Section 5 Flashcards
(196 cards)
1
Q
What are acute phase reactants?
A
- proteins that increase due to infection, injury, trauma (c-reactive protein, alpha-1 antitrypsin, haptoglobin, fibrinogen, ceruplasmin, alpha-1 acid glycoprotein, complement)
2
Q
What is a alloantibody?
A
- antibody formed in response to antigen from individuals of same species
3
Q
What is an antigen (Ag)?
A
- foreign substance that can stimulate antibody production
- most often a large, complex molecule (molecular weight >10,000), usually protein or polysaccharide
4
Q
What is antibody (Ab)?
A
- immunoglobulin (Ig) produced by plasma cells in response to Ag
5
Q
What is an autoantibody?
A
- Ab against self
6
Q
What is avidity?
A
- strength of bond between Ag and Ab
7
Q
What is chemokines?
A
- cytokines that attract cells to particular site
- important an inflammatory response
8
Q
What is chemotaxis?
A
-migration of cells toward chemokine
9
Q
What is cluster of differentiation (CD)?
A
- antigenic features of leukocytes
10
Q
What is a cytokine?
A
- chemicals produced by activated immune cells that affect function of other cells
- includes interferons, chemokines, tumor necrosis factors
- transforming growth factors, transforming growth factors, colony stimulating factors, interleukins
11
Q
What is a epitope?
A
- determinant site on Ag
12
Q
What is a hapten?
A
- low molecular weight substance that can bind to Ab once its formed, but is incapable of stimulating Ab production unless bound to larger carrier molecule
13
Q
What is hypersensitivity?
A
- heightened state of immune responsiveness that can cause tissue damage in host
14
Q
What is immunity?
A
- resistance to infection
15
Q
What is an immunogen?
A
- any substance capable of inducing immune response
16
Q
What is an immunoglobin (Ig)?
A
- antibody
17
Q
What is inflammation?
A
- cellular and humoral mechanisms involved in reaction to injury or infection
18
Q
What are interferons?
A
- cytokines with antiviral properties
- also active against certain tumors and inflammatory processes
19
Q
What are interleukins?
A
- cytokines produced by leukocytes that affect inflammatory response through increased in soluble factors or cells
20
Q
What is a ligand?
A
- molecule that binds to another molecule of complementary configuration; the substance being measured in an immunoassay
21
Q
What is a lysozyme?
A
- enzyme found in tears and saliva that attacks cell walls of microorganisms
22
Q
What is Major Histocompatibility Complex (MCH)?
A
- system of genes that control expression of MHC molecules found on all nucleated cells; originally referred to as human leukocyte antigens (HLA)
23
Q
What is a monoclonal lab?
A
- Ab derived from a single B-cell clone, frequently used in clinical laboratory assays such as ELISA
24
Q
What is opsonin?
A
- serum protein that attach to foreign substance and enhance phagocytosis
- most often complement or Ab
25
What is phagocytosis?
- step-wise engulfment of cells or particulate matter by neutrophils and macrophages
26
What are plasma cells?
- transformed B cells that secure Ab
27
What is a polyclonal Ab?
- Ab produced by many B cell clones
28
What is postzone?
- reduces ag/ab complexes due to Ag excess
- can cause a false NEG in serological test for Ab.
- repeat test in 1-2 weeks
29
What is prozone?
- reduced ag/ab complexes due to Ab excess
- can cause false NEG in serological test for Ab.
- dilute serum and retest
30
What is seroconversion?
- change of serological test from NEG to POS due to development of detectable Ab
31
What is the thymus?
- small, flat bilobed organs found in thorax; site of T-lymphocyte development
- one to two primary lymphoid organs
32
What are the primary Lymphoid organs?
- Bone marrow
- thymus
33
What is titer?
- means of expressing Ab concentration; reciprocal of highest dilution with POS rxn
34
What is a vaccination?
- injection of immunogenic material to induce immunity
35
What is zone of equivalence?
- when # of multivalent sites of Ag and Ab are approximately equal
- results in optimal precipitation
36
What are the 2 branches of the immune system?
- cellular
- humoral
37
Describe cellular immunity
- cell mediated
- defense against = viruses, fungi, mycobacteria, other intracellular pathogens, tumor cells
- T cells, macrophages are involved
- examples = graft rejection, hypersensitivity rxn, elimination of tumor cells
38
Describe humoral immunity
- Ab mediated
- defense agaisnt = bacteria (extracellular)
- B cells, and plasma cells are involved
- example = Ab production
39
Describe innate immunity
- defense mechanism present at birth
- not Ag specific
- external defense system: intact skin, mucous membrane, cilia and mucus in respiratory tract, stomach acid, flushing of urine, lactic acid in vagina, lysozyme in tears and saliva, normal Floral
- internal defense system: neutrophils, macrophages, acute phase reactants, complement, chemokines
- memory = NO
40
Describe acquired immunity
- also known as acquired
- defense mechanism that are Ag specific
- T cells, B cells, plasma cells, Ab, and cytokines involved
- memory = YES
41
Describe naturally acquired active immunity
- individual infected with microorganism produces Ab
- example = clinical or sub clinical infection
- specific
- not immediate
- long term
42
Describe artificially acquired active immunity
- individual exposed to Ag through vaccine develops immunity without having infection
- example = diphtheria, tetanus, pertussis vaccine (DTaP), measles, mumps, rubella, vaccine (MMR), polio, tetanus, haemophilus influenza type B (Hib) vaccine
- specific
- not immediate
- long term
43
Describe Naturally acquired passive immunity
- individual protected by Abs produced by another person
- examples = material Abs that cross placenta and are present in breast milk
- specific
- immediate
- not long term
44
Describe artificially acquired passive immunity
- individual receives immune globulin containing Abs produced by another person
- examples = Rh immune globulin, convalescent plasma, antitoxins
- specific
- immediate
- not long term
45
What are the cells of the innate immune system?
