Development of β-lactoglobulin-specific chimeric human IgEκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates.
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 wit...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Public Library of Science (PLoS)
2014-01-01
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| Series: | PLoS ONE |
| Online Access: | http://europepmc.org/articles/PMC4143325?pdf=render |
| _version_ | 1818846563239198720 |
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| author | Karen Knipping Peter J Simons Laura S Buelens-Sleumer Linda Cox Marcel den Hartog Niels de Jong Reiko Teshima Johan Garssen Louis Boon Léon M J Knippels |
| author_facet | Karen Knipping Peter J Simons Laura S Buelens-Sleumer Linda Cox Marcel den Hartog Niels de Jong Reiko Teshima Johan Garssen Louis Boon Léon M J Knippels |
| author_sort | Karen Knipping |
| collection | DOAJ |
| description | 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.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.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.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.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. |
| first_indexed | 2024-12-19T05:47:32Z |
| format | Article |
| id | doaj.art-a150c17cbb3d48fe9f30c148a09fa9b3 |
| institution | Directory Open Access Journal |
| issn | 1932-6203 |
| language | English |
| last_indexed | 2024-12-19T05:47:32Z |
| publishDate | 2014-01-01 |
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| series | PLoS ONE |
| spelling | doaj.art-a150c17cbb3d48fe9f30c148a09fa9b32022-12-21T20:33:47ZengPublic Library of Science (PLoS)PLoS ONE1932-62032014-01-0198e10602510.1371/journal.pone.0106025Development of β-lactoglobulin-specific chimeric human IgEκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates.Karen KnippingPeter J SimonsLaura S Buelens-SleumerLinda CoxMarcel den HartogNiels de JongReiko TeshimaJohan GarssenLouis BoonLéon M J KnippelsCow'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.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.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.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.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.http://europepmc.org/articles/PMC4143325?pdf=render |
| spellingShingle | Karen Knipping Peter J Simons Laura S Buelens-Sleumer Linda Cox Marcel den Hartog Niels de Jong Reiko Teshima Johan Garssen Louis Boon Léon M J Knippels Development of β-lactoglobulin-specific chimeric human IgEκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates. PLoS ONE |
| title | Development of β-lactoglobulin-specific chimeric human IgEκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates. |
| title_full | Development of β-lactoglobulin-specific chimeric human IgEκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates. |
| title_fullStr | Development of β-lactoglobulin-specific chimeric human IgEκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates. |
| title_full_unstemmed | Development of β-lactoglobulin-specific chimeric human IgEκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates. |
| title_short | Development of β-lactoglobulin-specific chimeric human IgEκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates. |
| title_sort | development of β lactoglobulin specific chimeric human igeκ monoclonal antibodies for in vitro safety assessment of whey hydrolysates |
| url | http://europepmc.org/articles/PMC4143325?pdf=render |
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