| Summary: | <p>Abstract</p> <p>Background</p> <p><it>Plasmodium berghei</it> rodent malaria is a well-known model for the investigation of anti-malarial drug efficacy <it>in vivo</it>. However, the availability of drug <it>in vitro</it> assays in <it>P. berghei</it> is reduced when compared with the spectrum of techniques existing for <it>Plasmodium falciparum</it>. New alternatives to the current manual or automated methods described for <it>P. berghei</it> are attractive. The present study reports a new ELISA drug <it>in vitro</it> assay for <it>P. berghei</it> using two monoclonal antibodies against the parasite lactate dehydrogenase (pLDH).</p> <p>Methods</p> <p>This procedure includes a short-<it>in vitro</it> culture, the purification of schizonts and the further generation of synchronized mice infections. Early stages of the parasite are then incubated against different concentrations of anti-malarial drugs using micro-plates. The novelty of this procedure in <it>P. berghei</it> relies on the quantification of the drug activity derived from the amount of pLDH estimated by an ELISA assay using two monoclonal antibodies: 14C1 and 19G7. The IC<sub>50</sub>s obtained through the ELISA assay were compared with those from the micro-test.</p> <p>Results</p> <p>The initial parameters of the synchronized samples used in the <it>in vitro</it> assays were a parasitaemia of 0.5% and haematocrit of 1%, with an incubation period of 22 hours at 36.5°C. pLDH detection using a 14C1 coating at 10 μg/ml and 19G7 at 2.5 × 10<sup>-3</sup> μg/ml provided good readouts of optical densities with low background in negative controls and specific detection levels for all parasite stages. IC<sub>50</sub>s values derived from the ELISA assay for artesunate, chloroquine, amodiaquine and quinine were: 15, 7, 2, and 144 nM, respectively. When artesunate and chloroquine IC<sub>50</sub>s were evaluated using the micro-test similar values were obtained.</p> <p>Conclusion</p> <p>This ELISA-based <it>in vitro</it> drug assay is easy to implement, fast, and avoids the use radioisotopes or expensive equipment. The utility of this simple assay for screening anti-malarial drug activity against <it>P. berghei in vitro</it> is demonstrated.</p>
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