Multiplexed micronutrient, inflammation, and malarial antigenemia assessment using a plasma fractionation device

Collecting, processing, and storing blood samples for future analysis of biomarkers can be challenging when performed in resource limited environments. The preparation of dried blood spots (DBS) from heel or finger stick collection of whole blood is a widely used and established method. DBS pose less risk of infection from blood borne pathogens, do not require immediate specimen processing and tolerate a wider range of storage temperatures, and are easier to ship. As such, DBS are commonly used in large-scale surveys to assess infectious disease status and/or micronutrient status in vulnerable populations. Recently, we reported that DBS can be used with a multiplexed immunoassay, the Q-plex™ Human Micronutrient 7-plex Array (MN 7-plex). This tool can simultaneously quantify seven protein biomarkers related to micronutrient deficiencies (iodine, iron and vitamin A), inflammation and malarial antigenemia using plasma or serum. Serum ferritin, a key iron biomarker, cannot be measured from DBS due to red blood cell (RBC) ferritin confounding the results. In this study, we demonstrate the performance of a simple and rapid blood fractionation tool that passively separates serum from cellular components via diffusion through a membrane into a plasma collection disc (PCD) to produce plasma spots. We evaluated the concordance of MN 7-plex analyte concentrations from matched panels of eighty-eight samples of PCD, DBS, and wet plasma prepared from anticoagulated venous whole blood. The results show high correlation between eluates from PCD and DBS and wet plasma for each analyte. Serum ferritin measures from the PCD eluates were highly correlated to wet plasma samples. This suggests that surveillance for iron deficiency may be improved over the current methods restricted to only measuring sTfR in DBS as when used in combination with the MN 7-plex, all seven biomarkers can be simultaneously measured using PCDs.

在资源受限环境中，为后续生物标志物分析而开展血液样本的采集、处理与存储工作往往颇具挑战。通过足跟或指尖采血获取全血后制备干血斑（Dried Blood Spots, DBS）是一种应用广泛且成熟的方法。干血斑（DBS）的血源性病原体感染风险更低，无需即刻进行样本处理，可耐受更广范围的存储温度，且运输更为便捷，因此常被用于大规模人群调查，以评估脆弱人群的传染病感染状态和/或微量营养素营养状况。近期我们的研究表明，干血斑（DBS）可与多重免疫检测技术Q-plex™人类微量营养素7重检测阵列（MN 7-plex）配合使用。该工具可利用血浆或血清同时定量检测7种与微量营养素缺乏（碘、铁及维生素A）、炎症及疟疾抗原血症相关的蛋白质生物标志物。血清铁蛋白作为关键的铁代谢生物标志物，无法通过干血斑（DBS）进行检测，原因是红细胞（Red Blood Cell, RBC）内的铁蛋白会干扰检测结果。本研究中，我们验证了一款简易快速的血液分离工具的性能：该工具可通过膜扩散被动分离血清与细胞成分，使血清渗透进入血浆收集盘（Plasma Collection Disc, PCD），进而制备血浆斑。我们针对88组匹配样本开展了检测一致性评估，这些样本分别来自血浆收集盘（PCD）、干血斑（DBS）以及由抗凝静脉全血制备的湿血浆，均采用MN 7-plex检测分析物浓度。检测结果显示，针对所有分析物，血浆收集盘（PCD）与干血斑（DBS）的洗脱液与湿血浆的检测结果均呈现高度相关性，血浆收集盘（PCD）洗脱液的血清铁蛋白检测结果与湿血浆样本同样呈现高度相关性。这表明，相较于当前仅能通过干血斑（DBS）检测可溶性转铁蛋白受体（sTfR）的方法，使用血浆收集盘（PCDs）结合MN 7-plex检测技术，可同时检测全部7种生物标志物，从而优化缺铁相关的监测工作。