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Results and harmonization guidelines from two large-scale international Elispot proficiency panels conducted by the Cancer Vaccine Consortium (CVC/SVI)

机译：癌症疫苗协会（CVC / SVI）开展的两个大型国际Elispot水平专家小组的结果和统一指南

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The Cancer Vaccine Consortium of the Sabin Vaccine Institute (CVC/SVI) is conducting an ongoing large-scale immune monitoring harmonization program through its members and affiliated associations. This effort was brought to life as an external validation program by conducting an international Elispot proficiency panel with 36 laboratories in 2005, and was followed by a second panel with 29 participating laboratories in 2006 allowing for application of learnings from the first panel. Critical protocol choices, as well as standardization and validation practices among laboratories were assessed through detailed surveys. Although panel participants had to follow general guidelines in order to allow comparison of results, each laboratory was able to use its own protocols, materials and reagents. The second panel recorded an overall significantly improved performance, as measured by the ability to detect all predefined responses correctly. Protocol choices and laboratory practices, which can have a dramatic effect on the overall assay outcome, were identified and lead to the following recommendations: (A) Establish a laboratory SOP for Elispot testing procedures including (A1) a counting method for apoptotic cells for determining adequate cell dilution for plating, and (A2) overnight rest of cells prior to plating and incubation, (B) Use only pre-tested serum optimized for low background: high signal ratio, (C) Establish a laboratory SOP for plate reading including (C1) human auditing during the reading process and (C2) adequate adjustments for technical artifacts, and (D) Only allow trained personnel, which is certified per laboratory SOPs to conduct assays. Recommendations described under (A) were found to make a statistically significant difference in assay performance, while the remaining recommendations are based on practical experiences confirmed by the panel results, which could not be statistically tested. These results provide initial harmonization guidelines to optimize Elispot assay performance to the immunotherapy community. Further optimization is in process with ongoing panels.
机译：萨宾疫苗研究所（SVC）的癌症疫苗协会（CVC / SVI）正在通过其成员和附属协会进行一项正在进行的大规模免疫监测协调计划。通过在2005年由36个实验室组成的国际Elispot能力验证小组，使这项工作作为外部验证计划得以实现，随后在2006年又由29个参与实验室的第二个小组进行了应用，从而可以应用第一个小组的经验。通过详细的调查评估了关键方案的选择以及实验室之间的标准化和验证实践。尽管小组成员必须遵循一般准则以比较结果，但每个实验室都能够使用自己的方案，材料和试剂。第二小组记录了总体上显着改善的性能，通过正确检测所有预定义响应的能力来衡量。确定了可能对整体测定结果产生重大影响的方案选择和实验室操作，并提出以下建议：（A）建立用于Elispot测试程序的实验室SOP，包括（A1）凋亡细胞的计数方法以确定足够的细胞稀释液用于铺板，以及（A2）在铺板和孵育之前过夜放置细胞，（B）仅使用针对低背景：高信号比优化的经过预先测试的血清，（C）建立用于平板读数的实验室SOP，包括（ C1）在阅读过程中进行人工审核，以及（C2）对技术工件进行适当的调整，并且（D）仅允许经过实验室SOP认证的训练有素的人员进行测定。发现（A）中所述的建议在化验性能方面具有统计学上的显着差异，而其余建议则基于专家组结果确认的实践经验，无法进行统计学检验。这些结果提供了最初的协调指导，以优化Elispot分析对免疫疗法界的作用。正在进行中的小组正在进一步优化。

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