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Immunology and Serology [LEC] > [DISCUSSION] MODULE 1 UNIT 5 > Flashcards

[DISCUSSION] MODULE 1 UNIT 5 Flashcards

1
Q

-MAJOR HISTOCOMPATIBILITY COMPLEX IS ALSO CALLED

A

Human Leukocyte antigens (HLA)

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2
Q

-MAJOR HISTOCOMPATIBILITY COMPLEX PRPONENT

A

Dausset

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3
Q
  • First defined by discovering an antibody response to circulating wbcs
A

MAJOR HISTOCOMPATIBILITY COMPLEX

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4
Q

– Determine whether transplanted tissue is histocompatible and thus accepted or recognized as foreign and rejected

A

MAJOR HISTOCOMPATIBILITY COMPLEX

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5
Q

-all nucleated cells in the body

A

(Class I)

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6
Q

-professional APC’s (dendritic cells, macrophages, B cells)

A

(Class II)

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7
Q

-Play a pivotal role in the development of both humoral and cellular immunity

A

MAJOR HISTOCOMPATIBILITY COMPLEX

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8
Q

MHC Main Function

A
  • Bring antigen to the cell surface for recognition by T cells, because T cell activation will occur only when antigen is combined with only MHC molecules
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9
Q

– Clinical relevance of MHC

A

transfusion reactions
graft rejection
autoimmune diseases

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10
Q

– Genes controlling expression of these molecules are actually a system of genes known as the

A

MAJOR HISTOCOMPATIBILITY COMPLEX (MHC)

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11
Q

-Most polymorphic system found in humans

A

Genes coding for MHC molecules

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12
Q

– Found on the short arm of chromosome 6 (6p)

A

GENES CODING FOR MHC MOLECULES

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13
Q

CATEGORIES OF GENES CODING FOR MHC MOLECULES

A

Class I molecules
Class II molecules
Class III molecules

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14
Q

classical class I molecules

A

HLA-A, HLA-B and HLA-C antigens

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15
Q

classical class II molecules

A

D region → HLA-DR, HLA-DQ, HLA-DP

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16
Q

classical class Ill molecules

A

Code for complement proteins and cytokines such as tumor necrosis factor

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17
Q
  • Polymorphic
A

MAJOR HISTOCOMPATIBILITY COMPLEX

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18
Q
  • There are so many possible alleles at each location
  • E.g., at least 580 alleles of HLA-A
A
  • Polymorphic
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19
Q
  • alleles of HLA-B
A

921

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20
Q
  • alleles of HLA-C
A

312

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21
Q
  • Probability that any two individuals will express the
    same MHC molecules is
A

very low

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22
Q

GENES ARE

A

CODOMINANT

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23
Q

Consists of a structurally distinct a chain associated with a second, shorter polypeptide called b2
-microglobulin

A

CLASS I MHC MOLECULES

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24
Q

is organized in three folded domains ( a1
, a2
, and a3
) has a
carboxy-terminal membrane anchor

A

Class I a chain

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25
Q

(?), with one folded domain, is linked to the
membrane only indirectly through its association with the a chain

A

smaller b2
-microglobulin

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26
Q

Critical for stabilizing the class I molecule and for facilitating its transport to the cell surface

A

smaller b2
-microglobulin

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27
Q

The peptide-binding site in a class I protein is formed by the

A

a1 and a2 domains

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28
Q

Can only accommodate peptides that are (?) amino acids long

A

8 – 10

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29
Q

are similar to the constant regions found in immunoglobulin molecules

A

a3 and b2 regions

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30
Q

reacts with CD8 on cytotoxic T cells

A

a3 region

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31
Q

Expressed on all nucleated cells, but differ in the level of expression

A

CLASS I

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32
Q

Highest on lymphocytes and lowest on liver
hepatocytes, neural cells and muscle cells

