TY - JOUR
T1 - Development of a quantitative, competitive polymerase chain reaction-enzyme-linked immunosorbent assay for the detection of Wuchereria bancrofti DNA
AU - Fischer, Peter
AU - Liu, Xiaole
AU - Lizotte-Waniewski, Michelle
AU - Kamal, Ibrahim H.
AU - Ramzy, Reda M.R.
AU - Williams, Steven A.
N1 - Funding Information:
Acknowledgements The authors would like to thank Dr. T. B. Nutman for the supply of W. bancrofti microfilariae. This study was supported by a scholarship from the German Academic Exchange (DAAD) to P.F. and by grants from the National Academy of Sciences Blakeslee Fund for Genetics Research and the WHO to S.A.W.
PY - 1999
Y1 - 1999
N2 - A quantitative, competitive polymerase chain reaction (QC-PCR) assay for the sensitive detection of Wuchereria bancrofti DNA was developed. A competitor sequence was constructed by an exchange of nucleotides in the Wuchereria-specific Ssp I repeat. The PCR products were hybridized to specific DNA probes and their amounts, determined by an enzyme-linked immunosorbent assay (ELISA). In laboratory-prepared samples the QC-PCR-ELISA assay was capable of detecting the amount of DNA equivalent to 0.1 microfilaria (mf) added to 200 μl of blood lysate. The assay was also tested on 78 blood samples collected in endemic areas in Egypt. All 28 samples that were positive both for mf and for circulating antigen were also QC-PCR-ELISA-positive. In addition, one mf-negative but antigen-positive sample was also positive as determined by QC-PCR-ELISA. A positive correlation of mf density with the QC-PCR-ELISA was observed. Samples containing 10 or fewer mf/ml had a mean relative amount of Ssp IPCR product of 19.7 units, whereas samples with 11-100 mf/ml had a mean of 36.3 units and those with more than 100 mf/ml had a mean of 84.6 units. Because of the high standard deviation within each group, estimates of worm burdens in infected individuals using the QC-PCR-ELISA are not recommended. However, we present data indicating that the W. bancrofti QC-PCR-ELISA is a powerful new tool for evaluation of parasitic loads for community-based diagnosis of bancroftian filariasis.
AB - A quantitative, competitive polymerase chain reaction (QC-PCR) assay for the sensitive detection of Wuchereria bancrofti DNA was developed. A competitor sequence was constructed by an exchange of nucleotides in the Wuchereria-specific Ssp I repeat. The PCR products were hybridized to specific DNA probes and their amounts, determined by an enzyme-linked immunosorbent assay (ELISA). In laboratory-prepared samples the QC-PCR-ELISA assay was capable of detecting the amount of DNA equivalent to 0.1 microfilaria (mf) added to 200 μl of blood lysate. The assay was also tested on 78 blood samples collected in endemic areas in Egypt. All 28 samples that were positive both for mf and for circulating antigen were also QC-PCR-ELISA-positive. In addition, one mf-negative but antigen-positive sample was also positive as determined by QC-PCR-ELISA. A positive correlation of mf density with the QC-PCR-ELISA was observed. Samples containing 10 or fewer mf/ml had a mean relative amount of Ssp IPCR product of 19.7 units, whereas samples with 11-100 mf/ml had a mean of 36.3 units and those with more than 100 mf/ml had a mean of 84.6 units. Because of the high standard deviation within each group, estimates of worm burdens in infected individuals using the QC-PCR-ELISA are not recommended. However, we present data indicating that the W. bancrofti QC-PCR-ELISA is a powerful new tool for evaluation of parasitic loads for community-based diagnosis of bancroftian filariasis.
UR - http://www.scopus.com/inward/record.url?scp=0032946196&partnerID=8YFLogxK
U2 - 10.1007/s004360050531
DO - 10.1007/s004360050531
M3 - Article
C2 - 9951959
AN - SCOPUS:0032946196
VL - 85
SP - 176
EP - 183
JO - Parasitology Research
JF - Parasitology Research
SN - 0044-3255
IS - 3
ER -