Immuno Part 2 Lec Flashcards
1
Q
Immunoassays have been developed to detect either antigen or
antibody, and they vary from easily performed manual tests to
highly complex automated assays.
A
Precipitation Reaction
2
Q
involves combining soluble antigen with soluble antibody to
produce insoluble complexes that are visible.
A
Precipitation
3
Q
is the process by which particulate antigens aggregate to form
larger complexes when a specific antibody is present.
A
Agglutination
4
Q
is the initial force of attraction that exists between a
single Fab site on an antibody molecule and a single epitope or
determinant site on the corresponding antigen.
A
Affinity
5
Q
School-based laboratory manual test
A
Precipitation reaction
6
Q
Most efficent immmunoglobulin in this type of reaction because of its small molecules which is much efficient in Precipitation reaction
A
IgG
7
Q
indicator for agglutination reaction
A
Rbc
8
Q
most efficient; antibody involved in large agglutination reaction. Has 10 valence
A
IgM
9
Q
Large complex of agglutination
A
Clumping
10
Q
Antibodies are capable of reacting with antigens that are structurally similar to the original antigen that induced antibody production.
A
Cross-reactivity
11
Q
It is inline to affinity
A
Cross-reactivity
12
Q
initial force of attraction of antigen and antibody is called
A
Cross-reactivity
13
Q
represents the sum of all the attractive forces between an antigen and an antibody.
A
Avidity
14
Q
involves the strength with which a multivalent antibody binds a multivalent antigen, and it is a measure of the overall stability of an antigen–antibody complex
A
Avidity
15
Q
is essential to detecting the presence of an unknown, whether it is antigen or antibody
A
Avidity
16
Q
A high avidity can actually compensate for a ______ (low or high) affinity.
A
Low
17
Q
stabilization ng antigen-antibody complex binding
A
Avidity
18
Q
antigen-antibody binding
A
Affinity
19
Q
Precipitin curve, we are dealing with the
A
Zone of equivalence
20
Q
Zone of equivalence where it is inlined with the
A
Prozone and postzone
21
Q
the optimum precipitant curve in which the number of the multivalent sites of the antigen and antibody are approximately equal
A
Zone of equivalence
22
Q
phenomenon wherein we are dealing with antibody excess.
A
Prozone
23
Q
The antigen combines with only 1 or 2 antibody molecules and no cross linkages are formed
A
Prozone
24
Q
antigen excess. No lattice network formed.
A
Postzone
25
Every available antibody site is bound with single antigen and no crosslinks are formed
Postzon
26
Measurement of precipitation by
light scattering
27
Determination of Precipitation via
Passive immunodiffusion techniques
Immuno electrophoretic techniques
28
Light scattering test involves
Turbidimetry
Nephelometry
29
Passive immunodiffusion techniques test involve
Radial immunodiffusion
Ouchterlony double diffusion/immunodiffusion
30
Immunoelectrophoretic technique test involved
Rocket immunoelectrophoresis
Immunoelectrophoresis
Immunofixation electrophoresis
Countercurrent immunoelectrophoresis
31
is a measure of the turbidity or cloudiness of a solution.
Turbidimetry
32
A detection device is placed in direct line with the incident light.
Collecting light after it has passed through the solution.
Turbidimetry
33
turbidimetry has the intensity of light transmitted thru the medium, so that the unscattered light is measured at _______ degrees angle from the incident light beam
180 degrees
34
the intensity of scattered light is measured. It is usually not necessarily at right angles to the incident light beam
Nephelometry
35
It can detect antigen or antibody but we usually use antibody as reagent and the patient sample to detect the unknown antigen
Nephelometry
36
Nephelometers measure light scatter at angles ranging from ______degrees to about ______degrees.
10 degrees to 90 degrees
37
This would be the reaction allowed to run essentially to completion
which means that large particles tend to fall out of solution and decreases the amount of the scatter
End point nephelometry
38
provides accurate and precise quantitation of serum proteins, and due to automation, the cost per test is typically lower than other methods.
nephelometry
39
Nephelometry Can also be used as a quantitation of immunoglobulins such as _____, ___, and _____, ______ even the Kappa and Lambda light chain and it is exclusively done by nephelometry.
IgG, IgA, and IgM, IgE
40
Order of nephelometry device
light source→ monochromator → cuvette → lens → detector
41
The precipitation of antigen–antibody complexes can also be determined in a support medium such as a ______
Gel
42
Passive immunodiffusion techniques A high-molecular-weight complex polysaccharide derived from __________, and ___________, a___________, are used for this purpose.
seaweed, agarose and purified agar
43
The precipitation of antigen–antibody complexes can also be determined in a support medium such as a gel.
