An antibody-free dual-biomarker rapid enrichment workflow (AnDREW) improves the sensitivity of malaria rapid diagnostic tests

• Published in: Analytical biochemistry 2021-01, Vol.612, p.114020-114020, Article 114020
• Main Authors: Kantor, Andrew G., Markwalter, Christine F., Nourani, Armin, Wright, David W.
• Format: Article
• Language: English
• Subjects: Aptamer; Immobilized metal affinity chromatography (IMAC); Malaria; Plasmodium vivax; Rapid diagnostic test (RDT)
• ISSN: 0003-2697; 1096-0309
• DOI: 10.1016/j.ab.2020.114020

Rapid diagnostic tests (RDTs) are critical to the success of malaria elimination campaigns. These tests are rapid, user-friendly, and field-deployable to resource-limited regions. However, RDTs demonstrate poor sensitivity because they can only tolerate a small (5 μL) volume of blood, which limits the amount of protein biomarker delivered to the test. We have developed the Antibody-free Dual-biomarker Rapid Enrichment Workflow (AnDREW) for purifying histidine-rich protein 2 (HRP2) and Plasmodium lactate dehydrogenase (PLDH) from large volume (150 μL) blood samples. We used Zn(II)NTA and aptamer-conjugated magnetic beads to capture HRP2 and PLDH, respectively. Both biomarkers were then eluted into RDT-compatible volumes using ethylene diamine tetraacetic acid (EDTA). We optimized both bead conjugates individually by enzyme-linked immunosorbent assays (ELISAs) and then combined the optimized capture and elution assays for both biomarkers to produce the AnDREW. The AnDREW-enhanced RDTs exhibited a 11-fold and 9-fold improvement in analytical sensitivity for detection of HRP2 and PLDH, respectively, when compared to unenhanced RDTs. Moreover, the limit of detection for PLDH was improved 11-fold for the AnDREW-enhanced RDTs (3.80 parasites/μL) compared to unenhanced RDTs (42.31 parasites/μL). Importantly, the AnDREW utilizes a pan-specific PLDH aptamer and improves upon existing methods by eluting both biomarkers without complexed antibodies. [Display omitted] •Magnetic beads capture malaria biomarkers from large volume blood samples.•EDTA can elute biomarkers in metal-dependent diagnostic assays.•Sample preparation improves the sensitivity & detection limit of malaria RDTs.