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Development of β-Lactoglobulin-Specific Chimeric Human IgEκ Monoclonal Antibodies for In Vitro Safety Assessment of Whey Hydrolysates.
- Source :
- PLoS ONE; Aug2014, Vol. 9 Issue 8, p1-14, 14p
- Publication Year :
- 2014
-
Abstract
- Background: Cow’s milk-derived whey hydrolysates are nutritional substitutes for allergic infants. Safety or residual allergenicity assessment of these whey hydrolysates is crucial. Currently, rat basophilic leukemia RBL-2H3 cells expressing the human IgE receptor α-chain (huFcεRIα-RBL-2H3), sensitized with serum IgE from cow’s milk allergic children, are being employed to assess in vitro residual allergenicity of these whey hydrolysates. However, limited availability and inter-lot variation of these allergic sera impede standardization of whey hydrolysate safety testing in degranulation assays. Objective: An oligoclonal pool of chimeric human (chu)IgE antibodies against bovine β-lactoglobulin (a major allergen in whey) was generated to increase sensitivity, specificity, and reproducibility of existing degranulation assays. Methods: Mice were immunized with bovine β-lactoglobulin, and subsequently the variable domains of dissimilar anti-β-lactoglobulin mouse IgG antibodies were cloned and sequenced. Six chimeric antibodies were generated comprising mouse variable domains and human constant IgE/κ domains. Results: After sensitization with this pool of anti-β-lactoglobulin chuIgEs, huFcεRIα-expressing RBL-2H3 cells demonstrated degranulation upon cross-linking with whey, native 18 kDa β-lactoglobulin, and 5–10 kDa whey hydrolysates, whereas a 3 kDa whey hydrolysate and cow’s milk powder (mainly casein) showed no degranulation. In parallel, allergic serum IgEs were less sensitive. In addition, our pool anti-β-lactoglobulin chuIgEs recognized multiple allergenic immunodominant regions on β-lactoglobulin, which were also recognized by serum IgEs from cow’s milk allergic children. Conclusion: Usage of our ‘unlimited’ source and well-defined pool of β-lactoglobulin-specific recombinant chuIgEs to sensitize huFcεRIα on RBL-2H3 cells showed to be a relevant and sensitive alternative for serum IgEs from cow’s milk allergic patients to assess safety of whey-based non-allergic hydrolyzed formula. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 19326203
- Volume :
- 9
- Issue :
- 8
- Database :
- Complementary Index
- Journal :
- PLoS ONE
- Publication Type :
- Academic Journal
- Accession number :
- 97803081
- Full Text :
- https://doi.org/10.1371/journal.pone.0106025