A trial with 80 subjects in each arm would have at least 80% power (using a significance level of 0.10) to detect a difference between the two combinations if the failure rate detected after treatment is 20% in the AS/SP arm (based on the reported failure of SP of approximately 15 – 20% in Sudan) and 5% in AL arm, allowing for 10% loss to follow-up.
Study population and field survey
The study was carried out in three villages in eastern Sudan, southwest of Gedarif town (Abunaja Juama, Abunaja Bargo and Asar). A community-based survey was conducted during the transmission seasons (October to December) of 2004 and 2005 to identify symptomatic patients that were microscopically diagnosed with mono infection of P. falciparum. Patients that satisfied the enrolment criteria according to the WHO protocol for the assessment and monitoring of antimalarial drug efficacy for the treatment of uncomplicated falciparum malaria  were recruited after they provided written informed consent to participate. However, due to the low endemicity of malaria in this area all age groups were recruited. Patients were randomly allocated to receive either AS/SP or AL by a simple random technique of a hat draw. Pregnant women were not enrolled as per the national protocol for uncomplicated malaria recommendations.
Finger prick blood samples were collected and thick and thin smears were prepared on a patient's first attendance. When the patient was enrolled to the study, a finger prick sample was collected on days 0, 1, 2, 3, 7, 14, 21, and 28 on a glass slide and a parallel drop of blood was collected on filter paper. The filter paper was air-dried and stored in self-sealing plastic bags for molecular analysis.
Thick and thin blood smears were stained with Giemsas' stain for ten minutes. The slides were read by two independent experienced microscopists. Parasite count was performed by counting asexual parasites against 200 white blood cells (WBC), and parasite density calculated assuming an average of 6,000 WBC in the population. Ten percent of the slides were read by a third microscopist in Khartoum for quality control. In case of disagreement the majority result was taken.
Treatment and follow up schedule
Each patient was allocated randomly to either AS/SP or AL, treatment was administered under supervision for the AS/SP group, while only one dose of the AL was supervised and the second dose was given to the patient for self-administration. Patients were asked to return to the clinic on days 1, 2, 3, 7, 14, 21, and 28 or whenever they did not feel well. When a patient failed to come to the clinic on any scheduled day, a member of the team visited the home, gave them the dose and obtained the sample unless the patient was not found or refused to give the sample. Those who refused to give their sample (withdrawn) or missed more than one day of follow-up were excluded from the analysis.
Patients, who were positive by microscopy and refused to participate in the study or did not meet the enrolment criteria, were provided with AS/SP for self-administration whereas pregnant women were referred to Gedarif Teaching Hospital as recommended by the national protocol.
In vivo analysis
A per protocol analysis was performed employing the WHO excel sheet .
Samples that were positive by microscopy on any of the follow-up days were classified as early treatment failure (ETF), late parasitological failure (LPF), or late clinical failure (LCF). Adequate clinical and parasitological response (ACPR) was defined as complete absence of microscopically detectable parasitaemia during the follow-up period, according to the protocol .
Samples that were classified as ETF, LCF or LPF were genotyped by PCR as previously described for msp-1  and msp -2 [13, 14] on day 0, 1, 2, 3 and failure (day 7 to 28) to differentiate recrudescence from reinfection. Unpublished observations indicate the necessity of day 0 to day 3 sampling to accurately identify recrudescence from re-infection.
The study was reviewed and ethical clearance was obtained from the National Ethical Committee, Federal Ministry of Health, Sudan. Individual written informed consent was obtained from all participants or their legal guardians.