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STORAGE DURATION AND POLYMERASE CHAIN REACTION DETECTION OF PLASMODIUM FALCIPARUM FROM BLOOD SPOTS ON FILTER PAPER

To evaluate the effect of long-term storage of sample filters on the sensitivity of polymerase chain reaction (PCR) detection of malaria, 252 blood spots from patients with microscopically confirmed Plasmodium falciparum malaria were analyzed and stratified by storage duration. The spots were collected between 1996 and 2000 on filter paper and stored at room temperature. A Chelex-based method was used to extract the DNA. Unexpectedly, after the first purification, the sensitivity of the PCR from recently stored samples was low and showed progressively increased with time storage (P = 0.003, by chi-square test for linear trend). This suggested that PCR inhibitors were easier to dissolve from the more recent blood spots (< 4 years old) than from blood spots 4 years old, thus leading to a time-dependent increase in PCR sensitivity. However, if DNA was purified again (when the first PCR result was negative), the cumulative sensitivity was not influenced by storage duration. This indicated that length of storage is not a critical issue providing purification is sufficient.

Received November 18, 2002. Accepted for publication April 22, 2003.

Acknowledgments: We thank Bongkot Soonthornsata (Faculty of Tropical Medicine, Mahidol University) and Rujada Wanotayan and Arporn Hongyantarachai (Queen Sirikit National Institute of Child Health, Bangkok, Thailand) for their kind assistance.

Financial support: S. Chaorattanakawee and J. Patarapotikul were supported by the 2000 Royal Golden Jubilee Ph.D. research grant from the Thailand Research Fund. Part of the project was supported by a Mahidol University grant.

Authors’ addresses: S. Chaorattanakawee, O. Natalang, H. Hananantachai, M. Nacher, S. Looareesuwan, and J. Patarapotikul, Faculty of Tropical Medicine, Mahidol University, 420/6 Rajvithi Road, 10400 Bangkok, Thailand. A. Brockman, Shoklo Malaria Research Unit, 736/2 Intrakiri Road, PO Box 46, Mae Sot, 63110 Tak, Thailand, Telephone/Fax: 66-2-55-531-531, E-mail: shoklo{at}cscoms.com.

Reprint requests: J. Patarapotikul, Faculty of Tropical Medicine, Mahidol University, 420/6 Rajvithi Road, 10400 Bangkok, Thailand, Telephone: 66-2-246-9000, extension 1668, Fax: 66-2-246-8340, E-mail: tmjpt{at}mahidol.ac.th.

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