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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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mSPE-7 IgE trimer displays cytokinergic activity. (A) Improved size fractionation of mSPE-7 IgE was achieved with a Superdex 200 Increase HPLC column. (B) SEC-MALLS analysis of peaks A, B and C from the column determined the percentage of high molecular weight aggregates, trimers and monomers in Peaks A, B and C, respectively. The percentages of each in the three fractions are indicated. (C) No RBL-SX38 cell degranulation above buffer background control was induced by incubation with the IgE in peak A preparation (65.6% aggregated IgE), in the absence of antigen. (D) Significant, but low-level degranulation, compared to buffer background control was induced by incubation with 5 μg/ml of the IgE in peak C (95.1% monomeric IgE and 4.1% trimeric IgE) in the absence of antigen. Incubation with unpurified Sigma mSPE-7 IgE, in absence of antigen, resulted in significant RBL-SX38 degranulation compared to buffer background control in both experiments. (E) Significant RBL-SX38 cell degranulation was induced by incubation with NS1 mSPE-7 IgE trimer in the absence of antigen (peak B in purification profile). (F) Degranulation induced by peaks A, B and C at 5 μg/ml is compared. Means of 3 independent experiments ± SEM are shown. Statistically significant difference to background control (unless otherwise indicated) was determined by one-way ANOVA with Dunnett’s post-test; ****P < 0.0001, **P = 0.001 to 0.01, ns P > 0.05.