- neutrophils
- eosinophils
- basophils
- monocytes
- mast cells
- macrophages
- dendritic cells
- Natural killer cells (NK)
46
Describe neutrophils in innate immunity
- phagocytosis, inflammatory response
- respond to chemotaxins
- Granules contain bactericidal enzymes
47
Describe eosinophils of the innate immunity
- neutralization of basophil and mast cell products; destruction of some helminths; hypersensitivity rxns
- some phagocytic ability
48
Describe basophils of the innate immunity
- hypersensitivity rxn
- granules contain histamin, heparin, eosinophil chemotactic factor A
- in allergic rxn, bind IgE
- granules release contents in presence of Ag
49
Describe monocytes of the innate immunity
- phagocytosis
- migrate to tissues, become macrophages
- respond to chemotaxins
50
Describe mast cells of the innate immunity
- hypersensitivity rxn
- connective tissue cells
- resemble basophils but larger and more granules
- bind IgE
51
Describe macrophages of the innate immunity
- phagocytosis; eliminations of bacteria, intracellular parasites and tumor cells; secretion of cell mediators; Ag presentation
- activated by contact with microorganisms or cytokines from T cells
52
Describe dendritic cells of innate immunity
- phagocytosis; presentation of Ag to T cells
- initiate adaptive immune response
53
Describe natural killer cells of the innate immunity
- 1st line of defense against tumor cells and cells infected with viruses
- lymph without T or B markers
- no unique surface Ag, but CD16+ and CD56+
- bridge between innate and acquired immunity
- lack specificity
- stimulated by cytokines
- respond early in infection
- provide time for T and B cell to be activated
- <20% of lymph’s
- key cell in antibody- dependent cellular cytotoxicity (ADCC)
54
What are the cells for the acquired immunity?
- T lymphocytes (T cells)
- helper T cells
- cytotoxic T cells
- T regulatory cells
- B lymphocytes (B cells)
- plasma cells
- memory cells
55
Describe T lymphocytes (T Cell) of the acquired immunity
- cell-mediated immunity
- CD3+
- derived from cells in bone marrow
- develop T-cells-specific Ags in thymus
- 60-80% of lymph’s
56
Describe Helper T cells of the acquired immunity
- orchestrate cell-mediated immunity
-activate B cells, cytotoxic cells, and NK cells
- CD4+
- 2/3 peripheral T cells
- Normal CD4+ = 1,000/uL
- in acquired immunodeficiency syndrome (AIDS), <200/uL
57
Describe cytotoxic T cells of the acquired immunity
- suppressor cells inhibits T helper cells
- cytotoxic cells kill other cells
- CD8+
- 1/3 peripheral T cells
- normal CD4/CD8 ratio 2:1 in AIDS, < 0.5:1)
58
Describe T regulatory cells of the acquired immunity
- suppress immune response to self
- CD4+ and CD25+
59
Describe B lymphocytes (B cells) of the acquired immunity
- after antigenic challenge, transform into blasts that give rise to plasma cells and memory cells
- develop in bone marrow
- when mature, have surface Igs( IgM, IgD) tag acts as receptors for Ags
- 10-20% of lymphs
60
Describe plasma cell of the acquired immunity
- Ab production
- in peripheral lymphoid organs
- nondividing
61
Describe memory cells of the acquired immunity
- responses to Ag when encountered again with increased speed and intensity
- in peripheral lymphoid organs
- live months to years
- can be B or T cells
62
Describe subpopulatuons of lymphocytes
- lymphocyte stem cells will either travel to the thymus or Bone marrow
- 60-80% of T cells travel to thymus and will either be CD4+ or CD8+
- the lymphocyte cells in the bone marrow will either become B cells or NK cells
- 10-20% will become B cells and <20% are NK killer cells
63
What are the secondary lymphoid organs?
