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IMMUNE1 LEC MIDTERM TRANSES > CHAPTER 6.2: AGGLUTINATION > Flashcards

CHAPTER 6.2: AGGLUTINATION Flashcards

1
Q

Technique in which molecules with a net charge are separated when an electric field is applied

A

ELECTROPHORESIS

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2
Q

Negative charged particles migrate to the

A

ANODE (+ Pole)

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3
Q

Positive charged particles migrate to

A

CATHODE (- Pole)

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4
Q

FACTORS THAT INFLUENCE RATE OF PROTEIN MIGRATION

A
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5
Q

• The bigger and the larger the size, it will be hard to migrate

A

SIZE AND SHAPE OF PROTEIN

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6
Q

• Used agarose gel and the amount of solvation has a great impact with regards to the rate of protein migration

A

AMOUNT OF SOLVATION

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7
Q

• alkaline pH

A

PH OF BUFFER: >8

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8
Q

• Room temperature

A

TEMPERATURE

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9
Q

• Protein will denature once it is exposed to high temperature

A

TEMPERATURE

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10
Q

• flow of ions goes toward the cathode and can impede movement of proteins toward the anode

A

ENDO-OSMOSIS

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11
Q

DIFFERENT TESTS FOR ELECTROPHORESIS

A
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12
Q

Laurell Technique (1960)

A

ROCKET IMMUNOELECTROPHORESIS

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13
Q

Radial immunodiffusion (RID) + electrophoresis

A

ROCKET IMMUNOELECTROPHORESIS

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14
Q

Single reactant moving in one dimension

A

ROCKET IMMUNOELECTROPHORESIS

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15
Q

Electrophoresis is used to facilitate migration of the antigen into the agar

A

ROCKET IMMUNOELECTROPHORESIS

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16
Q

End result: precipitin line that is conical in shape, resembling a rocket

A

ROCKET IMMUNOELECTROPHORESIS

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17
Q

The height of the rocket, measured from the well to the apex, is directly in proportion to the amount of antigen in the sample.

A

ROCKET IMMUNOELECTROPHORESIS

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18
Q

This technique has been used to quantitate immunoglobulins, using a buffer of pH 8.6

A

ROCKET IMMUNOELECTROPHORESIS

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19
Q

ROCKET IMMUNOELECTROPHORESIS Procedure:
1. Antigen is pushed through antibody containing gel under influence of an (?)
2. When they are equivalence, precipitation will occur forming a (?)

A

applied electric field

cone/ rocket shape band

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20
Q

• Ressler’s method

A

CROSSED IMMUNOELECTROPHORESIS

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21
Q

• Single reactant moving in 2 dimensions

A

CROSSED IMMUNOELECTROPHORESIS

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22
Q

CROSSED IMMUNOELECTROPHORESIS Procedure
1. Proteins are separated by (?)
2. Proteins are subjected to a 2nd electrophoresis where they will move through a (?) until rocket is formed (Ag-Ab reach equivalence)

A

electrophoresis

Ab-containing agarose

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23
Q

• Countercurrent electrophoresis

A

COUNTER IMMUNOELECTROPHORESIS

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24
Q

• Voltage Facilitated double immunodiffusions

A

COUNTER IMMUNOELECTROPHORESIS

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25
Q

• Double reactants moving in one dimension

A

COUNTER IMMUNOELECTROPHORESIS

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26
Q

COUNTER IMMUNOELECTROPHORESIS Use:

A

Identify bacterial, fungi or virus in fluids

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27
Q

COUNTER IMMUNOELECTROPHORESIS Procedure:
1. Ag and Ab are added to separate parallel wells cut out in an (?)
2. When an electric field is applied, the Ag will migrate to the (?) and Ab to the (?)
3. Zone of equivalence will form a (?)

A

agar gel

Anode; cathode

precipitate

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28
Q

• Grabar and Williams

A

CLASSIC IMMUNOELETROPHORESIS

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29
Q

• Double reactants moving in 2 dimensions

A

CLASSIC IMMUNOELETROPHORESIS

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30
Q

• Two-step process

A

CLASSIC IMMUNOELETROPHORESIS

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31
Q

• Used as a screening tool for the differentiation of many serum proteins, including the major classes of immunoglobulins.

A

CLASSIC IMMUNOELETROPHORESIS

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32
Q

• It is both a qualitative and a semiquantitative technique and has been used in clinical laboratories for the detection of myelomas, Waldenström’s macroglobulinemia, malignant lymphomas, and other lymphoproliferative disorders.

