteria dark brown to black
135
Legionella pneumophila acid-fast stain positivity
L. micdadei is modified acid-fast positive (unique among Legionella species)
136
Microscopic features of Legionella infection
Leukocytoclastic inflammatory infiltrate of macrophages and neutrophils+Diffuse alveolar damage+Septic vasculitis+Focal septal disruption
137
Common differential diagnoses for pneumonia
Klebsiella pneumoniae (Gram-negative rod)+Streptococcus pneumoniae (Gram-positive cocci)+Haemophilus influenzae (Gram-negative coccobacillus)+Respiratory viruses+Chlamydia pneumoniae+Mycoplasma pneumoniae+Pseudomonas aeruginosa
138
Recommended antibiotics for Legionella
Macrolides (azithromycin
139
Nucleic acids composition
Alternate sugar and phosphate groups with nitrogenous bases
140
DNA sugar and bases
Deoxyribose sugar with purines (adenine guanine) and pyrimidines (thymine cytosine)
141
RNA sugar and bases
Ribose sugar with purines (adenine guanine) and pyrimidines (uracil cytosine)
142
Acidic dyes stain
Acidophilic substances (cationic at physiologic pH)
143
Basic dyes stain
Basophilic substances (anionic at physiologic pH)
144
Hematoxylin and eosin nuclear stain
Aluminum ion-hematin complex (hemalum) stains nuclei purple
145
Eosin stains
Eosinophilic structures in red pink and orange shades
146
Histochemical protein identification targets
Amino groups (lysine)+Phenyl groups (tyrosine)+Disulfides and sulfhydryl linkages (cystine cysteine)+Indole groups (tryptophan tryptamine)+Guanidyl groups (arginine)
147
Common fixative for amino acid histochemistry
Neutral buffered formalin saline
148
Embedding medium for amino acid histochemistry
Glycol methacrylate plastic
149
Acid dyes used for ribboned epon sections
Aniline Blue Black+Coomassie Brilliant Blue R-250
150
Alkaline fast-green method stains
Basic proteins such as protamines and histones
151
Peracetic acid-alcian blue stain detects
Cystine and cysteine oxidized to cysteic acid stained blue-green
152
Alcian blue-PAS staining differentiates
Acidic mucins (alcian blue) and neutral mucins (PAS positive)
153
Feulgen stain target
Nuclear DNA via Schiff reagent reaction with aldehydes
154
Feulgen stain specificity
DNA-specific; RNA not hydrolyzed by acid
155
Feulgen stain fixation
Zenkers+Carnoy’s+Formalin+Kelly’s+Regaud’s+Flemming’s+Susa (except Bouin’s and Brasil’s)
156
Feulgen stain results
DNA red-purple+Cytoplasm green
157
Methyl green-pyronin stain
DNA green+RNA pinkish red+Granules dark rose-red+Plasma cell cytoplasm purple
158
Methyl green-pyronin fixation
10% formalin+Absolute alcohol+Zenker’s+Carnoy’s
159
Fluorescein fluorescence
Apple green emission with blue light excitation
160
Rhodamine fluorescence
Orange-red emission with green light excitation
161
Acridine orange fluorescence
DNA yellow-green+RNA brick to orange-red+Used for cervical smear screening
162
Acriflavine use
Alternative Schiff reagent in Feulgen technique+DNA fluorescent yellow stain
163
Electron microscopy stain
Phosphotungstic acid aqueous solution for aldehyde-fixed tissue
164
Polyacrylamide gel electrophoresis (PAGE)
Separates proteins or nucleic acids by electrophoretic mobility
165
Common PAGE protein stains
Coomassie Brilliant Blue R-250+Ethidium bromide+Silver stain
166
Basis of PAGE separation
Movement of proteins from cathode to anode based on charge and size
167
Most sensitive technique for identifying DNA
In situ hybridization (ISH)
168
Probe types used in ISH
Labeled complementary DNA RNA or modified nucleic acid strands
169
Purpose of ISH
Localize specific DNA RNA nucleic acid sequences or gene expression within cells
170
Pre-treatment step in ISH
Unmask target nucleic acids for probe hybridization
171
Two main detection methods in ISH
Fluorescence in situ hybridization (FISH)+Chromogenic in situ hybridization (CISH)
172
Visualization method in FISH
Fluorescence microscopy detecting fluorophore signals
173
Visualization method in CISH
Bright-field microscopy detecting chromogen (brown stain)
174
HER2 status detection methods
Immunohistochemistry (IHC) for protein expression+FISH for gene amplification confirmation
175
HER2 gene amplification visualization in FISH
Fluorescent probes light up and signals counted per cell
176
Clinical importance of HER2 detection
HER2-positive patients can receive targeted therapy like trastuzumab (Herceptin)
177
PCR amplification steps
Denaturation at 95°C+Primer annealing at 54°C+Extension at 72°C
178
RT-PCR unique feature
RNA converted to complementary DNA by reverse transcriptase before amplification
179
In situ PCR advantage
Detects minute quantities of nucleic acid sequences within tissue sections localized in cells
180
Difference between FISH and CISH
FISH uses fluorescent probes and fluorescence microscopy+CISH uses chromogenic probes and bright-field microscopy
181
Advantages of CISH over FISH
Lower cost+Easier use+Stable reagents allowing sample storage+Simultaneous tissue morphology assessment
182
FISH advantage
High sensitivity and resolution+Multiplexing capability for multiple targets