- spleen
- lymph nodes
- tonsils
- appendix
- cutaneous-associated lymphoid tissue (CALT)
- Mucosal-associated lymphoid tissue (MALT), including Peyer patches in lower ileum
64
Describe isolation of lymphocytes
- density gradient centrifugation with separation media
- layers plasma, mononuclear cells, separation media, RBCs and granulocytes (Neutrophil, eosinophil, and basophil)
65
Describe ID of lymphocytes
- flow cytometry
- fluorescent-labeled monoclonal Abs against specific surface Ags
- each Ab to has different fluorescent tag
- light scattering measured as cells flow through laser beam
- common T cell Ags tested: CD 2, 3, 4, 7, and 8
- B cell Ags = CD 19, 20, 22, surface Ig
66
Describe the basic Immunoglobulin structure
- 2 heavy chains and 2 light chains held together by disulfide (S-S) bonds
- heavy chains determine Ig class (IgG, IgA, IgM, IgD, IgE)
- light chains are either lambda and kappa (only 1 type per Ig)
— free light chains are known are Bence jones protein
- has fab fragment, Fc fragment, constant region, variable region, hinge region, and joint chain
67
Describe FAB fragment
- fragment Ag binding
- consists of 1 light chain and 1/2 heavy chain held together by S-S bonds
- 2 per Ig
- each can bind antigen
68
Describe Fc fragment
- fragment crystallizable
- carboxyl-terminal halves of 2 heavy chains held together by S-S bonds
- site of Ab biological activity such as osponization and complement fixation
69
Describe the constant region of an Ig
- carboxyl-terminals of Heavy and light chains where amino acid sequence is safe for all chains of that type
70
Describe the variable region of Ig
- amino-terminal ends of H and L chains where amino acids sequence varies
- also known as Ag-recognition unit
- responsible for Ig specificity
71
Describe hinge region of Ig
- flexible portion of H chains between first and second constant regions
- allows molecule to bend to that 2 Ag-binding sites can operate independently
72
Describe joining chain of Ig
- glycoprotein that links Ig monomers in IgM and secretory IgA
73
Describe the characteristics of IgG
- monomer
- 150,000 daltons
- 2 gamma
- 70-75% of total Ig
- serum concentration = 800-1,600 mg/dL
- 2 binding sites
- complement fixation = YES
- crosses placenta
- defense against bacteria and viruses
- neutralizes toxins
- opsonin
- passive immunity in newborns
- more efficient at precipitation than agglutination
- main Ig produced during memory (recall) response to Ag
74
Describe the characteristics of IgM-
- pentameter
- 900,000 daltons
- 2 Mu (u) chains
- 10% of total Ig
- serum concentration = 120-150 mg/dL
-10 binding sites
- complement fixation = YES
- does not cross placenta
- neutralizes toxins, opsonins
- first Ig produced in immune response
- only Ig produced in immune response
- most efficient Ig at initiating complement cascade
- more efficient at agglutination than IgG
- destroyed by sulfydryl compounds
75
Describe are the characteristics of IgA
- monomer and dimer
- 160,000-400,000 daltons
- 2 alpha chains
- 10-15% of total Ig
- serum concentration (mg/dL) = 70-350%
- 2-4 binding sites
- complement fixation = NO
- does not cross placenta
- 1st line of defense
- patrols mucosal surfaces
- prevents adherence of bacteria and neutralizes toxins
- In tears, sweat, saliva, respiratory and GI mucosa, breaks milk
76
Describe the characteristics of IgD
- monomer
- 180,000 daltons
- 2 delta chains
- <1% of total Ig
- serum concentration = 1-3
- 2 binding sites
- complement fixation = NO
- does not cross the placenta
- may play role in B-cell maturation
- on surface of B cells
77
Describe the characteristics of IgE
- monomer
- 190,000 daltons
- 2 epsilon chains
- 0.002% of total Ig
- serum concentration = 0.005 mg/dL
- 2 binding sites
- complement fixation = NO
- does not cross placenta
- role in allergic rxn
- binds to basophils and mast cells
- when 2 adjacent molecules on mast cell bind Ag, degranulation of cell with release of histamine and heparin
- type I immediate hypersensitivity rxn
78
Describe complement
- group of >30 proteins involved in phagocytosis and clearance of foreign Ag
- most are inactive Andy me precursors that are converted to active enzymes in precise order (cascade)
- functions = inflammation, opsonization, chemotaxis, cell lysis
- classical pathway = triggered by Ag/Ab rxn. IgM is most efficient activator. Single molecule attached to 2 adjacent Ags can initiate cascade. IgG1, 2 and 3 can activate complement but at least 2 molecules required
- recognition unit = C1 (first to bind)
- activation unit = C4, 2, 3
- membrane attack complex (MAC) = C5, 6, 7, 8, 9 (cell lysis)
- alternative pathway = Ab independent. Activated by bacteria, fungi, viruses, tumor cells, some parasites
- lectin pathway = Ab independent. Initiated by mannose binding lectin (MBL). Nonspecific recognition of sugars on microorganisms. Important defense mechanism in infancy
- C3 presents highest concentration
- increase susceptibility to infection.
- accumulation of immune complexes, which can lead to glomerulonephritis
- Calcium and Magensium required
- inactivation = 56C for 30 minutes
79
What are the types of hypersensitivity reactions?
