Component-resolved in vitro diagnosis in carrot allergy: does the use of recombinant carrot allergens improve the reliability of the diagnostic procedure?
BACKGROUND
In Europe, pollen-related food allergy is the most frequent form of food allergy in adults. Reliability of current diagnostic procedures, however, is poor and therapeutic options are not available.


OBJECTIVE
In the present study, we created a panel of recombinant allergens from carrot and evaluated its potential in component-resolved in vitro diagnosis of carrot allergy.


METHODS
Recombinant (r) Dau c 1.0104, Dau c 1.0201 and Dau c 4 were cloned by a polymerase chain reaction strategy, expressed in Escherichia coli and purified. Carrot lipid transfer protein (LTP) was expressed in the yeast Pichia pastoris. Sera from 40 carrot-allergic patients were investigated. Twenty-one birch pollen-allergic subjects with negative open provocation to carrot and 20 non-allergic subjects were included as controls. IgE binding to recombinant allergens as well as to cross-reactive carbohydrate determinants (CCD) was measured by ELISA. Cross-reactivity between Dau c 1 isoforms and Bet v 1 was assayed by ELISA inhibition. Biological activity of the recombinant carrot allergens was assessed by histamine release assay and peripheral blood mononuclear cells stimulation.


RESULTS
Ninety-eight percent of the carrot-allergic patients were positive to at least one recombinant allergen; 98% reacted to rDau c 1.0104, 65% to rDau c 1.0201, 38% to rDau c 4 and 20% had IgE against CCD. Specificity using the recombinant allergens was high when compared with non-allergic controls, but low compared with birch-sensitized subjects without carrot allergy. Sensitization to Dau c 1.0201, however, proved to be highly specific for clinically relevant sensitization. Inhibition assays indicated the absence of LTP in carrot root extract, and epitope diversity between Dau c 1.0104, Dau c 1.0201 and Bet v 1.


CONCLUSIONS
Our panel of recombinant allergens from carrot can provide a standardized tool for in vitro diagnosis of carrot allergy, and for epitope studies.
Ballmer-Weber
B K
BK
Allergy Unit, Department of Dermatology, University Hospital, Zürich, Switzerland. barbara.ballmer@usz.ch
Wangorsch
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A
Bohle
B
B
Kaul
S
S
Kündig
T
T
Fötisch
K
K
van Ree
R
R
Vieths
S
S
D000328:Adult; D000485:Allergens / Q000276:immunology*; D052179:Antigens, Plant; D029662:Betula / Q000276:immunology; D002352:Carrier Proteins / Q000032:analysis; D003429:Cross Reactions / Q000276:immunology; D018552:Daucus carota / Q000737:chemistry / Q000276:immunology*; D000939:Epitopes / Q000276:immunology; D005512:Food Hypersensitivity / Q000175:diagnosis* / Q000276:immunology; D006636:Histamine Release / Q000276:immunology; D006801:Humans; D007073:Immunoglobulin E / Q000276:immunology; D007963:Leukocytes, Mononuclear / Q000276:immunology; D010940:Plant Proteins / Q000032:analysis / Q000276:immunology*; D011058:Pollen / Q000276:immunology; D020033:Protein Isoforms / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology
D000485:Allergens; D052179:Antigens, Plant; D002352:Carrier Proteins; C119925:Dau c 1 protein, Daucus carota; C507284:Dau c 4 protein, Daucus carota; D000939:Epitopes; D010940:Plant Proteins; D020033:Protein Isoforms; D011994:Recombinant Proteins; C511920:lipid transfer proteins, plant; C061312:Bet v 1 allergen, Betula; D007073:Immunoglobulin E