April 23, 2013
ISAC and Component-resolved Diagnosis
Douwe de Boer, Chris Nieuwhof, Judith Bons en Paul Menheere
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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Allergy: clinical relevance  “in vitro” testing
intra-allergenic heterogeneity
allergen = mixture of allergen components
Component-resolved
Diagnosis (CRD)
B
A
E
C
Ebo et al., University of Antwerp
D
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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RAST
classical ImmunoCAP assay
RAST1
40 µL
Single-analyte assay: single analysis of specific IgE directed against
single allergens, single allergen components or mixtures of allergens
ISAC (Immuno Solid-phase Allergy Chip )
ImmunoCAP ISAC (16 spots per mm2)
Multiplex assay: multiple analysis of specific
IgE directed against allergen components in one
single run
ISAC112
30 µL
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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Storage protein
Storage protein
intra-allergenic heterogeneity
Storage protein
Storage protein
Profilin
Storage protein
Storage protein
Pathogenesis Related-10 protein
non-specific Lipid Transfer Protein
Component-resolved
Diagnosis (CRD)
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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Complications Detecting specific IgEs - 1
intra-allergenic heterogeneity
inter-allergenic homogeneity
non-specific Lipid Transfer Proteins
Ara h 9
Pathogenesis Related-10 proteins
Ara h 8
Profilins
Ara h 5
Pan-allergens
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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Complications Detecting specific IgEs - 2
Component-resolved
Diagnosis (CRD)
intra-allergenic heterogeneity
inter-allergenic homogeneity
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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Pan-allergens
– risks
and severity
initially
CCD was considered
to be important
to explain cross-reactivity
Cross-reactive
Carbohydrate
Determinants
specific
glycan
epitope
mainly “plant” derived pan-allergens
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
Profilins
Cytoskeleton
proteins
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Highly conserved proteins
True pan-allergen
(high degree of cross-reactivity)
rMer a 1
rAra h 5
rBet v 2
rPhl p 12
rHev b 8
rPru p 4
rMal d 4
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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Pathogenesis Related-10 Protein (PR-10)
Large family and conserved proteins
Heat and acid labile proteins
rGly m 4
rAra h 8
Unknown
function
rBet v 1
rCor a 1.01
rCor a 1.04
rApi g 1
rDau c 1
rPru p 1
rPyr c 1
rMal d 1
rPru ar 1
rPru av 1
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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non-specific Lipid Transfer Protein (LTP)
Relatively conserved proteins
Heat and acid stable protein
Shuttle
protein of
lipids
rZea m 4
rCor a 8
rPru av 3
rPru p 3
rAra h 9
rMal d 3
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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Storage Protein
Proteins for storage of energy mainly in seeds
Relatively less conserved proteins
pseudo-Pan-allergen (low degree of cross-reactivity)
“Voorraad
of opslag
eiwitten”
rSes i 3
rBer e 1
rCor a 9
rAra h 1
rAra h 2
rAra h 3
rAra h 6
nGly m 5
nGly m 6
CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
Pan-allergen sensitization in Europe
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CRD and ISAC | De Boer, Nieuwhof, Bons and Menheere | April 23, 2013
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ISAC and Component-resolved Diagnosis
Conclusions and remarks
 Component-resolved Diagnosis can give insights in
sensibilisations for separate allergen components
intra-allergenic heterogeneity
 Besides separate allergen components groups of panallergens can be distinguished
inter-allergenic homogeneity
 Components and pan-allergens are being associated with risks
 ISAC is a multiplex assay (an extensive number of components)
 RAST is a single analyte assay (limited number of components)