- type I = anaphylactic
- type II = cytotoxic
- type III = immune complex
- type IV = T-cell dependent
80
Describe type I hypersensitivity reaction
- anaphylactic
- IgE is key reactant
- release of mediators from eosinophils, mast cells, and basophils
- immediate onset of symptoms
- example = anaphylaxis hay fever, asthma, food, allergies
81
Describe type II
- cytotoxic
- IgG, IgM, complement, cellular Ag are key reactants
- cytolysis due to Ab and complement
- immediate onset of symptoms
- examples = transfusion reactions, hemolytic disease of newborn, autoimmune hemolytic anemia
82
Describe type III hypersensitivity reactions
- immune complex
- IgG, IgM, complement, soluble Ag
- deposits of Ag-Ab complexes in tissues
- immediate onset of symptoms
- examples = arthus rxn, serum sickness, SLE, and RA
83
Describe type IV hypersensitivity reactions
- T-cell dependent
- T cells, antigen presenting cells (APCs)
- release of cytokines
- delayed onset of symptoms (sensitization after 1st contact with Ag, symptoms upon reexposure
- examples = contact dermatitis, hypersensitivity pneumonia, Tuberculin skin test
84
Describe direct agglutination
- Naturally occurring Ags on particles
- particles aggluitnate in presence of corresponding Ab
- application = widal test for typhoid fever. Salmonella O and H Ags used to detect Abs in patient serum. test used in developing countries
85
Describe hemagglutination
- Ab-ab reaction that results in clumping of RBCs
- application = ABO typing
86
Describe passive (indirect) agglutination
- soluble Ags bound to particles
- particles agglutinate in presence of corresponding Ab
- application = Ab to group A streptococcus (GAS) or Ab to rotavirus or CMV
87
Describe reverse passive agglutination
- Ab attached to carrier particles
- particles agglutinate in presence of corresponding Ag
- application = kits available for rapid ID of bacteria such as GBS, staph aureus and cyrptococcus neoformans
88
Describe agglutination inhibition -
- competition between particulate Ag (reagent) and soluble Ag (in specimen) for sites on reagent Ab,
- lack of agglutination is POS result
- applications = detection of illicit drugs. Controls are crucial to confirm lack of agglutination
89
Describe hemagglutination inhibiton
- detects Abs to certain viruses that agglutinate RBCs
- in presence of Ab, virus in neutralized and hemagglutination doesn’t occur
- application = rubella and other viruses. Controls are crucial to confirm lack of hemagglutination
90
Describe precipitation
- soluble Ag combines with soluble Ab to produce visible complexes
- less sensitive than agglutination
91
Describe Ouchterlony double diffusion
- Ags and AB’s diffuse from wells in gel and form precipitin lines where they meet
- application = fungal Ag
92
Describe Radial immunodiffusion (RID)
- Ag diffuse out of well of gel containing Ab
- precipitin ring forms
- diameter proportional to concentration of Ag
- application = largely replaced by more sensitive methods such as nephlometry and ELISA
93
Describe immunofixation electrophoresis (IFE)
- proteins separated by electrophoresis
- antiserum placed directly on gel
- Ag-Ab complexes precipitate
- application = ID of Igs in monoclonal gammopathies, Bence jones proteins. Also helpful for detection of antigen present in serum, urine or CSF at low concentrations
94
Describe nephlometry
- light scattering by Ag-Ab complexes.
- amount of light scattered is proportional to concentration
- applications = Igs, complement, C-reactive protein (CRP), haptoglobin, and ceruplasmin
95
What is it ligand?
- substance being measured in immunoassay
- can be Ag or Ab
96
What is competitive immunoassay?
- patient Ag and labeled reagent Ag compete for binding sites on reagent Ab
97
What is noncompetitive immunoassay?
- doesn’t involve competition for binding sites
- more sensitive tan competitive assays
98
What is heterogenous immunoassays?
- separation step to remove free from bound analyte
- more sensitive than homogenous assays
99
What is homogenous immunoassay?
- Doesn’t require separation step
- easier to automate
100
Describe EIA
- any immunoassay that uses an enzyme as label
- enzyme labeled ligand and unlabeled patient ligand compete for binding sites on Ab attached to solid phase
- free labeled ligand remove by washing. Subtrtate added.
- Color inversely proportional to concentration of ligand in specimen
- used to measure small relatively pure
- heterogenous
- competitive
- direct
101
Describe direct EIA
- 1st type of of EIA developed.
- competitive
- Enzyme-labeled reagent is part of initial Ag-Ab rxn
- all reactants added at same time
- 1 incubation and 1 wash
102
Describe indirect EIA
- noncompetitive EIA
- enzyme labeled reagent isn’t involved in initial Ag-Ab rxn
- 2 incubation and 2 washes
- more sensitive than direct assays
- also known as ELISA
103
Describe solid phase immunoassay
- reagent Ag or Ab bound to support medium
104
Describe ELISA
- heterogenous
- noncompetitive
- indirect
- Ag attached to solid phase. Ab in specimen attaches. Unbound Ab removed by washing. Enzyme-labeled antiglobulin added. attaches to Ab on solid phase, Substrate added
- color directly proportional to Ab concentration
- more sensitive than competitive EIA
- one of most common immunoassays
- used to detect Abs to viruses (HIV), HAV, HCV, EBV)
105
Describe sandwich ELISA or capture assay
- heterogenous
- noncompetitive
- indirect
- Ab attached to solid phase
- Ag in specimen attaches
- Enzyme-labeled Ab added, attaches to different determinant
- enzymatic activity is directly proportional to amount of Ag in sample
- Ags must have multiple determinants
- used to measure hormones, proteins, and detect tumor markers, viruses, parasites, fungi
- high concentration of Ag can cause hook effect
- too much Ag for binding sites so undiluted sample has lower absorbance than dilutions
106
Describe Rapid ELISA (lateral flow)
- membrane based
- reagent Ag or Ab bound to membrane in single use cassette. Sample added. Presence of Ag-Ab complex indicated by colored rxn
- may have built in control
- usually qualitative and designed primarily for POC or at home testing
- examples = pregnancy test, cardiac troponin, and SARS-CoV-2
107
Describe EMIT
- “enzyme - multiplied immunoassay technique
- homogenous
- Ag specimen and enzyme-labeled Ag compete for binding site on reagent Ab
- when enzyme-labeled Ag binds, enzyme activity inhibited
- enzyme activity is directly proportional to concentration of Ag in specimen
- used for determination of low molecular weight analytes not readily measured by other methods
- automate
108
Describe directly antibody (DFA) staining
- specimen on glass slide overlaid with fluorescein-labeled Ab
- if corresponding Ag present, labeled Ab binds
- fluorescence observed with fluorescent microscope
- detects Ags.
