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The Effect of Storage and Extraction Methods on Amplification of Plasmodium falciparum DNA from Dried Blood Spots
- Source :
- American Journal of Tropical Medicine and Hygiene, 92, 922-5, American Journal of Tropical Medicine and Hygiene, 92, 5, pp. 922-5
- Publication Year :
- 2015
- Publisher :
- American Society of Tropical Medicine and Hygiene, 2015.
-
Abstract
- Contains fulltext : 154115.pdf (Publisher’s version ) (Open Access) Extraction and amplification of DNA from dried blood spots (DBS) collected in field studies is commonly used for detection of Plasmodium falciparum. However, there have been few systematic efforts to determine the effects of storage and extraction methods on the sensitivity of DNA amplification. We investigated the effects of storage conditions, length of storage, and DNA extraction methods on amplification via three PCR-based assays using field samples and laboratory controls. Samples stored as DBS for 2 or more years at ambient temperature showed a significant loss of sensitivity that increased with time; after 10 years only 10% samples with parasite densities > 1,000 parasites/muL were detectable by nested polymerase chain reaction (PCR). Conversely, DBS and extracted DNA stored at -20 degrees C showed no loss of sensitivity with time. Samples with low parasite densities amplified more successfully with saponin/Chelex compared with spin-column-based extraction, though the latter method performed better on samples with higher parasite densities stored for 2 years at ambient temperature. DNA extracted via both methods was stable after 20 freeze-thaw cycles. Our results suggest that DBS should be stored at -20 degrees C or extracted immediately, especially if anticipating 2 or more years of storage.
- Subjects :
- Time Factors
Plasmodium falciparum
Biology
Polymerase Chain Reaction
Sensitivity and Specificity
Specimen Handling
law.invention
chemistry.chemical_compound
law
Virology
Freezing
Parasite hosting
Malaria, Falciparum
Polymerase chain reaction
Models, Statistical
Spots
Extraction (chemistry)
Temperature
Articles
DNA, Protozoan
biology.organism_classification
DNA extraction
Molecular biology
lnfectious Diseases and Global Health Radboud Institute for Health Sciences [Radboudumc 4]
Infectious Diseases
chemistry
Parasitology
Nested polymerase chain reaction
DNA
Subjects
Details
- ISSN :
- 14761645 and 00029637
- Volume :
- 92
- Database :
- OpenAIRE
- Journal :
- The American Journal of Tropical Medicine and Hygiene
- Accession number :
- edsair.doi.dedup.....dc7099a97ff1ea97f58c39bf742be034