Development of an IFNγ ELISPOT for the analysis of the human T cell response against mumps virus

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• 作者：Wanda G.H. Han ; ^{wanda.han@rivm.nl" class="auth_mail" title="E-mail the corresponding author} ; Maarten E. Emmelot ; Haziz Jaadar ; Hinke I. ten Hulscher ; Cé ; cile A.C.M. van Els ; Patricia Kaaijk
• 关键词：T cell ELISPOT ; Mumps virus ; Optimization ; NK cells ; IFNγ
• 刊名：Journal of Immunological Methods
• 出版年：2016
• 出版时间：April 2016
• 年：2016
• 卷：431
• 期：Complete
• 页码：52-59
• 全文大小：1006 K

文摘

In the last decade, mumps virus (MuV) causes outbreaks in highly vaccinated populations. Sub-optimal T cell immunity may play a role in the susceptibility to mumps in vaccinated individuals. T cell responses to mumps virus have been demonstrated, yet the quality of the MuV-specific T cell response has not been analyzed using single cell immunological techniques. Here we developed an IFNγ ELISPOT assay to assess MuV-specific T cell responses in peripheral blood mononuclear cells (PBMC) of healthy (vaccinated) donors and mumps patients. Various in vitro MuV-specific stimulation methods of PBMC were compared, using either live or inactivated MuV alone or MuV-infected autologous antigen presenting cells, i.e. Epstein Barr Virus-transformed B lymphoblastoid cell lines (EBV-BLCL) or (mitogen pre-activated) PBMC, for their ability to recall IFNγ-producing responder cells measured by ELISPOT. For the detection of MuV-specific T cell responses, direct exposure (24 h) to live MuV was the preferred stimulation method when assay sensitivity and practical reasons were considered. Notably, flowcytometric confirmation of data revealed that primarily T cells and NK cells produce IFNγ upon live MuV stimulation. Depleting PBMC from CD56⁺ NK cells prior to stimulation with live MuV led to the enumeration of MuV-specific T cell responses by ELISPOT. Our assay constitutes a tool to evaluate memory MuV-specific T cell responses in MuV vaccinated or infected persons. Furthermore, this study provides evidence that live MuV not only induces IFNγ production by T cells, but also by NK cells.