- fluorescent isothiocynate or rhodamine B isothiocyanate
- examples of analytes = bacterial, viral AG
109
Describe Indirect fluorescent antibody (IFA) staining
- reagent Ag on glass slide overlaid with patient serum
- if corresponding Ab present in serum, attaches to Ag
- when fluorescein-labeled Antihuman globulin added, attaches to Ab
- fluorescence observed with fluorescent microscope
- “sandwich technique”
- Detects Abs in serum
- antinuclear antibody (ANA), fluorescent treponemal antibody (FTA)
110
Describe fluorescence polarization immunoassay (FPIA)
- labeled Ag competes with Ag in specimen for sites n reagent Ab
- free labeled Ag rotates rapidly, emits little polarized light
- bound labeled Ag rotates more slowly, emits more polarized light
- amount of polarized light is inversely proportional to concentration of Ag in specimen
- competitive
- homogenous
- automated
- therapeutic drugs, hormones
111
What are characteristics of EIA
- labels = alkaline phosphatase, horseradish peroxide, alpha-D-galactosidase, G6PD
- detection = enzymes react with substrate to produce color change
- types of assays available = mostly noncompetitive now. Heterogenous and homogenous
- advantages = sensitivity. Specificity. No health hazard or disposal problems. Reagents with long shelf life. Can be automated
- disadvantages = natural inhibitors in some specimens. Cross reactivity of some substances
112
What are the characteristics of of FIA?
- labels = fluorescein, rhodamine
- detection = fluorochromes absorb energy from light source, concerto long wavelength (lower energy)
- types of assay available = usually competitive, heterogenous and homogenous
- advantages = sensitivity. Specificity. No health hazard or disposal problems. Reagents with long shelf life. automated
- disadvantages = autofluorescence from organic substances in serum. Non specific binding to substances in serum. Expensive, dedicated instrumentation
113
What are the characteristics of Chemiluminescent immunoassay (CIA)?
- labels = luminol, acridinium esters, ruthenium derivatives, nitrophenyl oxalates
- detection = chemiluminescent molecules produce light from chemical reaction
- types of assays available = competitive and noncompetitive. Heterogenous and homogenous
- advantages = sensitivity. Specificity. No health hazard or disposal problems. Reagents with long shelf life. automated
- disadvantages = quenching of light emission by some biological materials
114
What are the nontreponemal tests for syphilis?
- VDRL
- RPR
115
Describe VDRL
- method = flocculation
- detect Reagin (Ab against cardiolipin that is in serum of patients with syphilis)
- Ag = cardiolipin
- POS rxn = microscopic clumps
- specimens = inactivated serum, CSF
- reactivity during disease = may be NEG in primary stage. Titers usually peak during secondary or early late stages. More rapid decline with treatment. Becomes nonreactive in 1-2 years following successful treatment
- false POS = biologic false POS with infectious mononucleosis (IM), infectious hepatitis, malaria, leprosy, lupus erythrematous, RA, advanced age, pregnancy.
- reactive in other treponemal infections such as yaws and piñta
- screening test
- reactives should be confirmed by treponemal test.
- replaced by RPR for serum.
- Still performed on CSF for diagnosis (Dx) of neurosyphilis
116
Describe RPR
- method = flocculation
- detect = reagin
- Ag = cardiolipin with charcoal
- POS rxn = macroscopic agglutination
- specimen = serum (inactivated not required), plasma
- reactivity during disease = may be NEG in primary stage. Titers usually peak during secondary or early late stages. More rapid decline with treatment. Becomes nonreactive in 1-2 years following successful treatment
- false POS = biologic false POS with infectious mononucleosis (IM), infectious hepatitis, malaria, leprosy, lupus erythrematous, RA, advanced age, pregnancy.
- used for screening and treatment monitoring
- screening test
- reactives should be be confirmed by treponemal test
117
What are the treponemal tests for syphilis?
- FTA-ABS
- TP-PA
- ELISA
118
Describe the Fluorescent treponemal antibody absorption (FTA-ABS)
- detects Ab to T. Pallidum
- reagents = sorbent (nonpathogenic treponemes-Reiter strain), slides with Nichols strain of T. Pallidum, fluorescein-labeled Antihuman globulin
- POS rxn = fluorescence
- specimens = serum, CSF
- reactivity during disease = usually POS before nontreponemal tests. Some false NEGs in primary syphilis. Usually POS for life
- false POS = fewer than nontreponemal tests. Reactive with other treponemal diseases (yaws, pinta)
- absorbent removes nonspecific Ab
- used to confirm reactive nontreponemal test
- not good for treatment monitoring
119
Describe treponema pallidum particle agglutination (TP-PA)
- detects Ab to T. Pallidum
- regents = colored gelatin particles coated with treponemal Ag
- POS rxn = agglutination of sensitized gel particles. Smooth mat over surface of well
- reactivity during disease = not as sensitive in primary syphilis as FTA. Sensitivity close to 100% in secondary syphilis
- usually POS in late stages
- specimens = serum
- false POS = fewer than nontreponemal tests
- used to confirm reactive nontreponemal tests
- a smooth mat will form in the base of the well of a microtiter plate,indicating a POS results
- not good for treatment monitoring
120
Describe Antibody capture enzyme-linked immunosorbent assay (ELISA)
- detects Ab to T. Pallidum
- reagents = enzyme-labeled treponemal Ag.
- POS rxn = color development following addition of substrate
- specimens = serum
- reactivity during disease = high sensitivity
- false POS = similar to other treponemal tests
- can be automated
- used for screening and for confirmation of reactive nontreponemal tests.
- when used for screening, reactives should be confirmed with nontreponemal test (reverse sequence screening), followed by TP-PA or FTA-ABS if nontreponemal test is nonreactive
- IgM capture assay for diagnosis for congenital syphilis
121
What does RPR reactive and FTA reactive indicate?
