An Integrated System Using Immunomagnetic Separation, Polymerase Chain Reaction, and Colorimetric Detection for Diagnosis of Plasmodium falciparum

Naowarat Seesod Department of Biochemistry and Biotechnology, Royal Institute of Technology, Malaria Center, Region 5, Division of Parasitology, National Institute for Medical Research, Department of Biology, Faculty of Science, Chulalongkorn University, Stockholm, Sweden

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Pornthip Nopparat Department of Biochemistry and Biotechnology, Royal Institute of Technology, Malaria Center, Region 5, Division of Parasitology, National Institute for Medical Research, Department of Biology, Faculty of Science, Chulalongkorn University, Stockholm, Sweden

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Anders Hedrum Department of Biochemistry and Biotechnology, Royal Institute of Technology, Malaria Center, Region 5, Division of Parasitology, National Institute for Medical Research, Department of Biology, Faculty of Science, Chulalongkorn University, Stockholm, Sweden

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Anthony Holder Department of Biochemistry and Biotechnology, Royal Institute of Technology, Malaria Center, Region 5, Division of Parasitology, National Institute for Medical Research, Department of Biology, Faculty of Science, Chulalongkorn University, Stockholm, Sweden

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Sodsri Thaithong Department of Biochemistry and Biotechnology, Royal Institute of Technology, Malaria Center, Region 5, Division of Parasitology, National Institute for Medical Research, Department of Biology, Faculty of Science, Chulalongkorn University, Stockholm, Sweden

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Mathias Uhlen Department of Biochemistry and Biotechnology, Royal Institute of Technology, Malaria Center, Region 5, Division of Parasitology, National Institute for Medical Research, Department of Biology, Faculty of Science, Chulalongkorn University, Stockholm, Sweden

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Joakim Lundeberg Department of Biochemistry and Biotechnology, Royal Institute of Technology, Malaria Center, Region 5, Division of Parasitology, National Institute for Medical Research, Department of Biology, Faculty of Science, Chulalongkorn University, Stockholm, Sweden

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An integrated system for sample preparation and DNA detection of the malaria parasite using immunomagnetic separation in combination with the polymerase chain reaction (PCR) and colorimetric analysis is described. A cocktail of three monoclonal antibodies towards merozoite surface antigen-1 was used for magnetic capture of parasites from microliter amounts of whole blood. A sensitivity down to a parasitemia of 10-6% was achieved using cultured parasites as a model. The integrated approach was evaluated in a field study. A total of 410 blood samples from patients attending malaria clinics in Trat province and Kanchanaburi province in Thailand were analyzed. The samples were processed by immunomagnetic separation and transferred to central laboratory for PCR-based detection. Microscopic examinations on blood smears were done in parallel; 53% were positive using the DNA-based assay, while only 32% were positive using conventional microscopic analysis. This field study suggests a possible model for large-scale testing of malaria with an increased sensitivity compared with conventional methods.

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