A

CLASSIC IMMUNOELETROPHORESIS

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33
Q

CLASSIC IMMUNOELETROPHORESIS Use:

A

Differentiate the Ig Class, identify abnormal proteins, myeloma proteins, Monitor purity of pharmaceutical products

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34
Q

CLASSIC IMMUNOELETROPHORESIS Procedure:
1. Ag is introduced in a well and an electric field is applied resulting in separation of proteins
2. Ab is introduced in a trough parallel to the separated protein
3. Ag-Ab complex form

A
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35
Q

CLASSIC IMMUNOELETROPHORESIS Procedure:
1. Ag is introduced in a well and an (?) is applied resulting in separation of proteins
2. Ab is introduced in a (?) parallel to the separated protein
3. (?) form

A

electric field

trough

Ag-Ab complex

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36
Q

CLASSIC IMMUNOELETROPHORESIS

Sequence:
• Cathode (+) to Anode (–)

A
  1. Albumin
  2. Alpha-1 globulin
  3. Alpha-2 globulin
  4. Beta globulin
  5. Gamma globulin
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37
Q

Immunoglobulin

A

Gamma globulin

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38
Q

STEPS IN AGGLUTINATION

A
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39
Q

published the first report about the ability of antibody to clump cells, based on observations of agglutination of bacterial cells by serum.

A

Gruber and Durham

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40
Q

This finding gave rise to the use of serology as a tool in the diagnosis of disease, and it also led to the discovery of the ABO blood groups (1902)

A

Gruber and Durham

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41
Q

Process by which (?) such as cell aggregate to form larger complexes when a (?) is present

A

particulate antigens (agglutinogen)

specific antibody (agglutinin)

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42
Q

Antigen-Antibody reaction

A

SENSITIZATION

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43
Q

Stabilization of agglutinogen + agglutinin

A

SENSITIZATION

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44
Q

Stabilization of antigen–antibody complexes with the binding together of multiple antigenic determinants.

A

SENSITIZATION

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45
Q

is affected by the nature of the antibody molecules themselves

A

SENSITIZATION

46
Q

Best antibody for agglutination is IgM

A

SENSITIZATION

47
Q

IgM

potential valence of 10 is over (?) more efficient in agglutination than is IgG with a valence of 2

A

700 times out of

48
Q

Cross linking

A

LATTICE FORMATION

49
Q

Representing the sum of interactions between antibody and multiple antigenic determinants on a particle (Avidity)

A

LATTICE FORMATION

50
Q

There is visible agglutination

A

LATTICE FORMATION

51
Q

FACTORS THAT AFFECT AGGLUTINATION

52
Q
  1. Buffer pH Routine:

(close to physiological pH)

53
Q

Affects the zoning phenomenon

A

2.Relative concentration of Ag and Ab

54
Q

• Abs will not detect determinants buried within the particle

A
  1. Location and concentration of Antigenic determinants of the particle
55
Q

• More number of determinants, the higher the likelihood of cross bridging

A
  1. Location and concentration of Antigenic determinants of the particle
56
Q

Non covalent interaction

A
  1. Electrostatic interactions between particles
57
Q

• in the buffer plays an important role in agglutination

A
  1. Electrolyte concentration (ionic strength)
58
Q

• Electrolytes reduce (?) that interfere with lattice formation

A

electrostatic charges

59
Q
  1. Antibody isotope Best:
60
Q
  1. Temperature
    : Cold reacting (range 4-22oC)
    : Warm reacting with optimum temperature at 37oC
61
Q
  1. Time of incubation of coated particles with patient’s serum Incubation times ranges from
A

15-60 minutes

62
Q

0

A

No agglutinates
Dark, turbid, homogenous

63
Q

W+

A

Many tiny agglutinates, many free cells, may not be visible without microscope
Dark, turbid

64
Q

1+

A

Many small agglutinates, many free cells (25% are agglutinated)
Turbid

65
Q

2+

A

Many medium sized agglutinins, moderate number of free cells (50% are agglutinated)
Clear

66
Q

3+

A

Several large agglutinates, few free cells (75% are agglutinated)
Clear

67
Q

4+

A

One large solid agglutination, no free (100% are agglutinated)
Clear

68
Q

MAJOR CATEGORIES OF AGGLUTINATION REACTIONS

69
Q

It will not use any carrier particle

A

DIRECT/ ACTIVE AGGLUTINATION

70
Q

Detecting the presence of antigen

A

DIRECT/ ACTIVE AGGLUTINATION

71
Q

Occurs when antigens are found naturally on a particle

A

DIRECT/ ACTIVE AGGLUTINATION

72
Q

Reaction is due to an Ag-Ab reaction where in the Ag is inherent native to the cell

A

Direct Immune

73
Q

Example: ABO grouping (hemagglutination), Widal Test
• ABO antigens are found in the RBC
• Reagent: Antisera