- POS for syphilis
122
What does RPR reactive and FTA nonreactive indicate?
- NEG for syphilis
123
What does ELISA reactive ad RPR reactive indicate?
- POS for syphilis
124
What does ELISA reactive, RPR nonreactive, and FTA-ABS reactive indicate?
- late, latent, or previous syphilis
125
Describe Anti-streptolysin O (ASO)
- diagnosis = sequelae of GAS infection: rheumatic fever, poststreptococcal glomerulonephritis
- common methods = nephelometry
- use recombinant streptolysin Ag.
- if Ab present, Ag-Ab complexes form and increase light scatter
126
Describe DNase-B test
- diagnosis = sequelae of GAS infection; rheumatic fever, glomerulonephritis following skin infection
- common methods = EIA, nephelometry
- highly specific
- May be POS when ASO is NEG
127
Describe streptozyme test
- diagnosis = sequelae of GAS infection
- common method = slide agglutination
- use BAP coated with several streptococcal Ags
- more false POS and NEG
- should be used in conjunction with ASO and anti-DNase
- serial titers should be performed
128
Describe heliobacter pylori Ab test
- diagnosis = gastric and duodenal ulcers caused by H. Pylori
- common method = method of choice: ELISA. Rapid tests, PCR available
- most tests detect IgG
- 25% decrease in titer = successful treatment
- Abs remain for years
- POS rapid tests should be confirmed by ELISA
129
Describe Mycoplasma pneumonia Ab test
- diagnosis = primary atypical pneumonia (PAP)
- common methods =
— most common = EIA. Also agglutination, IFA.
— molecular methods are also available
- most tests for IgM and IgG Abs
130
Describe Rickettsial Ab test
- diagnosis = typhus, RMSF, other rickettsial infections
- common methods =
—gold standard = IFA, micro-IF
— PCR available
131
What are the serological tests for IM?
- Heterophile antibodies
- EBV-specific antibodies
132
Describe heterophile antibody test
- non specific Abs that agglutinate horse, sheep, and bovine RBCs
- heterophile Abs are Abs that react with similar Ags from different species
- occurrence = 90% of patients develop in 1st month of illness
- can persist for 1 year NEG in 10% of adults and up to 50% of children with IM
- if symptomatic and heterophile NEG, test for EBV-specified Abs
- tests = rapid latex agglutination, solid phase immunoassays. Ag is purified bovine RBC extract. Screening tests
133
Describe EBV-specifid antibodies
- specific Abs against EBV Ags present in different phases of infection — early: early Ag (EIA), CIA
- IFA is gold standard but time consuming and harder to interpret
- molecular tests can be used for immunocompromised patients who don’t produce Abs
134
What tests detect HAV?
- total anti-HAV
- IgM anti-HAV
- HAV RNA
135
Describe total anti-HAV test
- past infection and immunity
136
Describe IGM anti-HAV test
- acute infection
137
Describe HAV RNA test
- current infection
- used to detect HAV in food and water
138
What are the tests that detect HBV?
- Hepatitis B surface antigen (HBsAg)
- hepatitis B e antigen (HBeAg)
- total anti-hepatitis B core (HBc)
- IgM anti-HBc
- anti- HBe
- anti-HBs
- hepatitis B virus (HBV) DNA
139
Describe Hepatitis B surface antigen (HBsAg)
- acute or chronic infection, infectivity
- 1st serological marker to appear, indicating early acute infection
- used to screen donor blood
- POS should be confirmed by repeat testing and another assay such as hepatitis B deoxyribonucleic acid (DNA) PCR
140
Describe Hepatitis B e antigen (HBeAg)
- acute or chronic infection
- indicates high degree of infectivity
141
Describe total anti-hepatitis B core (HBc)
- current or past infection or carrier
- predominately IgG, which persists for life
142
Describe IgM anti-HBc
- current or recent infection
- 1st Ab to appear. Useful for detecting HBV infection when HBsAg is no longer detectable (“window period”)
- used to screen blood donors
143
Describe anti-HBe test
- recovery, reduced infectivity
144
Describe Anti-HBs test
- recovery and immunity
- Ab that develops following immunization
145
Describe Hepatitis B virus (HBV) DNA test
- current infection
- decectable 21 days before HBsAg
- used to monitor viral load and therapy
146
What are the tests that detect HCV?
- Anti-HCVh
- HCV ribonucleic acid (RNA)
147
Describe anti-HCVh test
- acute, chronic, or previous infection
- POS should be confirmed by recombinant immunoblot assay (RIBA) or molecular method
148
Describe HCV ribonuclieic acid (RNA) test
- current infection
- used for viral load testing, blood/organ donor screening
- HCV genotyping to determine optimal treatment
149
What are the test that detect HDV?
- IgM anti-HDV
- IgG anti-HDV
- HDV RNA
150
Describe IgM anti-HDV
- acute or chronic infection
- HDV is a defective virus that can only occur in presence of HBV
151
Describe the IgG anti-HDV test
- recovery or chronic infection
152
Describe HDV RNA test
- current infection
- marker of active viral replication
- used to monitor therpay
153
What tests are used to detect HEV?
- test are currently not approved by food and drug administration (FDA) for use in US
154
What is the serological profile of acute HAV?
- IgM anti-HAV +
155
What is the serological profile of recovery from HAV?
- total anti-HAV +
156
What is the serological profile of acute HBV?
