Authors

N Bahamontes-RosaA Rodriguez AlejandreV GomezS VieraM G Gomez-Lorenzo L M Sanz-AlonsoA Mendoza-Losana ¹ GSK, Spain

Discussion

Background
Quantitative real-time PCR (qPCR) is now commonly used as a method to confirm diagnosis of malaria and primarily to differentiate recrudescence from re-infection, especially in clinical trials and in reference laboratories where precise quantification is critical. Although antimalarial drug discovery makes use of in vivo murine efficacy models, the use of molecular analysis from the models has been quite limited. The aim of our study was to develop qPCR as a methodology to support pre-clinical antimalarial models making use of material maintained in filter papers for qPCR analysis. Results were compared with traditional methods.
Methods

FTA technology (Whatman) is a rapid and safe method for extracting nucleic acids from infected blood. Peripheral blood samples from mice infected with Plasmodium berghei, P. yoelii, or P. falciparum were kept as frozen samples or as spots on FTA cards. The extracted genetic material from both types of samples was assessed for quantification by qPCR using sets of specific primers specifically designed for Plasmodium 18S rRNA, LDH, and CytB genes.
Results
The optimal conditions for nucleic acid extraction from FTA cards and qPCR amplification were set up, and were confirmed to be suitable for par