This study was designed to assess the relative reliability of microscopy, the dipstick technique based on the detection of Plasmodium falciparum-specific histidine rich protein II (HRPII) (Parasight-F) and PCR assays in diagnosing falciparum malaria infections in Nigerian children. The prevalence of P. falciparum infections in enrolled patients was 100% by microscopy. Parasite density ranged from 329 to 81,194 parasites/microL of blood, with a geometric mean parasite density of 5168 parasites/microL of blood. The sensitivity of the HRPII based dipstick, PCR and microscopy were 80%, 92% and 100% respectively. A false negative rate of 20% was observed with Parasight-F as compared with microscopy. The parasitemia in patients with false negative Parasight-F tests ranged from 319 to 54,680 parasites/microL of blood. Detailed PCR analysis of Isolates obtained from five out of the eight patients who exhibited a negative Parasight-F test, showed that the average numbers of P. falciparum clones in these five isolates were: 1.7 +/- 1.02 with MSPI, 3.2 +/- 1.3 with MSP2 and 1.4 +/- 1.72 with GLURP. Comparison of microscopy and HRPII results showed a significant (p=0.009) difference as opposed to microscopy and PCR (p=0.239). This study showed that caution should be exercised when excluding P. falciparum infections on the basis of HRPII based dipstick results alone. Microscopy or PCR diagnosis where possible, should be carried out in order to confirm negative P. falciparum HRPII-based dipstick tests.