- HBsAg +
- total anti-HBc +
- IgM anti-HBc +
- Anti-HBs -
157
What is the serological profile of recovery from HBV?
- HBsAg -
- Total anti-HBc +
- anti-HBs +
158
What is the serological profile of chronic hepatitis B/carrier?
- HBsAg +
- total anti-HBc +
- IgM anti-HBc -
- Anti-HBs -
159
What is the serological profile for hepatitis B immunization?
- HBsAg -
- Anti-HBc -
- Anti-HBs +
160
Describe appearance of HIV markers
- Viral RNA = detectable within days of infection
- p24 Ag = core court for nucleic acids. detectable in 2-3 weeks. becomes undetectable as Abs develop, then detectable again in late stages as immune system fails and virus replicates
- IgM Ab = usually detectable in 2-8 weeks. Transient. Peaks in about 1-2 weeks, undetectable about 1-2 weeks later
- IgG Ab = detectable shortly after IgM. Increased in titer over several months. Long lasting
161
Describe the ELISA/CLIA testing for HIV
- 1st gen = IgG Ab to HIV-1 (6-12 weeks)
- 2nd gen = IgG Ab to HIV-1/2 (6-12 weeks)
- 3rd gen = IgG and IgM Ab to HIV-1/2 (3-4 weeks)
- 4th gen = IgG and IgM Ab to HIV-1/2 and P24 Ag (2 weeks)
- 5th gen = IgG and IgM Ab to HIV but differentiates HIV-1 from HIV-2. Also detects p24 Ag
* P24 Ag without HIV Ab = acute infection. P24 Ag and HIV Ab = established infection. Currently not approved for screening blood donors. 5th generation tests not widely implemented yet
162
Describe rapid tests of HIV screening
- detects IgG and IgM Ab to HIV
- window period = 4-12 weeks
- immunochromatographic assays
- can be performed on whole blood, serum, oral fluid
163
Describe NAAT of HIV screening
- detects = HIV RNA
- window period = 5 days
- not cost effective for screening in most settings but used if initial ELISA/CLIA combo test is POS while HIV-1 and HIV-2 differentiation immunoassays are NEG
- used for screening blood donors and sometimes infants but PCR to HIV-1 preferred for infants)
- Ab tests aren’t reliable in children <18 months of age.
- Abs from infected mother can be present even if child isn’t infected
164
What can cause false POS with HIV-antibody ELISA testing?
- heat inactivation of serum
- repeated freezing/thawing of serum
- autoantibodies
- multiple pregnancies
- liver disease
- administration of Ig
- administration of certain vaccines
- some malignancies
165
What can cause false NEG with HIV-antibody ELISA testing?
- blood drawn before seroconversion (window period)
- hypogammaglobulinemia
- immunosuppressive therapy
- strain of HIV not detected by assay
- technical errors
166
Describe CDA T-cell count test to stage and monitor HIV
- HIV infects CDa cells
- number declines as disease progresses.
- <200/uL defines stage 3 infection according to CDC
- also used to monitor therapy
- perform every 3-6 months
- flow cytometry is gold standard
167
Describe HIV-1 viral load assays: PCR; branched chain DNA assay (bDNA) test to stage and monitor HIV
- quantitative methods to determine plasma HIV RNA
- used to predict disease progression, determine when to start antiretroviral therapy
- qPCR most frequently performed, bDNA assays are used in labs with high testing volumes
- test 2-8 weeks after start of therapy and then every 3-4 months
- same 3-4 months
- same assay should be used in order to to assess changes
168
Describe Western Blot (WB) for HIV testing
- traditional confirmatory test but not as sensitive as 4th gen EIA or NAAT.
- interpretation is controversial but most labs report POS if at least 2 of the following 3 bands are present: p24, gp41, gp120/160
- NAAT required following NEG or indeterminant results.
- time-consuming, difficult to interpret
169
What are the screening tests for SLE?
- ANA
- C3 EIA
- urinalysis
170
Describe antinuclear antibody (ANA) for SLE
- indirect immunofluorescence (IIF). Substrate is human epithelial cell line (Hep-2)
- high sensitivity (95-100%) but low specificity
- dilutions tested to eliminate low titer reaction in normal population
- cutoff dilution to report POS usually >=1:80
- endpoint titer may be reported
- generally higher in SLE
- detects auto Abs-to nuclear-Ag
- staining patterns reported but not considered as significant as in part
- labor intensive
- subjective
- still considered gold standard by rheumatologists
171
Describe Complement (C3) EIA test for SLE
- method = EIA
- C3 is turned over rapidly in SLE patients, especially during flare-ups will see a decrease in serum C3 levels
172
Describe urinalysis test for SLE
- method = dipstick
- looking for RBCs and protein to screen for kidney damage caused by Ag-auto Ab complexes
173
Describe Anti-dsDNA test for specific ANA
- low sensitivity for SLE
- high specificity for SLE. Uncommon in other diseases or normal individuals
- titer correlates with disease activity
- peripheral or homogenous fluorescent pattern with IIF
- other methods: EIA
174
Describe anti-sm tests for specific ANA?
- low sensitivity for SLE
- high specificity for SLE. Uncommon in other diseases or normal individuals
- coarsely speckled patterns with IIF
- the methods = EIA, immunodiffusion
175
Describe antihistone, anti-DN, anti-SS-A/Ro, anti-SS-B/La, anti-nRNP test for specific ANA
- low sensitivity for SLE
- low specificity for SLE
- generally not useful for diagnosis of SLE
- used to diagnosis other connective tissue diseases (Sjorgren Syndrome)
- methods = IIF, EIA, immunodiffusion
176
Describe extra table nuclear antigen (ENA) for specific ANA
- low sensitivity for SLE
- anti-sm is specfic for SLE
- immunodiffusion (Ouchterlony double diffusion) test panel that typically yields tests for Abs to Smith (Sm), SS-A/Ro, SS-B/La, ribonucleoprotein (RNP).