Passive immunodiffusion technique
44
The rate of diffusion is affected by the size of the particles, the temperature, the gel viscosity, and the amount of hydration.
Passive immunodiffusion techniques
45
involves the stabilization and visualization
Agar or agarose gel
46
A modification of the single-diffusion technique was the
Radial immunodiffusion
47
who was the first to use gels for precipitation reactions, and he pioneered the technique known as single diffusion.
James Oudin
48
Radial immunodiffusion has
positive result would be the formation of the
precipative bonds
49
Radial Immunodiffusion is also called as
RID
50
Antibody is uniformly distributed in the support gel, and antigen is applied to a well cut into the gel.
RID
51
The area of the ring obtained is a measure of antigen concentration, and this can be compared to a standard curve obtained by using antigens of known concentration.
RID
52
Two techniques for measurement of RID
Mancini method
Fahey and Mckelvey method
53
DIAMETER OF THE RING = antigen concentration
Mancini method
54
Mancini method occurs between _____ and ______ hours
24 and 72 hours
55
Mancini method determines
IgM and IgG
56
Determination of IgM:
completed at 50-72hrs
57
Mancini method forms:
Ring formation
58
Determination of IgG:
Complete at 24 hours
59
Mancini method involve in method of
End point method
60
the diameter is proportional to the log of the concentration
Fahey and mckelvey method
61
Fahey and mckelvey method involves method We are using measurement taken before the point of equivalence is reached
Kinetic method
62
Fahey and mckelvey methods Readings is taken for about
18 hours
63
Sources of error include in radial immunodiffusion
● overfilling or under filling the wells,
● nicking the side
● spilling sample
● improper incubation time and temperature,
● incorrect measurement.
64
Radial immunodiffusion has been used to measure
IgG, IgM, IgA, and complement components.
65
In this technique, both antigen and antibody diffuse independently through a semisolid medium in two dimensions, horizontally and vertically.
Ouchterlony double diffusion
66
has a smooth curve which means it have the same antigen that is present
Category A
67
demonstrates a cross line pattern which shows two separate reactions and indicates that the compared antigens share no common epitotes
Category B
68
there’s a fusion of 2 lines with a spur indicating partial identity.
Category C
69
Also called as ‘Laurel Technique” because it was developed by Laurel in early 1960s
Rocket immunoelectrophoresis
70
Rocket immunoelectrophoresis Also called as ‘_____________” because it was developed by Laurel in early 1960s
Laurel Technique
71
The end result of rocket immunoelectrophoresis
Conical line
72
Double-diffusion technique that incorporates electrophoresis current to enhance results.
Immunoelectrophoresis
73
Immunoelectrophoresis Introduced by __________ and __________ in 1953, this is performed as a two-major step process and can be used for semi quantitation of a wide range of antigens.
Grabar and Williams
74
this is performed as a two-major step process and can be used for semi quantitation of a wide range of antigens.
Immunoelectrophoresis
75
2 Step process of immunoelectrophoresis
Electrophoresis
Diffusion of antigen and antibodies
76
Immunoelectrophoresis Reaction takes place within_________
18-24hrs.
77
procedure has been used as screening tool for the differentiation of many serum proteins including the major classes of immunoglobulins.
Immunoelectrophoresis
78
Immunoelectrophoresis used in the laboraratory for the determination of
Myelomas
79
Immunoelectrophoresis can detect
waldenstroms
macroglobulinemia
malignant lymphomas
other lymphoproliferative disorders
80
causes increased bacterial infection
IgG deficiency
81
Either increase value of antigen or deficient
B lymphocyte deficiency
82
as first described by Alper and Johnson.
Immunofixation electrophoresis
83
Similar to immunoelectrophoresis
Immunofixation electrophoresis
84
Immunofixation electrophoresis meduim can be use
Agarose
Cellulose acetate
85
Immunofixation electrophoresis Reaction Time:
Less than 1 hour
86
Immunodiffusion takes place in a shorter time and results in a higher resolution than when antibody diffuses from a trough
Immunofixation electrophoresis
87
Modification of immunoelectrophoresis.
Countercurrent immunoelectrophoresis
88
Based on the movement of antigen towards the
anode and of the antibody towards the cathode during the passage of the electric current through agar.
Countercurrent immunoelectrophoresis
89
Countercurrent immunoelectrophoresis reaction to take place
30-60 mins
90
are designed for antigens and antibodies that may be small in size or present in very low concentrations.
