Background: Ovomucoid (Gal d 1) has been demonstrated to be the most important allergen in IgE-mediated egg allergy. Peptide microarray analysis is a novel method that can provide useful information on the nature of specific allergens. Methods: A peptide microarray immunoassay was performed using a 15- and 20-amino acid (aa) library of overlapping peptides (3-offset) of the primary sequence of ovomucoid. Sera from 50 patients with IgE-mediated egg allergy and reactivity to ovomucoid, with more than 1 year of follow-up, and sera from 10 controls were tested. Peptides were considered major epitopes when the average weighted Z-score was greater than 3 and recognized by at least 20% of the patient’s sera. Specific IgE epitopes were established on the basis of the IgE/IgG4 Z-score ratio. Results: The IgE and IgG4 recognition pattern was similar in both sets of peptides, but the signal intensity was generally higher in the 20-aa set. Thirty-four percent of the patients did not recognize any IgE sequential peptide and 20% of the patients recognized more than 10 sequential peptides. We identified 3 major IgE B-cell epitopes in domains I and II of ovomucoid. IgE/IgG4 ratio analysis showed that peptides 1–2 (aa 4–20) and peptides 29–31 (aa 91–104) were specific IgE epitopes. Conclusion: By using peptide microarray immunoassay in egg-allergic patients, we established that 34% of the patients do not have any linear epitope recognized by IgE. Further studies are needed to determine the clinical relevance of this finding.

Keywords:
Egg allergens, Epitope mapping, Ovomucoid, Peptide microarrays
1.
Benhamou AH, Caubet JC, Eigenmann PA, Nowak-Wegrzyn A, Marcos CP, Reche M, Urisu A: State of the art and new horizons in the diagnosis and management of egg allergy. Allergy 2010;65:283–289.
2.
Burks AW, Tang M, Sicherer S, Muraro A, Eigenmann PA, Ebisawa M, Fiocchi A, Chiang W, Beyer K, Wood R, Hourihane J, Jones SM, Lack G, Sampson HA: Icon: food allergy. J Allergy Clin Immunol 2012;129:906–920.
3.
Boyano-Martinez T, Garcia-Ara C, Diaz-Pena JM, Martin-Esteban M: Prediction of tolerance on the basis of quantification of egg white-specific IgE antibodies in children with egg allergy. J Allergy Clin Immunol 2002;110:304–309.
4.
Sampson HA, Scanlon SM: Natural history of food hypersensitivity in children with atopic dermatitis. J Pediatr 1989;115:23–27.
5.
Savage JH, Matsui EC, Skripak JM, Wood RA: The natural history of egg allergy. J Allergy Clin Immunol 2007;120:1413–1417.
6.
Cooke SK, Sampson HA: Allergenic properties of ovomucoid in man. J Immunol 1997;159:2026–2032.
7.
Järvinen KM, Beyer K, Vila L, Bardina L, Mishoe M, Sampson HA: Specificity of IgE antibodies to sequential epitopes of hen’s egg ovomucoid as a marker for persistence of egg allergy. Allergy 2007;62:758–765.
8.
Takagi K, Teshima R, Okunuki H, Itoh S, Kawasaki N, Kawanishi T, Hayakawa T, Kohno Y, Urisu A, Sawada J: Kinetic analysis of pepsin digestion of chicken egg white ovomucoid and allergenic potential of pepsin fragments. Int Arch Allergy Immunol 2005;136:23–32.
9.
Urisu A, Yamada K, Tokuda R, Ando H, Wada E, Kondo Y, Morita Y: Clinical significance of IgE-binding activity to enzymatic digests of ovomucoid in the diagnosis and the prediction of the outgrowing of egg white hypersensitivity. Int Arch Allergy Immunol 1999;120:192–198.
10.
Dieguez MC, Cerecedo I, Muriel A, Zamora J, Abraira V, Camacho E, Anton M, de la Hoz B: Utility of diagnostic tests in the follow-up of egg-allergic children. Clin Exp Allergy 2009;39:1575–1584.
11.
Urisu A, Ando H, Morita Y, Wada E, Yasaki T, Yamada K, Komada K, Torii S, Goto M, Wakamatsu T: Allergenic activity of heated and ovomucoid-depleted egg white. J Allergy Clin Immunol 1997;100:171–176.
12.
Kato I, Schrode J, Kohr WJ, Laskowski M Jr: Chicken ovomucoid: Determination of its amino acid sequence, determination of the trypsin reactive site, and preparation of all three of its domains. Biochemistry 1987;26:193–201.
13.
Matsuda T, Watanabe K, Nakamura R: Immunochemical and physical properties of peptic-digested ovomucoid. J Agric Food Chem 1983;31:942–946.
14.
Pomes A: Relevant B cell epitopes in allergic disease. Int Arch Allergy Immunol 2010;152:1–11.
15.
Lin J, Bardina L, Shreffler WG: Microarrayed allergen molecules for diagnostics of allergy. Methods Mol Biol 2009;524:259–272.
16.