A

CLASS I

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33
Q

are expressed at a lower level
than HLA-A and HLA-B antigens so the latter two
are the most important to match for transplantation

A

HLA-C antigens

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34
Q

Another group of molecules called the nonclassical class I antigens

A

HLA-E, HLA-F, HLA-G

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35
Q

are not expressed on cell surfaces and do not
function in antigen recognition but may play other roles in the immune
response

A

HLA-E, HLA-F

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36
Q

Expressed on trophoblast cells during the first trimester of pregnancy

A

HLA-G

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37
Q

Found primarily on antigen-presenting cells (B cells, dendritic cells, monocytes, macrophages)

A

CLASS II

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38
Q

Consist of two (2) noncovalently bound polypeptide chains that are both encoded by genes in the MHC complex

A

CLASS II

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39
Q

DR is expressed at the highest level

A

CLASS II

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40
Q

Accounts for ½ of the all the class II molecules on a particular cell

A

DR

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41
Q

Both the a chain (MW = 33 kD) and the b chain (MW = 27 kD) are anchored to the cell membrane

A

CLASS II

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42
Q

Each has two domains

A

CLASS II

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43
Q

Peptide binding site →formed by the a1 and b1 domains

A

CLASS II

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44
Q

Both ends of the peptide-binding cleft are open

A

CLASS II

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45
Q

Allow the capture of longer peptides ( 9 – 20 amino acids) than is the case for class I molecules

A

CLASS II

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46
Q

CD4 contacts sequences in the b2 domain

A

CLASS II

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47
Q

Nonclassical class II genes

A

HLA-DM, HLA-DN, HLA-DO

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48
Q

Products of these genes play a regulatory role in antigen processing

A

HLA-DM, HLA-DN, HLA-DO

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49
Q

ENDOCYTIC PATHWAY Major antigen
sources

A

Endocytosed extracellular
proteins (host & foreign)
Membrane proteins (host &
foreign)

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50
Q

CYTOSOLIC PATHWAY Major antigen
sources

A

Cytosolic proteins of host or
intracellular pathogens (viral,
bacterial, parasitic)
Signal peptides (host & foreign)

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51
Q

ENDOCYTIC PATHWAY Processing machinery

A

Lysosomal enzymes

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52
Q

CYTOSOLIC PATHWAY Processing
machinery

A

Proteasomes (including low-
molecular-weight protein (LMPs)

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53
Q

ENDOCYTIC PATHWAY Cell types where active

A

Professional APCs

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54
Q

CYTOSOLIC PATHWAY Cell types where active

A

All nucleated cells

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55
Q

ENDOCYTIC PATHWAY Site of antigen –
MHC binding

A

Endocytic vesicles,
prelysosomes

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56
Q

CYTOSOLIC PATHWAY Site of antigen –
MHC binding

A

Rough endoplasmic reticulum

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57
Q

ENDOCYTIC PATHWAY MHC utilized

A

Class II

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58
Q

CYTOSOLIC PATHWAYbMHC utilized

A

Class I

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59
Q

ENDOCYTIC PATHWAY Presents to

A

CD4 (helper) T cells

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60
Q

CYTOSOLIC PATHWAY Presents to

A

CD8 (cytotoxic) T cells

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61
Q

CLINICAL SIGNIFICANCE OF MHC

A
  • MHC molecules can induce a response that leads to graft
    rejection
  • Play a role in development of autoimmune diseases
  • Determine the type of peptides to which an individual can mount
    an immune response
  • Presence of a particular MHC protein may confer additional
    protection (e.g., HLA-B8 and increased resistance to HIV)
  • Future developments to tailor vaccines to certain groups of
    molecules
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62
Q

