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.在资源受限环境中开展生物标志物检测相关的血液样本采集、处理与存储工作往往颇具挑战。通过足跟采血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。干血斑可降低血源性病原体引发的感染风险，无需即刻进行样本处理，可耐受更广范围的存储温度，且更便于运输。因此，干血斑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况和/或微量营养素营养状态。近期我们的研究表明，干血斑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、铁与维生素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，无法通过干血斑检测，因为红细胞（Red Blood Cell, RBC）内的铁蛋白会对检测结果造成干扰。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将血清与细胞成分被动分离，并收集至血浆收集盘（Plasma Collection Disc, PCD）中，从而制备血浆斑。我们对88份配对样本的MN 7-plex检测物浓度进行了一致性评估，这些样本分别来自血浆收集盘、干血斑以及由抗凝静脉全血制备的湿血浆。结果显示，针对每一种检测物，血浆收集盘与干血斑的洗脱液与湿血浆之间均存在高度相关性。血浆收集盘洗脱液所测得的血清铁蛋白水平与湿血浆样本具有高度相关性。这表明，相较于当前仅能通过干血斑检测可溶性转铁蛋白受体（soluble Transferrin Receptor, sTfR）的方法，使用血浆收集盘结合MN 7-plex检测技术，可同时检测全部7种生物标志物，从而改善缺铁监测工作。