The reproducibility of assessing potential biomarkers is crucial for their implementation. ONEST (Observers Needed to Evaluate Subjective Tests) has been recently introduced as a new additive evaluation method for the assessment of reliability, by demonstrating how the number of observers impact on interobserver agreement. Oestrogen receptor (ER), progesterone receptor (PR), and Ki67 proliferation marker immunohistochemical stainings were assessed on 50 core needle biopsy and 50 excision samples from breast cancers by 9 pathologists according to daily practice. ER and PR statuses based on the percentages of stained nuclei were the most consistently assessed parameters (intraclass correlation coefficients, ICC 0.918–0.996), whereas Ki67 with 5 different theoretical or St Gallen Consensus Conference–proposed cut-off values demonstrated moderate to good reproducibility (ICC: 0.625–0.760). ONEST highlighted that consistent tests like ER and PR assessment needed only 2 or 3 observers for optimal evaluation of reproducibility, and the width between plots of the best and worst overall percent agreement values for 100 randomly selected permutations of observers was narrow. In contrast, with less consistently evaluated tests of Ki67 categorization, ONEST suggested at least 5 observers required for more trustful assessment of reliability, and the bandwidth of the best and worst plots was wider (up to 34% difference between two observers). ONEST has additional value to traditional calculations of the interobserver agreement by not only highlighting the number of observers needed to trustfully evaluate reproducibility but also by highlighting the rate of agreement with an increasing number of observers and disagreement between the better and worse ratings.

评估潜在生物标志物的可重复性对其实施至关重要。ONEST（评估主观测试所需的观察者）最近被引入作为一种新的附加评估方法，用于评估可靠性，通过展示观察者的数量如何影响观察者间的一致性。9 名病理学家根据日常实践对 50 个核心穿刺活检和 50 个乳腺癌切除样本进行了雌激素受体 (ER)、孕激素受体 (PR) 和 Ki67 增殖标志物免疫组织化学染色评估。基于染色核百分比的 ER​​ 和 PR 状态是最一致的评估参数（组内相关系数，ICC 0.918-0.996），而具有 5 个不同理论或圣加仑共识会议提出的截止值的 Ki67 表现出中等至良好的可重复性（ICC：0.625-0.760）。ONEST 强调，像 ER 和 PR 评估这样的一致性测试只需要 2 或 3 名观察者即可对再现性进行最佳评估，并且对于 100 个随机选择的观察者排列，最佳和最差总体一致性百分比值的图之间的宽度很窄。相比之下，由于对 Ki67 分类的评估不太一致，ONEST 建议至少需要 5 名观察者才能更可靠地评估可靠性，并且最佳和最差图的带宽更宽（两个观察者之间的差异高达 34%）。