Ankylosing spondylitis

63
Q

Birdshot retinopathy

64
Q

Celiac disease

A

HLA-DR3, - DR5, - DR7

65
Q

Graves’ disease

66
Q

Narcolepsy

67
Q

Multiple sclerosis

68
Q

Rheumatoid arthritis

69
Q

Type 1 diabetes mellitus

A

HLA-DQ8, - DQ2, - DR3, - DR4

70
Q

HISTOCOMPATIBILITY
TESTING: APPLICATIONS

A
  1. Prevention of graft rejection / graft vs. host reaction
  2. Paternity exclusion
  3. Disease associations
  4. Prevent platelet refractoriness in Platelet transfusions
  5. Prevent Transfusion-related acute lung injury (TRALI)
  6. Hematopoietic stem cell transplantation
71
Q

HISTOCOMPATIBILITY
TESTING

A
  1. Tissue typing
  2. Antibody screening
  3. Tissue matching/ Crossmatching
72
Q
  1. Tissue typing
A

-HLA Phenotyping: Serologic techniques
-HLA Genotyping: Molecular methods

73
Q

Purified Lymphocyte suspension for Antigen detection
- Anticoagulated whole blood is overlaid into:

A

Ficoll-Hypaque reagent
Then centrifuge

74
Q

Purified Lymphocyte suspension for Antigen
Use T lymphocytes

A

Class I Antigens (HLA-A, HLA-B, HLA-C)

75
Q

Use B lymphocytes

A

Class II Antigens (HLA-DR, HLA-DP, HLA-DQ)

76
Q

B lymphocyte separation:

A
  1. Nylon wool separation
  2. Use immunomagnetic beads
  3. Fluorescent labeling (use FITC)
77
Q

Sources of Antibodies

A
  1. Multiparous women
  2. Patients who received multiple transfusions
    (WBC and platelets)
  3. Volunteers who were sensitized by blood
    transfusion or tissue grafts
  4. Patients who have rejected a transplanted
    kidney
78
Q

SEROLOGIC METHODS: TISSUE
TYPING

A

Lymphocytotoxicity Test (Complement-dependent
microlymphocytotoxicity)

79
Q

Expose unknown cell to a battery of antisera of known
HLA specificity

A

Lymphocytotoxicity Test (Complement-dependent
microlymphocytotoxicity)

80
Q

Lymphocytotoxicity Test (Complement-dependent
microlymphocytotoxicity)

A

-Incubate at room temperature for 30 minutes

81
Q

Lymphocytotoxicity Test (Complement-dependent
microlymphocytotoxicity)

A
  • Complement is added (rabbit serum)
82
Q

Lymphocytotoxicity Test (Complement-dependent
microlymphocytotoxicity)

A

-Incubated at room temperature for 60 minutes

83
Q

SEROLOGIC METHODS: TISSUE
TYPING

A

Lymphocytotoxicity Test (Complement-dependent
microlymphocytotoxicity)

84
Q

Lymphocytotoxicity Test

  • Expose unknown cell to a battery of antisera of known (?)
    -Incubate at room temperature for (?)
  • Complement is added (?)
    -Incubated at room temperature for
  • Then add (?) (eosin Y)
  • Take an aliquot from well and examine under light microscope using (?)
  • (?) take up the dye
A

HLA specificity
30 minutes
rabbit serum
60 minutes
trypan blue dye
hemocytometer
Dead cells

85
Q

Dead cells

A

Flattened, appear large, dark and nonrefractile

86
Q

Unaffected cells

A

small, bright and refractile

87
Q

1 Negative
2 Doubtful positive
4 Weak positive
6 Positive
8 Strong positive

A

0 – 10
11 – 20
21 – 50
51 – 80
81 – 100

88
Q

1 Negative
2 Doubtful positive
4 Weak positive
6 Positive
8 Strong positive

A

0 – 10
11 – 20
21 – 50
51 – 80
81 – 100

89
Q

SEROLOGIC METHODS: TISSUE TYPING:
Complement-Dependent Lymphocytotoxicity

90
Q

TISSUE TYPING: Complement-
Dependent Lymphocytotoxicity

To increase sensitivity:

