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. 2017 Aug 15;7:8164. doi: 10.1038/s41598-017-08212-6

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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SPE-7 IgE monomer is not cytokinergic. Monomeric IgE was isolated from (A) Sigma mSPE-7 IgE (Sigma mSPE-7), (B) NS1 hybridoma cell mSPE-7 IgE (NS1 mSPE-7), and (C) recombinant mSPE-7 and chSPE-7 IgE (Rec. mSPE-7 and Rec. chSPE-7) preparations by size-exclusion chromatography using the Superdex S200 HPLC column. Purification profiles (left panels) show selected monomeric fractions between dotted vertical lines. Incubation with unpurified Sigma mSPE-7 IgE in the absence of antigen resulted in significant RBL-SX38 degranulation compared to buffer background control in all experiments (right panels, black lower bars; ****P < 0.0001). Monomeric IgE from all preparations did not induce degranulation of RBL-SX38 cells compared to buffer background control (right panels, black lower bars; ns P > 0.05). Functionality of the monomeric IgE antibodies was confirmed by measurement of degranulation upon cross-linking with multimeric antigen, DNP-HSA (right panels, upper white bars). Means of 3 or 4 independent experiments ± SEM are shown. Statistically significant difference to background control was determined by one-way ANOVA with Dunnett’s post-test.