PMID- 31941161 OWN - NLM STAT- MEDLINE DCOM- 20200930 LR - 20200930 IS - 1422-0067 (Electronic) IS - 1422-0067 (Linking) VI - 21 IP - 2 DP - 2020 Jan 13 TI - Acid Stripping of Surface IgE Antibodies Bound to FcepsilonRI is Unsuitable for the Functional Assays that Require Long-Term Culture of Basophils and Entire Removal of Surface IgE. LID - 10.3390/ijms21020510 [doi] LID - 510 AB - Basophils are rare granulocytes and dysregulated functions of these cells are associated with several atopic and non-atopic allergic diseases of skin, respiratory system and gastrointestinal tract. Both cytokines and immunoglobulin E (IgE) are implicated in mediating the basophil activation and pathogenesis of these disorders. Several reports have shown that healthy individuals, and patients with allergic disorders display IgG autoantibodies to IgE and hence functional characterization of these anti-IgE IgG autoantibodies is critical. In general, anti-IgE IgG autoantibodies modulate basophil activation irrespective of allergen specificity by interacting with constant domains of IgE. Therefore, an ideal solution to prove the functions of such anti-IgE IgG autoantibodies would be to completely eliminate type I high affinity immunoglobulin E receptor (FcvarepsilonRI)-bound IgE from the surface of basophils and to demonstrate in an unequivocal manner the role of anti-IgE IgG autoantibodies. In line with previous reports, our data show that FcvarepsilonRI on peripheral blood basophils are almost saturated with IgE. Further, acetic acid buffer (pH 4) efficiently removes these FcvarepsilonRI-bound IgE. Although immediately following acetic acid-elution of IgE had no repercussion on the viability of basophils, following 24 hours culture with interleukin-3 (IL-3), the viability and yield of basophils were drastically reduced in acid-treated cells and had repercussion on the induction of activation markers. Lactic acid treatment on the other hand though had no adverse effects on the viability of basophils and IL-3-induced activation, it removed only a small fraction of the cell surface bound IgE. Thus, our results show that acid buffers could be used for the elution of FcvarepsilonRI-bound IgE on the basophil surface for the biochemical characterization of IgE antibodies or for the immediate use of basophils to determine their sensitivity to undergo degranulation by specific allergens. However, these methods are not utile for the functional assays of basophils that require longer duration of culture and entire removal of surface IgE to validate the role of anti-IgE IgG autoantibodies that interact with FcvarepsilonRI-bound IgE irrespective of allergen specificity. FAU - Galeotti, Caroline AU - Galeotti C AD - Institut National de la Sante et de la Recherche Medicale, Centre de Recherche des Cordeliers, Equipe-Immunopathologie et Immunointervention Therapeutique, Sorbonne Universite, Paris, F-75006, France. AD - Service de Rhumatologie Pediatrique, Centre de Reference des Maladies Auto-Inflammatoires Rares et des Amyloses, CHU de Bicetre, le Kremlin Bicetre, F-94270 Paris, France. FAU - Karnam, Anupama AU - Karnam A AD - Institut National de la Sante et de la Recherche Medicale, Centre de Recherche des Cordeliers, Equipe-Immunopathologie et Immunointervention Therapeutique, Sorbonne Universite, Paris, F-75006, France. FAU - Das, Mrinmoy AU - Das M AD - Institut National de la Sante et de la Recherche Medicale, Centre de Recherche des Cordeliers, Equipe-Immunopathologie et Immunointervention Therapeutique, Sorbonne Universite, Paris, F-75006, France. FAU - Kaveri, Srini V AU - Kaveri SV AD - Institut National de la Sante et de la Recherche Medicale, Centre de Recherche des Cordeliers, Equipe-Immunopathologie et Immunointervention Therapeutique, Sorbonne Universite, Paris, F-75006, France. AD - Universite Paris Descartes, Sorbonne Paris Cite, F-75006 Paris, France. FAU - Bayry, Jagadeesh AU - Bayry J AD - Institut National de la Sante et de la Recherche Medicale, Centre de Recherche des Cordeliers, Equipe-Immunopathologie et Immunointervention Therapeutique, Sorbonne Universite, Paris, F-75006, France. AD - Universite Paris Descartes, Sorbonne Paris Cite, F-75006 Paris, France. LA - eng GR - ANR-19-CE17-0021 (BASIN)/Agence Nationale de la Recherche/ PT - Journal Article DEP - 20200113 PL - Switzerland TA - Int J Mol Sci JT - International journal of molecular sciences JID - 101092791 RN - 0 (Receptors, IgE) RN - 37341-29-0 (Immunoglobulin E) RN - Q40Q9N063P (Acetic Acid) SB - IM MH - *Acetic Acid/chemistry/pharmacology MH - *Basophils/chemistry/immunology MH - *Biological Assay MH - Cell Culture Techniques MH - Humans MH - *Immunoglobulin E/chemistry/immunology MH - Receptors, IgE/*immunology PMC - PMC7014331 OTO - NOTNLM OT - CD123 OT - IL-3 OT - IgE OT - acetic acid OT - activation OT - allergy OT - atopy OT - basophil OT - human OT - inflammation OT - lactic acid OT - stripping OT - viability COIS- The work is supported in part by research grant from CSL Behring, Switzerland. The funder has no role in the design of the study. The authors have no other conflicts of interest and sponsors. EDAT- 2020/01/17 06:00 MHDA- 2020/10/02 06:00 PMCR- 2020/01/01 CRDT- 2020/01/17 06:00 PHST- 2019/12/22 00:00 [received] PHST- 2020/01/10 00:00 [accepted] PHST- 2020/01/17 06:00 [entrez] PHST- 2020/01/17 06:00 [pubmed] PHST- 2020/10/02 06:00 [medline] PHST- 2020/01/01 00:00 [pmc-release] AID - ijms21020510 [pii] AID - ijms-21-00510 [pii] AID - 10.3390/ijms21020510 [doi] PST - epublish SO - Int J Mol Sci. 2020 Jan 13;21(2):510. doi: 10.3390/ijms21020510.