A
  1. Extended incubation
  2. The Amos wash step
  3. Antihuman globulin
91
Q

Lymphocytotoxicity Disadvantages:

A
  1. It requires having cells available for testing.
  2. It is necessary to collect leukocytes and perform cellular testing in a timely manner to
    enable transplantation
  3. it is necessary to maintain reliable and consistent antigen panels that represent a broad range of HLA antigens
92
Q

Detect antibodies in patients who are candidates for
transplant

A

SEROLOGIC METHODS: Antibody
Screening

93
Q

Antibody
Screening

A
  1. Complement-dependent lymphocytotoxicity/
    Microlymphocytotoxicity
  2. ELISA
  3. Flow cytometry
94
Q

MICROLYMPHOCYTOTOXICITY
PERCENT PANEL REACTIVE ANTIBODY (%PRA)

A

Antibody Screening

95
Q

The proportion of lymphocytes in the panel that are killed by the patient’s serum

A

PERCENT PANEL REACTIVE ANTIBODY (%PRA)

96
Q

Utilize purified HLA antigens bound to the wells of microtiter plates.

97
Q

-Patient serum is added

98
Q

-If HLA-specific antibody is present, it will bind

99
Q

-Bound antibody is detected by addition of an enzyme-labeled anti immunoglobulin reagent

100
Q

Serves as a qualitative screen for the presence of HLA antibody in a serum

101
Q

-Recognizes false-positive reactions

102
Q

-Distinguishes Class I from Class II

103
Q

-Differentiates IgM from IgG antibodies

104
Q

-Increased specificity

105
Q

Detects antibody binding directly

A

Flow Cytometry

106
Q
  • Can distinguish between IgM and IgG
A

Flow Cytometry

107
Q
  • Uses T or B cells; or purified HLA antigens coated with microparticles
A

Flow Cytometry

108
Q
  • Bound antibody is detected by adding an FITC-labeled anti-IgGreagent
A

Flow Cytometry

109
Q
  • Percent PRA is determined
A

Flow Cytometry

110
Q
  • Most sensitive, most specific
A

Flow Cytometry

111
Q

CROSSMATCHING

A
  • Lymphocytotoxicity
    -Flow cytometry
    -ELISA being developed
112
Q

SEROLOGIC METHODS:

113
Q

MOLECULAR METHODS:

114
Q

HLA GENOTYPING:

A

A. Restriction Fragment Length Polymorphism (RFLP)
B. PCR- based

115
Q

PCR- based

A
  1. Sequence-Specific oligonucleotides (SSO)
  2. Sequence- Specific Primer(SSP)
  3. Sequence-based typing (SBT)
116
Q

Restriction enzymes (restriction endonucleases) are used

A

Restriction Fragment Length Polymorphism
(RFLP)

117
Q
  • Cleaves genomic DNA
A

Restriction Fragment Length Polymorphism
(RFLP)

118
Q
  • Obtain a pattern of fragmentation
A

Restriction Fragment Length Polymorphism
(RFLP)

119
Q
  • Degree of disparity between donor and recipient can be assessed by COMPARING patterns of fragmentation
A

Restriction Fragment Length Polymorphism
(RFLP)

120
Q

Automated, rapid and in vitro technique

121
Q

-Allows direct amplification of a particular
DNA sequence

122
Q

-Selected by the use of primers that border
the genes of interest

123
Q

PCR-amplification of a chosen sequence using primers flanking the sequence

A

Sequence-
Specific Oligonucleotides (SSO)

124
Q
  • The amplified DNA is immobilized on a membrane
A

Sequence-
Specific Oligonucleotides (SSO)

125
Q
  • Then hybridized with selected, labeled
    oligonucleotide probes
A

Sequence-
Specific Oligonucleotides (SSO)

126
Q

Oligonucleotide primers are designed to obtain amplification of specific alleles or groups of alleles