Lin J, Bardina L, Shreffler WG, Andreae DA, Ge Y, Wang J, Bruni FM, Fu Z, Han Y, Sampson HA: Development of a novel peptide microarray for large-scale epitope mapping of food allergens. J Allergy Clin Immunol 2009;124:315–322, 322.e1–3.
17.
Ruiter B, Knol EF, van Neerven RJ, Garssen J, Bruijnzeel-Koomen CA, Knulst AC, van Hoffen E: Maintenance of tolerance to cow’s milk in atopic individuals is characterized by high levels of specific immunoglobulin G4. Clin Exp Allergy 2007;37:1103–1110.
18.
Tomicic S, Norrman G, Falth-Magnusson K, Jenmalm MC, Devenney I, Bottcher MF: High levels of IgG4 antibodies to foods during infancy are associated with tolerance to corresponding foods later in life. Pediatr Allergy Immunol 2009;20:35–41.
19.
Cerecedo I, Zamora J, Shreffler WG, Lin J, Bardina L, Dieguez MC, Wang J, Muriel A, de la Hoz B, Sampson HA: Mapping of the IgE and IgG4 sequential epitopes of milk allergens with a peptide microarray-based immunoassay. J Allergy Clin Immunol 2008;122:589–594.
20.
Lemon-Mule H, Sampson HA, Sicherer SH, Shreffler WG, Noone S, Nowak-Wegrzyn A: Immunologic changes in children with egg allergy ingesting extensively heated egg. J Allergy Clin Immunol 2008;122:977–983.e1.
21.
Flinterman AE, Knol EF, Lencer DA, Bardina L, den Hartog Jager CF, Lin J, Pasmans SG, Bruijnzeel-Koomen CA, Sampson HA, van Hoffen E, Shreffler WG: Peanut epitopes for IgE and IgG4 in peanut-sensitized children in relation to severity of peanut allergy. J Allergy Clin Immunol 2008;121:737–743.e10.
22.
Wang J, Lin J, Bardina L, Goldis M, Nowak-Wegrzyn A, Shreffler WG, Sampson HA: Correlation of IgE/IgG4 milk epitopes and affinity of milk-specific IgE antibodies with different phenotypes of clinical milk allergy. J Allergy Clin Immunol 2010;125:695–702, 702.e1–702.e6.
23.
Mine Y, Wei Zhang J: Identification and fine mapping of IgG and IgE epitopes in ovomucoid. Biochem Biophys Res Commun 2002;292:1070–1074.
24.
Lin J, Sampson HA: The role of immunoglobulin E-binding epitopes in the characterization of food allergy. Curr Opin Allergy Clin Immunol 2009;9:357–363.
25.
Shreffler WG, Lencer DA, Bardina L, Sampson HA: IgE and IgG4 epitope mapping by microarray immunoassay reveals the diversity of immune response to the peanut allergen, Ara h 2. J Allergy Clin Immunol 2005;116:893–899.
26.
Takagi K, Teshima R, Okunuki H, Sawada J: Comparative study of in vitro digestibility of food proteins and effect of preheating on the digestion. Biol Pharm Bull 2003;26:969–973.
27.
Des Roches A, Nguyen M, Paradis L, Primeau MN, Singer S: Tolerance to cooked egg in an egg allergic population. Allergy 2006;61:900–901.
28.
Konstantinou GN, Giavi S, Kalobatsou A, Vassilopoulou E, Douladiris N, Saxoni-Papageorgiou P, Papadopoulos NG: Consumption of heat-treated egg by children allergic or sensitized to egg can affect the natural course of egg allergy: hypothesis-generating observations. J Allergy Clin Immunol 2008;122:414–415.
29.
Ayuso R, Sanchez-Garcia S, Lin J, Fu Z, Ibanez MD, Carrillo T, Blanco C, Goldis M, Bardina L, Sastre J, Sampson HA: Greater epitope recognition of shrimp allergens by children than by adults suggests that shrimp sensitization decreases with age. J Allergy Clin Immunol 2010;125:1286–1293.e3.
30.
Vereda A, Andreae DA, Lin J, Shreffler WG, Ibanez MD, Cuesta-Herranz J, Bardina L, Sampson HA: Identification of IgE sequential epitopes of lentil (Len c 1) by means of peptide microarray immunoassay. J Allergy Clin Immunol 2010;126:596–601.e1.
31.
Holen E, Bolann B, Elsayed S: Novel B and T cell epitopes of chicken ovomucoid (Gal d 1) induce T cell secretion of IL-6, IL-13, and IFN-gamma. Clin Exp Allergy 2001;31:952–964.
32.
Besler M, Petersen A, Steinhart H, Paschke A: Identification of IgE-binding peptides derived from chemical and enzymatic cleavage of ovomucoid (Gal d 1). Internet Symposium on Food Allergens (http://www.food-allergens.de) 1999;1:1–12.
33.
Hoffman DR: Immunochemical identification of the allergens in egg white. J Allergy Clin Immunol 1983;71:481–486.
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2012
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