Assessment of the prozone effect in malaria rapid diagnostic tests

Department of Clinical Sciences, Institute of Tropical Medicine (ITM), Nationalestraat 155, B 2000 Antwerp, Belgium.
Malaria Journal (Impact Factor: 3.49). 11/2009; 8:271. DOI: 10.1186/1475-2875-8-271
Source: PubMed

ABSTRACT The prozone effect (or high doses-hook phenomenon) consists of false-negative or false-low results in immunological tests, due to an excess of either antigens or antibodies. Although frequently cited as a cause of false-negative results in malaria rapid diagnostic tests (RDTs), especially at high parasite densities of Plasmodium falciparum, it has been poorly documented. In this study, a panel of malaria RDTs was challenged with clinical samples with P. falciparum hyperparasitaemia (> 5% infected red blood cells).
Twenty-two RDT brands were tested with seven samples, both undiluted and upon 10 x, 50 x and 100 x dilutions in NaCl 0.9%. The P. falciparum targets included histidine-rich protein-2 (HRP-2, n = 17) and P. falciparum-specific parasite lactate dehydrogenase (Pf-pLDH, n = 5). Test lines intensities were recorded in the following categories: negative, faint, weak, medium or strong. The prozone effect was defined as an increase in test line intensity of at least one category after dilution, if observed upon duplicate testing and by two readers.
Sixteen of the 17 HRP-2 based RDTs were affected by prozone: the prozone effect was observed in at least one RDT sample/brand combination for 16/17 HRP-2 based RDTs in 6/7 samples, but not for any of the Pf-pLDH tests. The HRP-2 line intensities of the undiluted sample/brand combinations with prozone effect (n = 51) included a single negative (1.9%) and 29 faint and weak readings (56.9%). The other target lens (P. vivax-pLDH, pan-specific pLDH and aldolase) did not show a prozone effect.
This study confirms the prozone effect as a cause of false-negative HRP-2 RDTs in samples with hyperparasitaemia.

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    • "Firstly, manufacturer recommendations on the appropriate sample volume to be applied should always be followed, as application of too large a volume can elicit the phenom‐ enon. [58] The prozone effect is also abated upon dilution of the patients' samples, and so for patients presenting severe clinical manifestations of malaria, health care workers can elect to verify a weak or negative test result using a diluted sample. [60] Finally, the use of alternative device architectures may prevent the occurrence of the prozone effect. "
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    ABSTRACT: Immunochromatographic rapid diagnostic tests (RDTs) have demonstrated significant potential for use as point-of-care diagnostic tests in resource-limited settings. Most notably, RDTs for malaria have reached an unparalleled level of technological maturity and market penetration, and are now considered an important complement to standard microscopic methods of malaria diagnosis. However, the technical development of RDTs for other infectious diseases, and their uptake within the global health community as a core diagnostic modality, has been hindered by a number of extant challenges. These range from technical and biological issues, such as the need for better affinity agents and biomarkers of disease, to social, infrastructural, regulatory and economic barriers, which have all served to slow their adoption and diminish their impact. In order for the immunochromatographic RDT format to be successfully adapted to other disease targets, to see widespread distribution, and to improve clinical outcomes for patients on a global scale, these challenges must be identified and addressed, and the global health community must be engaged in championing the broader use of RDTs.
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    • "The presence of polymorphisms in the PfHRP2 antigen (Mariette et al., 2008), or the absence of the gene and thus the targeted antigen sequence, means that alternative RDTs other than a PfHRP2-based RDT need to be considered for the diagnosis of a malaria infection in areas where deletions of the Pfhrp2 gene have been detected. False-negative findings can be explained by the absence of bands on an RDT either from excess antibodies or antigens – a phenomenon called the prozone effect, which appears to be restricted to PfHRP2-based RDTs (Gillet et al., 2009; Luchavez et al., 2011). "
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    ABSTRACT: In the last decade, there has been an upsurge of interest in developing malaria rapid diagnostic test (RDT) kits for the detection of Plasmodium species. Three antigens - Plasmodium falciparum histidine-rich protein 2 (PfHRP2), plasmodial aldolase and plasmodial lactate dehydrogenase (pLDH) - are currently used for RDTs. Tests targeting HRP2 contribute to more than 90 % of the malaria RDTs in current use. However, the specificities, sensitivities, numbers of false positives, numbers of false negatives and temperature tolerances of these tests vary considerably, illustrating the difficulties and challenges facing current RDTs. This paper describes recent developments in malaria RDTs, reviewing RDTs detecting PfHRP2, pLDH and plasmodial aldolase. The difficulties associated with RDTs, such as genetic variability in the Pfhrp2 gene and the persistence of antigens in the bloodstream following the elimination of parasites, are discussed. The prospect of overcoming the problems associated with current RDTs with a new generation of alternative malaria antigen targets is also described.
    Journal of Medical Microbiology 10/2013; 62(Pt 10):1491-505. DOI:10.1099/jmm.0.052506-0 · 2.27 Impact Factor
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    • "This might be related to several factors such as the variability of the target antigen, which is not uncommon in Africa (Koita et al., 2012), or the prozone effect observed in case of high P. falciparum or non–P. falciparum malaria parasitemia (Farcas et al., 2003; Fernando et al., 2004; Wongsrichanalai et al., 2003; Gillet et al., 2009). "
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    ABSTRACT: The Malaria Pf Rapid Test Device Acon® (Acon Labs) and the pan HRP2/aldolase RDT, Malaria P.f/Pan Rapid Test Device Acon® (Acon Labs), performances were evaluated for malaria species diagnosis in 592 febrile patients living in Gabon using microscopy as gold standard. Sensitivities were equal or above 96.0% for Plasmodium falciparum detection, of 62.5% for non-P. falciparum malaria species detection and higher in younger children (100%). Negative predictive values were greater than 97.0%. Acon®HRP2 had a higher specificity (96.6%) and lower false-positive (FP) rate (9.3%) compared to Acon®Pf/Pan, which had a specificity of 87.3% and a FP rate of 27.1% (P < 0.01). Overall, 32.5% of all Acon® Pf/Pan tests resulted in a "faint band" with only 2 resulted from samples with a parasitemia below 100 p/μL. The accuracy of Acon®HRP2 RDT for the diagnosis of P. falciparum infection is confirmed. However, the high FP rate observed with Acon®Pf/Pan is a limitation for its use.
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