A

Sequence-
Specific Primers (SSP)

127
Q
  • Assignment of allele is based on the presence or absence of amplified product
A

Sequence-
Specific Primers (SSP)

128
Q
  • Detected by agarose gel electrophoresis (AGE) and transillumination
A

Sequence-
Specific Primers (SSP)

129
Q

SBT Two Methods:

A
  • Sanger-based DNA sequencing
  • Next-generation DNA sequencing (NGS)
130
Q

Allows amplification of the most polymorphic regions of the HLA genes

A

Sequence
Based Typing (SBT)

131
Q

-Preferred method for hematopoietic stem cell transplantation

A

Sequence
Based Typing (SBT)

132
Q

Performed by terminal-end incorporation of fluorescently labeled nucleotides during PCR reactions

A

Sequence
Based Typing (SBT)

133
Q

-Allows amplification of the most polymorphic regions of the HLA genes

A

Sequence
Based Typing (SBT)

134
Q

-Preferred method for hematopoietic stem cell transplantation

A

Sequence
Based Typing (SBT)

135
Q

Donor and recipient cells are cultured together for several days

A

Mixed
Lymphocyte Reaction

136
Q
  • Allow CD4+ T cells to be activated and proliferate
A

Mixed
Lymphocyte Reaction

137
Q
  • In response to disparate Class II antigens
A

Mixed
Lymphocyte Reaction

138
Q
  • Amount of proliferation is measured and used to predict the magnitude of rejection
A

Mixed
Lymphocyte Reaction

139
Q
  • Can be done in a One-way MLR or a Two-way MLR

One-Way MLR → used to test for recipient’s
response to donor cells

A

Mixed
Lymphocyte Reaction

140
Q

-Donor cells are irradiated (using Cobalt or
Cesium) or treated with mitomycin C

A

Mixed
Lymphocyte Reaction

141
Q

-Most useful for bone marrow grafts and in
cases of living related donors

A

Mixed
Lymphocyte Reaction

142
Q

Radioactive label is added on day 5.

A

Mixed
Lymphocyte Reaction

143
Q

-Beta ray emissions are measured using a liquid scintillation counter (counts per minute)

A

Mixed
Lymphocyte Reaction

144
Q

-CPM correlates with the amount of
proliferation

A

Mixed
Lymphocyte Reaction

145
Q

-Depends on the degree of disparity between the recipient cells and potential donor cells

A

Mixed
Lymphocyte Reaction

146
Q

Types of Grafts

A
  1. Autograft
  2. Syngraft
  3. Allograft (Homograft)
  4. Xenograft (Heterograft)
147
Q

Most Common Tissues used for
Transplantation

A
  1. Kidney
  2. Heart
  3. Cornea
  4. Lung
  5. Skin
  6. Bone marrow
148
Q

HOST RESPONSE TO TRANSPLANTATION

A
  1. Hyperacute Rejection
  2. Acute or Accelerated rejection
  3. Chronic rejection
  4. Graft-versus-Host Disease
149
Q

Graft-versus-Host Disease

A
  • Acute GVHD
  • Chronic GVHD
150
Q

Induce intense immunosuppression in the
initial days post transplantation

A

Immunosuppressive Therapy

151
Q

-Maintenance of the graft

A

Immunosuppressive Therapy

152
Q

-Reversal of established rejection

A

Immunosuppressive Therapy

153
Q

Types of Immunosuppressive Therapy

A
  1. Corticosteroids
  2. Cyclosporine (Cyclosporin A)
  3. Tacrolimus
  4. Cytotoxic drugs
  5. Antilymphocyte (Antithymocyte) globulin
  6. Monoclonal antibodies
154
Q

Complications of Transplantation

A
  • Cancer
    -Osteoporosis
    -Diabetes
    -Hypertension
    -Hypercholesterolemia

Immunology and Serology [LEC] (15 decks)

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