A

Direct Immune

74
Q

Aggregation of indicator rod blood cells are NOT due to AgAb reactions

A

Direct Non Immune

75
Q

Example: Viral Hemagglutination test

A

Direct Non Immune

76
Q

Direct non immune agglutination

A

VIRAL HEMAGGLUTINATION

77
Q

Virus can stick to agglutinate RBC in the process

A

VIRAL HEMAGGLUTINATION

78
Q

Rubella virus, dengue virus, influenza virus, mumps virus

A

VIRAL HEMAGGLUTINATION

79
Q

Viral receptor: Peplomers

A

VIRAL HEMAGGLUTINATION

80
Q

Competitive binding Assay Procedure:
1. Patient serum incubated with (?)
2. Viral particles will bind to the (?)
3. (?) added to reaction mixture
4.
Positive result: Negative result:

A

viral particles (Commercially available)

Fab region of Anti-viral Abs

Indicator RBCs

Inhibition or Absence of Agglutination; (Presence of Ab)

Agglutination

81
Q

Reactions where Ag has been fixed or absorbed to a carrier/ inert particle

A

INDIRECT/ PASSIVE AGGLUTINATION

82
Q

Detecting the presence of antibodies

A

INDIRECT/ PASSIVE AGGLUTINATION

83
Q

Example: Antistreptolysin-O (ASTO)

A

INDIRECT/ PASSIVE AGGLUTINATION

84
Q

Different passive carriers:

A

o Human RBCs
o Clay (Bentonite)
o Latex particles
o Colloidal gold
o Charcoal particles

85
Q

Antibody is bound to the carrier

A

REVERSE PASSIVE AGGLUTINATION

86
Q

The antibody must still be reactive and is joined in such a manner that the active sites are facing outward.

A

REVERSE PASSIVE AGGLUTINATION

87
Q

Fluid is detected for the presence of Ag

A

REVERSE PASSIVE AGGLUTINATION

88
Q

Example: CRP, Reverse agglutination test for Candida and Nisseria

A

REVERSE PASSIVE AGGLUTINATION

89
Q

Patient sample (Ag) incubated with Ab in test kit

A

LATEX PARTICLE AGGLUTINATION INHIBITION

90
Q

Complex will form if the patient sample contains the corresponding Ag and the Fab sites are no longer available for the Ag-coated latex particles

A

LATEX PARTICLE AGGLUTINATION INHIBITION

91
Q

Reactions are based on competition between particulate and soluble antigens for limited antibody-combining sites, and a lack of agglutination is an indicator of a positive reaction

A

LATEX PARTICLE AGGLUTINATION INHIBITION

92
Q

If the patient sample has no free hapten, the reagent antibody is able to combine with the carrier particles and produce a visible agglutination. In this case, however, agglutination is a negative reaction

A

LATEX PARTICLE AGGLUTINATION INHIBITION

93
Q

Example: HCG/ pregnancy test

A

LATEX PARTICLE AGGLUTINATION INHIBITION

94
Q

Agglutination inhibition.
(?) is added to the patient sample. If patient antigen is present, (?) results. When (?) are added, no agglutination occurs, which is a positive test. If no patient antigen is there, the (?), and agglutination results, which is a negative test.

A

Reagent antibody

antigen–antibody combination

antigen-coated latex particles

reagent antibody combines with latex particles

95
Q

systems using bacteria as the inert particles to which antibody is attached

A

Coaglutination

96
Q

Coaglutination

(?) is most frequently used, because it has a protein on its outer surface, called protein A, which naturally adsorbs the fragment crystallizable (FC) portion of antibody molecules.

A

Staphylococcus aureus

97
Q

The active sites face outward and are capable of reacting with specific antigen

A

Coaglutination

98
Q

particles nonspecifically bind the FC portion of immunoglobulin molecules. When reagent antibody is used, combination with patient antigen produces a visible agglutination reaction.

A

Staphylococcus aureus

99
Q

detects non agglutinating antibody by means of coupling with a second antibody

A

ANTIGLOBULIN TEST

100
Q

Detects IgG Ab bound to Ag on Red cells (in-vivo)

A

Direct Antiglobulin Test

101
Q

Direct Antiglobulin Test Purpose:

A

• HDN investigation
• HTR investigation
• AIHA (Autoimmune Hemolytic Anemia)
• Drug induced Hemolytic Anemia

102
Q

Direct Antiglobulin Test Example:

A

Direct Coomb’s test

103
Q

• Detects presence of Abs in the serum that is still to be attached to an analyte

A

Indirect Antiglobulin Test

104
Q

Indirect Antiglobulin Test Purpose:

A

o Crossmatching
o Ab determination
o Ab identification
o RBC Ag phenotyping

105
Q

Example:

A

Indirect Coomb’s test

106
Q

QUANTITATIVE AGGLUTINATION REACTION Best

107
Q

Gold-inorganic colloidal particle

A

SPIA/ Sol Particle Immunoassay

108
Q

Dye-organic colloidal particle

A

DIA/ Disperse Dye Immunoassay

109
Q

Latex particle

A

IMPACT/ Immunoassay by Particle Counting

110
Q

IMPACT/ Immunoassay by Particle Counting

IMMUNE1 LEC MIDTERM TRANSES (4 decks)

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