Serum is not required for ex vivo IFN-gamma ELISPOT: a collaborative study of different protocols from the European CIMT Immunoguiding Program.
Serum is not required for ex vivo IFN-gamma ELISPOT: a collaborative study of different protocols from the European CIMT Immunoguiding Program.
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DOI：
10.1007/s00262-009-0814-4
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发表时间：
2010-04
期刊：
影响因子：
--
通讯作者：
Britten CM
中科院分区：
文献类型：
--
作者：
Mander A;Gouttefangeas C;Ottensmeier C;Welters MJ;Low L;van der Burg SH;Britten CM
癌症免疫治疗免疫引导计划已经进行了干扰素-γELISPOT熟练程度小组，以检查血清补充测试介质对分析性能的影响。16个欧洲实验室使用当地制定的不同方案对相同的PBMC样本进行了分析。参与者同时生成了两个数据集-一个使用添加了血清的培养液，另一个不使用血清。通过量化比较两种测试条件的性能：(1)活细胞的数量，(2)仅在介质中诱导的背景斑点形成，(3)检测抗原特异性T细胞反应的能力。研究表明，两种条件下回收的活细胞数量和总的本底斑点产量没有显著差异。此外，两种检测条件下的总体实验室表现相当；16个实验室中有11个报告使用无血清培养基时的检测率相同或更高，而5个实验室报告在无血清条件下的检测率下降。这些结果表明，干扰素-γELISPOT方法可以在无血清条件下获得良好的检测结果。对无血清条件的方案进行优化，应能获得良好的检测率，并消除血清批次和稳定性测试的要求，从而进一步统一检测方法。本文的在线版本(doi：10.1007/s00262-009-0814-4)包含补充材料，授权用户可以使用。
The Cancer Immunotherapy Immunoguiding Program has conducted an IFN-γ ELISPOT proficiency panel to examine the influence of serum supplementation of test media on assay performance. Sixteen European laboratories analyzed the same PBMC samples using different locally established protocols. Participants generated two simultaneous data sets—one using medium supplemented with serum and one without serum. Performances of the two test conditions were compared by quantifying: (1) the number of viable cells, (2) background spot formation induced in the medium only control and (3) the ability to detect antigen-specific T cell responses. The study demonstrated that the number of viable cells recovered and the overall background spot production were not significantly different between the two conditions. Furthermore, overall laboratory performance was equivalent for the two test conditions; 11 out of 16 laboratories reported equal or greater detection rates using serum-free medium, while 5 laboratories reported decreased detections rates under serum-free conditions. These results show that good performance of the IFN-γ ELISPOT assay can be achieved under serum-free conditions. Optimization of the protocol for serum-free conditions should result in excellent detection rates and eliminate the requirement of serum batch and stability testing, allowing further harmonization of the assay. The online version of this article (doi:10.1007/s00262-009-0814-4) contains supplementary material, which is available to authorized users.
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影响因子：
8.4
作者：
Copier, J.;Dalgleish, A. G.;Hakansson, L.
通讯作者：
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DOI：
10.1080/00365510701809235
发表时间：
2008-01-01
影响因子：
2.1
作者：
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通讯作者：
Morkrid, Lars
影响因子：
7.7
作者：
Elliott, Paul;Peakman, Tim C.
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