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Identification and isolation of a Fel d 1–like molecule as a major rabbit allergen  Christiane Hilger, PhD, Stéphanie Kler, MSc, Karthik Arumugam, PhD,

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Presentation on theme: "Identification and isolation of a Fel d 1–like molecule as a major rabbit allergen  Christiane Hilger, PhD, Stéphanie Kler, MSc, Karthik Arumugam, PhD,"— Presentation transcript:

1 Identification and isolation of a Fel d 1–like molecule as a major rabbit allergen 
Christiane Hilger, PhD, Stéphanie Kler, MSc, Karthik Arumugam, PhD, Dominique Revets, MSc, Claude P. Muller, MD, Catherine Charpentier, MD, Christiane Lehners, MD, Martine Morisset, MD, PhD, François Hentges, MD  Journal of Allergy and Clinical Immunology  Volume 133, Issue 3, Pages (March 2014) DOI: /j.jaci Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 Detection of IgE-reactive proteins in rabbit hair extract (A) and a preparation of purified lipophilins (B). Proteins were separated by means of 15% SDS-PAGE under denaturing conditions and analyzed in IgE immunoblots with sera of patients with rabbit allergy. Numbered lanes correspond to patients' sera in Table I. E, Coomassie-stained rabbit hair extract; L, Coomassie-stained purified Ory c 3; M, Coomassie-stained molecular weight marker; N, negative control. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 Mass spectrometric analysis of purified deglycosylated (A) and native (B) Ory c 3. The 3 peaks in Fig 2, B, show the heterogeneous molecular weight of Ory c 3 in comparison with the deglycosylated sample in Fig 2, A. The insert in Fig 2, B, shows a silver-stained SDS-PAGE of purified lipophilins without (−) or with (+) addition of N-Glycosidase F. M, Molecular weight marker; non red, nonreducing conditions; red, reducing conditions. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig 3 Detection of IgE reactivity to native or deglycosylated Ory c 3 by means of ELISA among 27 Ory c 3–positive patients. Results are shown as ODs. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig 4 Basophil activation test induced by Ory c 3. Patients' basophils were sensitized with increasing concentrations of native Ory c 3. A patient without IgE to Ory c 3 served as a negative control. Relative cell counts of CD63+ basophils, CD203c+ basophils, or both were determined by using flow cytometry. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig 5 Lipophilin secondary structure model. A, Model of the Ory c 3 CL2-AL heterodimer. B, Secondary structure superimposition comparison for Fel d 1 chain A (red) and lipophilin CL2 (yellow). C, Secondary structure superimposition comparison for Fel d 1 chain B (orange) and lipophilin AL (blue). Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

7 Fig 6 Sequence alignment of Ory c 3 and Fel d 1. Ory c 3A.0101 (lipophilin CL2, Q9GK63) is aligned with Fel d 1 chain A (P30438), and Ory c 3.B0101 (lipophilin AL, Q9GK67) is aligned with Fel d 1 chain B (P30440). Signal peptide cleavage sites are marked by arrows, and glycosylation sites are marked by stars. Identical amino acids are shaded in dark gray, and similar amino acids are shaded in light gray. Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

8 Fig E1 Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

9 Fig E2 Journal of Allergy and Clinical Immunology  , DOI: ( /j.jaci ) Copyright © 2013 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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