Biotechnology, Research Triangle Park, NC, 5BASF Plant Science,. Research Triangle Park, ND, 6Dow AgroSciences, Indianapolis, IN,. 7Bayer SAS, Sophia ...
Abstracts AB113
J ALLERGY CLIN IMMUNOL VOLUME 127, NUMBER 2
Post-Solid Organ Transplant Atopy Is Not Limited to Food Hypersensitivity P. Shroff1, R. S. Mehta2, J. Chinen2, S. J. Karpen2, C. M. Davis2; 1Baylor College of Medicine, Dept. of Medicine, Section of Immunology, Allergy, and Rheumatology, Houston, TX, 2Baylor College of Medicine, Dept. of Pediatrics, Houston, TX. RATIONALE: There is limited information in the literature regarding atopic disease in solid-organ transplant patients. Use of calcineurin inhibitors to prevent graft rejection may favor Th2 responses and lead to increased risk of allergen sensitization. We sought to analyze atopic manifestations of children receiving chronic immunosuppression after liver transplantation. METHODS: Chart review was performed on 176 liver transplant recipients for food allergy, asthma, allergic rhinitis, and allergic skin diseases. RESULTS: Twenty-seven (15.3%) patients were identified with atopic disease. Median age at transplant was 19 months (mean 26.3; IQR, 7-21 months) with a median follow up of 63 months (mean 77.0; IQR, 42-108 months). Tacrolimus was the main immunosuppressant. The first symptoms of atopy presented at a median of 16 months (mean 27.7, IQR 5-30 months) after transplant; and food allergy presented at a median of 14 months (mean 27.5; IQR, 6-31 months). Allergic skin disease was present in 14/27 cases (52%). Fourty-four percent of patients had asthma (n56) and/or allergic rhinitis (n59). Food allergy was suspected in 10/27 cases (37%), of which 7 had concomitant atopic disease. CONCLUSIONS: Atopy in solid organ transplant is not limited to food allergy and presents within the first three years after transplant. These data suggest a high proportion of non-food allergy atopic disease. In this population an older age of transplant may be related to a later onset of atopy than reported in the literature. These findings support the need to further elucidate the relationship of immunosuppressive drugs and atopy in children after liver transplantation.
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Relative and Absolute Quantitation of Ten Allergens in Twenty Commercial Soybean Varieties Using Tandem Mass Spectrometry N. L. Houston1, D. G. Lee1, S. E. Stevenson1, G. S. Ladics2, G. A. Bannon3, S. McClain4, L. Privalle5, N. Stagg6, C. Herouet-Guicheney7, J. J. Thelen1; 1University of Missouri, Columbia, MO, 2Pioneer Hi-Bred International, Inc., Wilmington, DE, 3Monsanto Co., St. Louis, MO, 4Syngenta Biotechnology, Research Triangle Park, NC, 5BASF Plant Science, Research Triangle Park, ND, 6Dow AgroSciences, Indianapolis, IN, 7 Bayer SAS, Sophia, FRANCE. RATIONALE: Some of the proteins expressed in the soybean (Glycine max) seed are allergenic to humans and animals; however, the concentration of these allergens and their variability in expression across germplasms is presently unknown. METHODS: Ten allergens were quantified from twenty non-genetically modified commercial soybean varieties using two parallel, label-free mass spectrometry approaches. Relative quantitation was performed by spectral counting and absolute quantitation was performed using multiple reaction monitoring (MRM) with synthetic, isotope-labeled peptides as internal standards. RESULTS: In the relative quantitation analysis, ten target allergens were identified; and five of these allergens showed concentration levels higher than the technical variation observed for the bovine serum albumin (BSA) internal standard (;11%), suggesting concentration differences among the varieties. To confirm this observation, absolute quantitation of these allergens from each variety was performed; and the concentrations of eight of the ten allergens in seed ranged from approximately 0.5 to 5.7 mg/mg of soy extract. MRM analysis reduced the technical variance of the BSA internal standard to approximately 7%, and confirmed differential concentrations for four allergens across the twenty varieties. CONCLUSIONS: This is the first quantitative assessment of its kind for a majority of the known soybean allergens and the data, taken together, show that while concentrations of individual allergens varied, the total concentration of allergens among the soy varieties was similar.
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Molecular Characterization of Ara h 1 Before and After Thermal Processing C. P. Mattison1, C. C. Grimm1, H. Wei2, S. J. Maleki1; 1USDA-ARSSRRC, New Orleans, LA, 2Waters Corp., Beverly, MA. RATIONALE: Heating and other processing techniques can alter food allergens depending upon the method and matrix involved. The increased allergenicity of Ara h 1 protein isolated from roasted peanuts is thought to be caused at least in part by chemical modifications. To identify what specific modifications are responsible for increased stability and recognition by IgE after thermal processing we are characterizing the roasting-induced molecular changes of the Ara h 1 protein. METHODS: Ara h 1 was isolated from raw and roasted peanuts, reduced with DTT, alkylated with iodoacetamide, digested with trypsin, and analyzed by liquid chromatrography coupled with mass-spectrometry on a Waters nano-Acquity UPLC, Xevo QTof and PLGS software. Additional rounds of analysis were performed on intact and digested Ara h 1 using an Agilent nano-LC with Agilent 1200 nano-pumps and a 6500 Q-Tof LC with the resulting spectra analyzed by Spectrum Mill software. RESULTS: We achieved greater than 60 percent sequence coverage of Ara h 1 isolated from either raw or roasted peanuts and find at least 16 peptides that are unique to Ara h 1 isolated from roasted peanuts. We detect peptide modification differences between Ara h 1 isolated from raw and roasted peanuts, and the protein modifications we observed include N-linked glycosylation, hydroxyproline, and carbamylated lysine residues. Some of these modifications were found within the context of previously characterized IgE epitopes. CONCLUSIONS: We have detected several chemical modifications on Ara h 1 protein isolated from raw and roasted peanuts that may be responsible for differences in IgE binding and immunogenicity.
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Contribution of Ara h 2 and Ara h 6 to the Effector Activity of Crude Peanut Extract X. Chen, Q. Wang, S. Dreskin; University of Colorado - Denver, Aurora, CO. RATIONALE: Our previous studies reported Ara h 2 and Ara h 6 account for the majority of the effector activity in crude peanut extracts (CPE). To further define the relative contributions of Ara h 2 and Ara h 6 towards the overall effector activity of crude peanut extract, we depleted CPE of Ara h 2 and Ara h 6 and then reconstituted the CPE by adding back purified allergens. METHODS: Ara h 2 and Ara h 6 were removed from CPE by immunodepletion with specific antibodies. CPE was fractioned by gel filtration. The fractions when recombined without a fraction that contained Ara h 2/6 is referred to as GF w/o 20kD. Native Ara h 2 and 6 were purified. The contents of Ara h 2 and Ara h 6 in the CPE were quantified by competitive ELISA. Reconstitution of the effector activity was measured with RBL SX-38 cells. RESULTS: Ara h 2 and Ara h 6 account for 460.3% and 7.561.5 % respectively of the total CPE protein. As previously reported, removal of Ara h 2/6 by immunodepletion and by gel filtration removed 70% and 90% of the activity, respectively. The addition of either purified Ara h 2 or Ara h 6 to GF w/o 20kd restored most of the original CPE effector activity. However, when both Ara h 2 and Ara h 6 were added the effector activity was completely restored. CONCLUSIONS: These allergen- depletion and reconstitution studies demonstrate that the two allergens together are necessary to account for the biologic activity of CPE.
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