Supplementary Material for: Suitability of Different Diagnostic Platforms for Virological Testing of Blood Samples from Cornea Donors

<b><i>Background:</i></b> To minimize the risk of disease transmission in cornea transplantation, donor screening for blood-derived viral infections is mandatory. Ideally, pre-mortem blood samples are used, but based on availability, cadaveric blood samples of cornea donors may also be used. However, serological and nucleic acid amplification tests (NATs) need to be validated for the use of cadaveric specimens. <b><i>Methods:</i></b> Hepatitis B virus (HBV), hepatitis C virus (HCV), human immunodeficiency virus (HIV), human T-lymphotropic virus (HTLV) 1/2, and <i>Treponema pallidum</i> (syphilis)-specific serological and/or NAT assays were validated on different platforms (Abbott Alinity i, Alinity m, Roche Cobas 6800, and Roche Cobas AmpliPrep/Cobas TaqMan (CAP/CTM)) using (un)spiked paired pre- and post-mortem cornea donor blood samples from the same individual (up to 23.83 h after death) of 28 individuals in accordance with the specifications of the German Federal Institute for Vaccines and Biomedicines (Paul-Ehrlich-Institut [PEI]). In addition, routinely HBV-, HCV- and HIV-PCR-negative tested post-mortem blood samples of 24 individuals were used to assess NAT specificity. <b><i>Results:</i></b> For the majority of serological parameters on the Abbott Alinity i (HBsAg, anti-HBc, anti-HBs, anti-HCV, anti-HIV, anti-HTLV 1/2, and anti-<i>Treponema pallidum</i>), ratios of generated test results of (un)spiked paired pre- and post-mortem blood samples differed ≤25%, with an agreement of qualitative pre- and post-mortem test results ranging from 91.2 to 100%. For NAT parameters (HBV, HCV, and HIV) on the Cobas 6800, Alinity m, and CAP/CTM, no significant deviation in virus concentrations (factor &gt;5) of spiked pre- and post-mortem blood samples could be observed. Ct-values of corresponding internal controls did also not differ significantly (&gt;1.5 Ct-values). In addition, no false-positive test results were generated when specificity was assessed. <b><i>Conclusion:</i></b> Overall, fluctuations of test results for serological and NAT parameters in pre- and post-mortem blood samples examined in this study, were only limited and within the range of what is also observed when routinely testing fresh patient specimens. We conclude that all examined assays are eligible for the screening of blood samples taken up to about 24 h after the occurrence of death.

**背景：** 为降低角膜移植中的疾病传播风险，对供体开展血液源性病毒感染筛查属于强制性要求。理想状态下应使用生前血液样本，但基于样本可获得性，也可采用角膜供体的死后血液样本。不过，针对死后样本的应用场景，血清学检测与核酸扩增试验（nucleic acid amplification tests, NATs）需完成验证工作。

**方法：** 本研究遵循德国联邦疫苗与生物制品研究所（Paul-Ehrlich-Institut [PEI]）的规范，针对28名个体的配对生前与死后角膜供体血液样本（死后采样时长最长可达23.83小时）的（未）加标样本，在多款检测平台（Abbott Alinity i、Alinity m、罗氏Cobas 6800、罗氏Cobas AmpliPrep/Cobas TaqMan [CAP/CTM]）上，对乙型肝炎病毒（hepatitis B virus, HBV）、丙型肝炎病毒（hepatitis C virus, HCV）、人类免疫缺陷病毒（human immunodeficiency virus, HIV）、人类T淋巴细胞白血病病毒1/2型（human T-lymphotropic virus 1/2, HTLV 1/2）以及梅毒密螺旋体（Treponema pallidum）的特异性血清学和/或核酸扩增试验进行验证。此外，本研究纳入24名个体的常规HBV、HCV及HIV-PCR检测呈阴性的死后血液样本，用于评估核酸扩增试验的特异性。

**结果：** 针对Abbott Alinity i平台上的多数血清学指标（乙型肝炎表面抗原[HBsAg]、抗乙型肝炎核心抗体[anti-HBc]、抗乙型肝炎表面抗体[anti-HBs]、抗丙型肝炎病毒抗体[anti-HCV]、抗人类免疫缺陷病毒抗体[anti-HIV]、抗人类T淋巴细胞白血病病毒1/2型抗体[anti-HTLV 1/2]以及抗梅毒密螺旋体抗体[anti-Treponema pallidum]），配对生前与死后（未）加标血液样本的检测结果比值差异≤25%，定性检测结果的符合率介于91.2%至100%区间内。针对Cobas 6800、Alinity m及CAP/CTM平台上的核酸扩增试验指标（HBV、HCV及HIV），未观察到加标生前与死后血液样本的病毒浓度存在显著差异（差异倍数＞5）。对应的内参Ct值同样未出现显著差异（差异＞1.5个Ct值）。此外，在特异性评估环节未产生假阳性检测结果。

**结论：** 总体而言，本研究中检测的血清学与核酸扩增试验指标在生前与死后血液样本中的检测结果波动幅度有限，且处于常规检测新鲜患者样本时的波动范围之内。综上，本研究涉及的所有检测方法均适用于死后约24小时内采集的血液样本的筛查工作。