Techniques for Studying Viruses Flashcards
(88 cards)
1
Q
direct viral test for HIV
A
light microscopy
electron microscopy
antigen detection
molecular testing
2
Q
indirect viral test for HIV
A
viral isolation (tissue culture, embryonated eggs, animal inoculation)
3
Q
Clinical specimen is examined
directly for the presence of virus
particles, virus antigen or viral
nucleic acids
what kind of isolation
A
direct examination
4
Q
direct examination iis used when clincal specimen is examined
directly for the presence of (3)
A
virus
particles, virus antigen or viral
nucleic acids
5
Q
- Histological appearance of cytopathic effect
- Viral antigens in infected cell cultures are
demonstrated by immuno-peroxidase staining
what kind of direct examination?
A
light microscopy
6
Q
- Viral antigens in infected cell cultures are
demonstrated by what staining in light microscopy
A
immuno-peroxidase staining
7
Q
direct microscopy is used to observe what
A
cytopathic effect of viruses
8
Q
interpretation when 1+ quantitation of monolayer
A
1 to 25 percent of monolayer exhibit CPE
9
Q
Most commonly used method for
direct detection of virus in clinical
specimens
A
electron microscopy
10
Q
electron distinctive virus or viral structures using what tsamples
A
biopsy
clinical specimens
11
Q
direct examination of antigen detection include (3)
A
immunofluorescence
enzyme immuno-assay
latex agglutination
12
Q
- Involves the use of a labeled antiviral antibody
(fluorescein isothiocyanate) which is layered over a
specimen suspected of containing a homologous
virus
A
immunofluorescence
13
Q
immunofluorescence involves the use of a labeled antiviral antibody
called ) which is layered over a
specimen suspected of containing a homologous
virus
A
fluorescein isothiocynate
14
Q
immunoflourescent can be
A
direct
indirect
15
Q
direct examination immunoflourescence intensity
1+
A
faint yet unequivocal apple green fluorescence
16
Q
direct examination immunoflourescence
2+
A
apple green fluorescence
17
Q
direct examination immunoflourescence
3+
A
bright apple green fluorescence
18
Q
direct examination immunoflourescence
4+
A
brilliant apple green fluorescence
19
Q
enzyme is covalently attached to an antigen or
antibody molecule, creating an immunological
tool with high specificity and high sensitivity.
A
ELISA (enzyme-linked immunosorbent assay)
20
Q
ELISA is where the enzyme is covalently attached to an __or
___molecule, creating an immunological
tool with high specificity and high sensitivity.
A
antigen
antibody
21
Q
enzymes typically bound to antigen or antibody
include
A
peroxidase
alkaline phosphatase
B-galactosidase
22
Q
- enzymes typically bound to antigen or antibody
include peroxidase, alkaline phosphatase, and βgalactosidase, all of which interact with enzymespecific substrates to form ___reaction
products that can be detected in very low
amounts.
A
colored
23
Q
if the antigen/antibody is present, then a complex is formed then bound to a color producing __
A
substrate
24
Q
Uses polystyrene microplates (e.g., 96-well plates) as the solid phase of ELISA
A
solid-phase ELISA
25
ELISA that Uses microporous membranes (e.g., nitrocellulose or nylon) as the solid phase
membrane ELISA
26
ELISA that remains the gold standard for quantitative analysis,
solid phase ELISA
27
ELISSA that is advantageous for point-of-care testing or specialized workflows requiring enhanced surface interactions
membrane elisa
28
* Detects - pathogen antigen
* Uses - enzyme-labeled pathogenspecific antibody (supplied)
what type
direct
29
detects - pathogen-specific antibodies
* Uses - enzyme-labeled antibody
directed (supplied) to pathogenspecific antibodies in patient samples
what type
indirect
30
Rapid tests in HIV for antibody is an __test
indirect
31
1.HIV p24 antigens manufactured
and attached to the bottom of a
plastic testing dish.
2. Add blood sample of patient; if
with Ab (Y-shaped), attaches to
p24.
