Bioluminescence DNA hybridization assay for Plasmodium falciparum based on the photoprotein aequorin.
A bioluminescence DNA hybridization assay for the detection of Plasmodium falciparum, the most deadly species of malaria, using the photoprotein aequorin as a bioluminescent label has been developed. The current gold standard for the detection of malaria is light microscopy, which can detect down to approximately 50 parasites/microL of blood, but has low-throughput, high costs, and requires high skill, which limit the applicability of the method, especially in the developing regions where malaria detection is mostly needed. The utilization of aequorin as a bioluminescence label offers the advantages of high signal-to-noise ratio and reliable detection down to attomole levels, allowing for the development of highly sensitive and miniaturized high-throughput bioluminescence assays. Herein, we developed a DNA hybridization assay for the detection of P. falciparum based on the competition between the target DNA and the signal generating DNA streptavidin-aequorin for hybridization with the probe DNA. This bioluminescence hybridization assay demonstrated a detection limit of 3 pg/microL and was employed for the detection of target DNA in standard and spiked human serum samples. The DNA hybridization assay was developed in a microplate format without the need for sample PCR amplification, showing the potential suitability of this method in the parallel analysis of samples by low-trained personnel, such as that typically encountered in developing regions.
Doleman
Leslie
L
Department of Chemistry, University of Kentucky, Lexington, Kentucky 40506-0055, USA.
Davies
Logan
L
Rowe
Laura
L
Moschou
Elizabeth A
EA
Deo
Sapna
S
Daunert
Sylvia
S
D000331:Aequorin / Q000032:analysis* / Q000235:genetics*; D000818:Animals; D015342:DNA Probes / Q000032:analysis* / Q000235:genetics*; D016054:DNA, Protozoan / Q000032:analysis* / Q000235:genetics*; D008163:Luminescent Measurements / Q000379:methods*; D009690:Nucleic Acid Conformation; D010963:Plasmodium falciparum
D015342:DNA Probes; D016054:DNA, Protozoan; D000331:Aequorin