Label immunoassay
91
The presence of such antigens or antibodies is determined indirectly by using a
Labeled reactant
92
The substance to be measured is known as the
Analyte
93
On this Labelled immunoassays here are some examples of analytes:
Bacterial antigen hormone
Drugs ( test kits)
Tumor markers
Specific immunoglobulins
94
is an antibody bonded to an analyte.
Labelled immunoassay
95
Labeled immunoassays have made possible rapid quantitative measurement of many important entities such as
Viral antigen
96
All the test kits that we encounter in immunology and serology are examples of Labelled immunoassays:
Mallaria test kits
Dengue NS1 for detecting recent dengue
infections
Dengue Duo for detecting IgM or IgG
Pregnancy test kits
97
Characteristic of labelled immunoassay
o Competitive Assay
o Non-Competitive Assay
98
Classification of labelled immunoassays
o Radioimmunoassay (RIA)
o Enzyme Immunoassay (EIA) o Fluorescent Immunoassay
99
All the reactants are mixed together simultaneously , and a labelled antigen competes with unlabeled patient antigen for a limited number of antibody-binding sites
Competitive immunoassay
100
Competitive immunoassay The amount of bound label is ______________ to the concentration of the patient antigen.
Inversely Proportional
101
Antibody, is first passively absorbed to a solid phase. (in solid phase we use wells)
Non-competitive immunoassay
102
Non-competitive immunoassay The amount of label measured is _________ to the amount of patient antigen
Directly proportional
103
Non-competitive immunoassay is often called as
Captured antibody
104
discovered by Georges Kohler and Cesar Milsten
Monoclonal antibodies
105
remains a constant source of highly specific antibody
Monoclonal antibodies
106
Antibodies in the Labelled immunoassays are characterized as
Monoclonal antibodies
107
Monoclonal antibodies aka
Calibrators
108
Are unlabeled analytes that are made up in known
concentrations of the substance to be measured
Calibrators
109
Most immunoassays used the change in absorbance measured by
Spectrophotometry
110
Quality Controls: should be run.
blank, negative control, and high and low positive control
111
Pioneered by Rosalyn Yalow and Solomon Berson.
Radioimmunoassay
112
Used radioactive substances as a label
Radio immunoassay
113
Most common Label for RIA is Iodine
125I (Iodine 125), 131I (Iodine 131), 3H (tritiated Hydrogen)
114
was originally based on the principle of competitive binding labelled antigen competes with unlabelled patient antigen for a limited number of antibody-binding sites.
Radioimmunoassay
115
Ria The amount of bound label is ______________ to the concentration of the labelled antigen.
Inversely Proportional
116
Advantages in using enzymes as label
cheap and readily available
Long shelf-life
easily adapted
Easy to dispose
little reagent is necessary
117
Common enzymes used in EIA:
Horseradish peroxidase (MOST COMMON)
ALP (MOST COMMON)
G-6-PD
Beta-D-galactosidase
118
Enzyme-labeled antigen competes with unlabeled patient antigen for a limited number of binding sites on antibody molecules
that are attached to a solid phase
Competitive Enzyme immuno assay
119
Similar to RIA
Enzyme immunoassay
120
Ezyme immunoassay aka
Direct elisa
121
referred to as indirect enzyme-linked immunosorbent assays (ELISA), because the enzyme-labeled reagent does not participate in
the initial antigen–antibody binding reaction.
Non-competitive enzyme immunoassay
122
Non-competitive EIA Enzymatic activity
Direct proportional
123
Membrane-based cassette assays are a relatively new type of enzyme immunoassay
Rapid immuno assay
124
The analyte is applied at one end of the strip and migrates toward the distal end, where there is an absorbent pad to maintain a constant capillary flow rate.
Immunochromatography
125
Cassette test 2 lines indicates
Positive results
126
1 line on control (C) means
Negative results
127
1 line on test (T) or no lines at all indicates
Invalid results
128
A form of immunoassay that uses label that fluoresce upon interaction with the desired analytes.
Fluorescent immunoassay
129
these fluorescent compounds are called
fluorophores or fluorochromes
130
fluorophores or fluorochromes compound
rhodamine, fluorescein, phycoerythrin
131
have a red-orange fluorescence
Phycoerythrin
132
Most commonly used fluorophores/fluorochromes:
Fluorescein
133
→ absorbs maximally at 490 to 495 nm and emits a “green color” at 517-520 nm
Fluorescein
134
absorbs at 550 nm and emits “ red colors” at 580 to 585 nm
Tetramethylrhodamine
135
there is a known antigen is fixed on the slide. This is for the antigen detection only.
Direct IFA
136
there is a known antigen fixed on the slide and it is used both for antigen and antibody detection. It involves a two step process.
Indirect IFA