3.Add second marker antibody.
4.Add dye to react to second
marker antibody (changes color.
what test
HIV antibody (indirect/serology)
32
is a biochemical technique used to detect and quantify specific antigens in a sample by exploiting the highly specific binding between an antigen and an antibody, combined with an enzyme-based signal amplification system
antigen enzyme immunoassay
33
1. Antibodies for p24 are
secured to the bottom of a
Petri dish.
* 2. Add blood sample of patient; if HIV pos, p24 in
blood attaches.
* 3. Another Ab for p24 is
attached.
* 4. Add second marker
antibody.
* 5. Add dye to react to
second marker antibody
(changes color)
what kind of HIV detection
HIV antigen (direct/serology)
34
has * Synthetic manufactured proteins mixed with patient’s
blood.
western blood
35
* Ab present in blood combines with their own antigen.
direct or indirect test for western blot?
indirect
36
* Proteins move down the gel, the heavier, the ___.
slower
37
western blot is * __ if at least three lines show.
positive
38
Observed when a sample containing the
specific antigen (or antibody) is mixed with an
antibody (or antigen) which is coated on the
surface of latex particles
antigen latex agglutination
39
is antigen latex aggutination direct or indirect?
direct
40
Determine the presence or absence of a
virus, * Ability to detect multiple viruses
simultaneously
viral gentionome detec
41
* Better sensitivity than cell culture and
DFA
* detection of viruses non-culturable by
cell culture (e.g., norovirus, hepatitis
viruses)
viral genome detection
42
non culturable by cell culture
example of viruses
norovirus
hepatitis viruses
43
potentially characterize the virus
genetically (genotype)
what examination
viral genome detection
44
viral genome detection is direct/indirect
direct
45
example of viral genome detection (5)
DNA probes
DNA probe hybridization
PCR
RT-PCR
Southern blot
northen blot
46
detects DNA sequences and requires DNA fragmentation by restriction enzymes before gel electrophoresis.
southern blot
47
detects RNA sequences directly, using denaturing gels to maintain RNA in a linear form for size separation, without the need for enzymatic digestion
northern blot
48
* Highly sensitive and specific tools used
for detection of virus in clinical
specimens (absence of replication)
dna probes
49
* DNA in clinical specimens are first
separated and then allowed to
hybridize with a labeled singlestranded DNA probe
dna probes
50
* Hybridized labeled probe can be
detected by ___or __
method depending on the label used
enzymatic
fluorescence
51
DNA probe is used for
CMV
papillomavirus
Epstein-Barr virus
52
In this method, the viral genome or restriction endonuclease cleavage
fragments of the genome are blotted onto nitrocellulose papers and then
detected on the paper by their hybridization to DNA probes.
southern blot analysis
53
* In this method, electrophoretically separated viral RNA are transformed onto
a nitrocellulose paper and then detected by their hybridization to DNA
probes.
nortern blot analysis
54
* It is a DNA amplification system that allows a millionfold amplification of a
target sequence of nucleic acids, at least 100,000-folds in a few hours
PCR
55
* Useful for diagnosis of viral infections caused by HIV-1, HIV-2, CMV, human
papilloma virus, HSV, hepatitis B, C, and D viruses, morbillivirus, echovirus,
rhinovirus, measles virus, rotavirus, etc.
PCR
56
indirect examination include
viral isolation
serology
57
viral isolation include
animal inoculation
embryonated eggs
tissue culture
58
limitation is that upkeep is expensive, variation between individuals, even if inbred means large numbers are needed
ethical considerations
limitations of culture system
animal culture system
59
limitation unnatural since cultures are no longer subject to homeostatic reponses
organ culture
60
three types of cell culture
primary cells
cell lines
permanent cell lines
61
derived from an organ or tissue, remain undifferentiated but survive for only a few passages
primary cells
62
are dedifferentiated but diploid and survive a larger number (about 50) passages before they die
cell lines
63
are dediffirentiated but immortal
continuous cell lines
64
* Experimental animals are used for cultivation of viruses
animal inoculation
65
animal inoculation d
direct/indirect?
