Quantification of malaria antigens PfHRP2 and pLDH by quantitative suspension array technology in whole blood, dried blood spot and plasma

Abstract

Background: Malaria diagnostics by rapid diagnostic test (RDT) relies primarily on the qualitative detection of Plasmodium falciparum histidine-rich protein 2 (PfHRP2) and Plasmodium spp lactate dehydrogenase (pLDH). As novel RDTs with increased sensitivity are being developed and implemented as point of care diagnostics, highly sensitive laboratory-based assays are needed for evaluating RDT performance. Here, a quantitative suspension array technology (qSAT) was developed, validated and applied for the simultaneous detection of PfHRP2 and pLDH in a variety of biological samples (whole blood, plasma and dried blood spots) from individuals living in different endemic countries. Results: The qSAT was specific for the target antigens, with analytical ranges of 6.8 to 762.8 pg/ml for PfHRP2 and 78.1 to 17076.6 pg/ml for P. falciparum LDH (Pf-LDH). The assay detected Plasmodium vivax LDH (Pv-LDH) at a lower sensitivity than Pf-LDH (analytical range of 1093.20 to 187288.5 pg/ml). Both PfHRP2 and pLDH levels determined using the qSAT showed to positively correlate with parasite densities determined by quantitative PCR (Spearman r = 0.59 and 0.75, respectively) as well as microscopy (Spearman r = 0.40 and 0.75, respectively), suggesting the assay to be a good predictor of parasite density. Conclusion: This immunoassay can be used as a reference test for the detection and quantification of PfHRP2 and pLDH, and could serve for external validation of RDT performance, to determine antigen persistence after parasite clearance, as well as a complementary tool to assess malaria burden in endemic settings.

Ultrasensitive antibody-aptamer plasmonic biosensor for malaria biomarker detection in whole blood

“…In this way, not only the interacting area of the biosensor is fully exploited thereby obtaining high sensitivity and low LOD, but also the specificity is inherently warranted by the presence of the antibodies (and even more increased in our scheme by the presence of the aptamers). As a result of the occurrence of optimal conditions for detecting PfLDH, we were able to measure a LOD of 18 fM (0.6 pg/mL), which is comparable to that recently measured by means of a much more complex Luminex technology 69 .…”

Plasmodium falciparum pfhrp2 and pfhrp3 gene deletions among patients in the DRC enrolled from 2017 to 2018

“…The use multiplex antigen detection provides a high-throughput, sensitive antigen screen that allows for identification of samples with low HRP2/3 expression requiring pfhrp2/3 deletion genotyping 22 , 23 , 29 – 31 and assays for the actual antigen target of the HRP2-based RDTs. Other laboratory assays are also available which may be able to provide sensitive multiplex data to screen for potential pfhrp2/3 deletions by antigen profiles 32 , 33 .…”

Prevalence and clinical impact of malaria infections detected with a highly sensitive HRP2 rapid diagnostic test in Beninese pregnant women

“…HRP2 was quantified using a highly sensitive laboratory based quantitative bead suspension array (qSA) based on Luminex technology [17]. After incubating 2'000 magnetic beads per analyte with blood sample at 1:5 dilution, beads were sequentially incubated with in-house biotinylated antibody α-HRP2 (MBS832975, MyBiSource, San Diego, CL) and streptavidin-PE (42250-1ML, Sigma Aldrich, St. Louis, MO).…”