- precipitin lines of ID/non-ID
- new method = multiplex bead assay
- immunoassay using specific Ag-coated beads and flow cytometry to detect multiple ANAs simutaneously
177
What are serological tests for RA?
- Rheumatoid factor (RF)
- Anti-cyclic citrullinated peptide antibody (anti-CCP)
178
Describe Rheumatoid factor tests for RA?
- common methods = agglutination, ELISA, nephelometry
- autoantibody (usually IgM) against IgG
- POS in 70-80% of patients with RA
- not specific for RA
- present with other autoimmune diseases infections, and in some normal individuals
- agglutination tests not detect IgM RF
- ELISA and nephelometry can also detect IgA and IgG classes of RF
- automated methods more common
179
Describe Anti-cyclic citrullinated peptide antibody (Anti-CCP)
- common method = ELISA
- more specific RA than RF
180
Describe Systemic Lupus Erythematosus (SLE)
- target cells and tissues = multiple; kidneys, joints, skin, brain, heart, lungs
- autoantibodies to double-stranded DNA and other nuclear components most common
- also see decreased serum C3, increased CRP and ESR
- RBCs/protein in urine in common
- abnormal CBC values may be observed: decreased hgb andht, low WBC and platelet count
181
Describe rheumatoid Arthiritis (RA)
- target cells and tissue = joints, bone, and other connective tissue
- Anti-CCP most diagnostic, Rheumtoid factor, POS ANA titer
- elevated CRP, ESR and decreased C3
- used to monitor treatment
182
Describe Wegener Granulomatosis
- also known as granulomatosis w/ polyangitis
- target cell and tissues = upper respiratory system, lungs and blood vessels
- POS antineutrophil cytoplasmic antibody (ANCA)
- glomerulonephritis common
- rare but severe due to chronic activation of neutrophils, T cells and B Cells
183
Describe Scleroderma
- target cell and tissues = skin and blood vessels; fibrosis can occur in vessels of most organs
- Scl-70 autoantibodies, speckled/Nucleolar ANA pattern
- Raynaud’s phenomenon common
184
Describe Sjorgren syndrome
- target cells and tissues = lacrimal and salivary glands
- autoantibodies toward RNA completed with cellular proteins (SS-A/Ro and SS-B/La)
- can occur with SLE or RA
185
Describe Graves’ disease
- thyroid glands (commonly thyroid stimulating hormone (TSH) receptors
- Decreased TSH, increased T4, TSH receptor autoantibodies most common
- may see thyroglobulin and thyroperoxidase (TPO) autoantibodies
- TSH receptor autoantibodies
- diagnostic in 99% of patients
186
Describe Hashimoto’s thyroiditis
- target cells and tissues = thyroid gland (epithelial cells)
- increased TSH, decreased T4
- TPO and thyroglobulin autoantibodies in most patients
- microsomal autoantibodies diagnostic
187
Describe Severe combined immunodeficiency (SCID)
- immune component deficient = adaptive arm, IL-2 receptor mutation
- decreased and/or nonfunctional T cells and B cells
- genetic tests will confirm
188
Describe Wiskott-Aldrich disease
- immune component deficient = cellular arm of immune system (mainly T cells but then B cells can’t form antibodies)
- decreased platelet # and size, decreased IgM (to ABO antigens, can be diagnostic), and elevated serum alpha Fetoprotein (AFP)
- patients often have severe eczema
189
Describe DiGeorge anomaly
- immune component deficient = chromosome 22 mutation, defective thymus, or lack of thymus
- partial to complete lack of T cells
-
190
Describe Ataxia Telangiectasia (AT)
- immune component deficient = chromosomal breaks occur, inhibiting VDJ rearrangement for T and B cells
- decreased circulating T cells and levels of IgA, IgE, and IgG2
- increased serum AFP
- physical manifestations include ataxia and capillary swelling/red blotches
191
Describe X-linked Burton Agammaglobulinemia
- tyrosine Kinase deficiency
- antibody immunodeficiency, lack CD19+ cells and all subsequent cells (plasma and memory cells)
- lack of CD19+ cells via flow cytometry, decreased or lack of IgA, IgE, IgG, IgM
192
Describe selective IgA deficiency
- immune component deficient = IgA only is deficient
- lack of serum IgA
- recurrent respiratory and GI infections
193
Describe Chediak Higashi syndrome
- immune component deficient = NK cells/neutrophils microbiocidal function
- differential shows WBC with enlarges granules, increased acute phase proteins and cytokines
194
Describe Chronic granulomatous disease
- immune component deficient neutrophil microbiocidal function
- DHR/flow cytometry (decreased fluorescence
- increased susceptibility to pyogenic infections
195
Describe Leukocyte Adhesion deficiency
- CD18 on phagocytic cells
- decreased CD18 on dendritic cells (measured by flow cytometry)
- delayed wound healing and chronic skin, intestinal, and respiratory tract infections
196
Describe intepretation of serological tests
- >= fold increase in titer from acute to convalescent specimen draw 10-14 days later is diagnostic
- IgM Ab is sign of recent infection
- IgG Ab is sig of immunity
- IgG Ab in newborn is maternal Ab