direct
66
* Play an essential role in study of pathogenesis of viral infections and that of viral oncogenesis
animal inoculation
67
animals used for inoculation
mouse, rabbits, hamsters, newborn or suckling rodents
68
s a laboratory technique where fertilized eggs containing developing embryos and their supporting membranes are used as host systems to isolate and propagate viruses.
viral isolation using embryonated eggs
69
* Initially used for the growth of viruses
embryonated eggs
70
first used fertilized chicken eggs (embryonated eggs) to cultivate viruses.
Ernest Goodpasture
71
expanded on Goodpasture’s work by identifying and utilizing different specific sites within the embryonated egg for virus cultivation.
sir Marcfarlane Burnet
72
Used mainly for cultivation of certain arboviruses like Japanese encephalitis virus, Saint Louis encephalitis virus, and West Nile virus, as well as for growth of some bacteria like chlamydia and rickettsia
what site of inoculation in embryonated egg
yolk sac
73
what site of inoculation in embryonated egg
Primarily used for the primary isolation of viruses such as influenza virus. The virus replicates in the amniotic fluid and embryo tissues accessible here
amniotic activity
74
what site of inoculation in embryonated egg
Commonly used for serial passages and production of large quantities of viruses like influenza, yellow fever (17D strain), and rabies virus. The allantoic cavity is a fluid-filled sac that supports high viral yields, making it valuable for vaccine production
allantoic cavity
75
what site of inoculation in embryonated egg
Some viruses, especially poxviruses (e.g., vaccinia, variola), produce visible lesions called pocks on the CAM, which can be used for virus identification and quantification
chorioallantoic membrane (CAM)
76
Demonstration of virus in appropriate clinical
specimens by culture establishes diagnosis of viral
diseases
tissue culture
77
Gold standard for establishing viral etiology of a
disease
tissue culture
78
tissue culture can be (3)
organ culture
explant culture
cell culture
79
* Routinely used for growing viruses
✓Suitable host cells
✓Cell culture media
✓Techniques in cell culture maintenance
tissue culture
80
* Monkey kidney cell culture
* Human embryonic kidney cell culture
* Chick embryo cell culture
primary cell cutlure
81
* Derived from cancerous tissue
* Capable of continuous serial
cultivation indefinitely without
senescing
* Usually derived from diploid cell
lines or from malignant tissues and
have altered and irregular number
of chromosomes
continuous cell lines
82
* Human carcinoma of cervix cell line (HeLa)
* Human epithelioma of larynx cell line (HEp-2)
* human alveolar adenocarcinoma (A549)
* Human carcinoma of nasopharynx cell line
(KB)
* Human synovial carcinoma cell line (McCoy)
* Sternal marrow cell line (Detroit-6)
* Vervet monkey cell line
* Baby hamster kidney cell line (Vero)/(ELVIS
cells)
what tissue culture
continuous cell lines
83
Less frequently isolated viruses by cell culture
varicella-zoster
measles
rubella
rhinoviruses
coxsackie A viruses
84
Based on detection of specific viral
antibodies in serum of the infected
human host
indirect examination serology
85
indirect examination using serology can be thru
* Hemagglutination
* Inhibition (HI) test
* Neutralization test (NT)
* Indirect fluorescent
* Antibody (IFA) test
* ELISA
* RIA
* Latex agglutination test (LAT)
* Western blot
86
Diagnosis of infections with nonculturable
agents, such as hepatitis viruses
indirect examination serology
87
Determination of immune status in regard to
rubella, measles, VZV, HAV, and HBV
* Monitoring of patients who are
immunosuppressed or have had transplants
* Epidemiologic or prevalence studies
indirect examinatio serology
88
is a method used to detect and quantify viruses that have the ability to bind to red blood cells (RBCs) through a viral surface protein called hemagglutinin
hemagluttination assay
