The results of the present study indicate that the Binax NOW® Malaria Test allows a semi-quantitative assessment of parasitaemia in travellers returning with P. falciparum malaria.
HRP-2 reactivity (T1) in absence of aldolase reactivity, proved a reliable predictive marker for a low (< 1%) P. falciparum parasitaemia. In the current study population HRP-2 reactivity without aldolase reactivity was applicable to 30% of returning travellers with P. falciparum. When clinical findings and routine laboratory results in these patients also are not indicative of severe disease, the patient most likely has uncomplicated malaria. This knowledge is important for further clinical decision-making.
Co-reactivity of aldolase and HRP-2 was present in all patients with a parasitaemia above 50,000 asexual parasites/μl (corresponding to approximately 1% infected red blood cells). Others reported co-reactivity with Binax NOW® in 80% of patients with P. falciparum parasitaemia > 40,000 parasites/μl . The authors suggested co-reactivity of HRP-2 and aldolase possibly could function as a semi-quantitative marker of high P. falciparum parasitaemia . In the present study this relation proved, however, not straightforward with co-reactivity of HRP-2 and aldolase also being present in patients with low parasitaemia (i.e. < 0.5%). As such co-reactivity of HRP-2 and aldolase is less reliable as marker for high parasitaemia.
The data of the present study suggest that aldolase and HRP-2 co-reactivity is present in all patients with severe malaria. Apparently HRP-2 and aldolase reactivity, which depends on the load of these antigens in the blood specimen, is preserved because these antigens are derived not only from circulating viable and non-viable malaria parasites, but also from sequestered parasites that are abundantly present in severe malaria. As a consequence of this sequestration, microscopic determination of peripheral blood smears might underestimate the total parasitaemia.
The relation between aldolase reactivity and P. falciparum parasitaemia, as observed with Binax NOW® in this study, could be dependent of the process of manufacturing. Other RDTs with a similar three-band configuration, therefore, should be studied in detail, to assess about the precise relationship between P. falciparum parasitaemia and aldolase reactivity.
HRP-2 and aldolase co-reactivity may also be a feature of a mixed Plasmodium infection. In a recent study of 2,847 cases of imported malaria in the Netherlands , 75% of the infections were solely caused by P. falciparum whereas the remainder was caused by P. vivax (15%), P. ovale (7%), and P. malariae (3%), respectively. Only 0,7% of all infections was attributable to mixtures of species, mostly involving P. falciparum. Thus, in the Dutch setting, HRP-2 and aldolase co-reactivity is far more likely to reflect a mono-parasitic P. falciparum infection rather than a mixed infection.
Results of RDT's may facilitate clinical decision making in patients suspected of having malaria. There are however also some drawbacks to consider. First, these tests cannot replace clinical assessment of the ill-returning patient and results of RDT tests should always be confirmed by thin or thick blood smears, including parasite counts in case of P. falciparum malaria [6, 12]. Second, the diagnostic power of this RDT test is dependent on the epidemiological setting, in particular the prevalence of the disease. The current findings may not simply be extrapolated to regions of malaria endemicity where low-grade malaria infections are far more prevalent and empirical anti-malarial treatment is common use, which may lead to false-negative and false-positive RDT findings, respectively. In addition, the majority of the travellers in this study contracted P. falciparum infection in Africa; other malaria-endemic continents like South-East Asia and South-America were underrepresented. Caution is warranted with extrapolating the applicability of the current findings to imported malaria acquired outside Africa. False negative results have been suggested for certain genetic polymorphisms of HRP-2 geographically confined to the Asia-Pacific region  and for P. falciparum isolates from South America lacking HRP-2 . In addition, false negative test results may occur at high parasitaemia due to a so-called prozone effect, defined as false-negative or false-low results in immunological reactions due to an excess of either antigens or antibodies. The prozone effect was observed for HRP-2 in 16 of 17 RDTs (including the Binax NOW® Malaria Test), resulting in a false low HRP-2 signal, whereas aldolase reactivity was not affected . Finally, the clinician must also consider the possibility of a P. knowlesi infection, which may give rise to severe disease and fatal complications as well . Even though early reports suggested that RDTs may not detect P. knowlesi infections, later studies demonstrated that P. knowlesi was reactive with the aldolase band in the Binax NOW® Malaria Test, but not with HRP-2 and that aldolase reactivity depended on the P. knowlesi parasitaemia .