Clinical Chemistry
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Clinical Chemistry 50: 51-57, 2004; 10.1373/clinchem.2003.020867
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(Clinical Chemistry. 2004;50:51-57.)
© 2004 American Association for Clinical Chemistry, Inc.

Molecular Diagnostics and Genetics

Immuno-PCR for Detection of Antigen to Angiostrongylus cantonensis Circulating Fifth-Stage Worms

Soi-Moi Chye1, Shiu-Ru Lin2, Ya-Lei Chen1, Lee-Yi Chung3 and Chuan-Min Yen3,a

1 Department of Medical Technology, Fooyin University, 151 Chinhsuen Road, Ta-Liao Hsiang, Kaohsiung, Taiwan, ROC.
Departments of2 MedicoGenomic Research Center and3 Parasitology, Kaohsiung Medical University, Shih-Chuan 1st Road, Kaohsiung 807, Taiwan, ROC.

aAuthor for correspondence. Fax 886-7-3218309; e-mail Chmiye{at}cc.kmu.edu.tw.

Background: Definitive diagnosis of infestation with Angiostrongylus cantonensis is difficult because the parasitic nematode is undetectable in the cerebrospinal fluid (CSF) of one-half of afflicted patients and the diagnostic sensitivity of ELISA for circulating worm antigens in patient sera is low. We studied immuno-PCR as a diagnostic tool.

Methods: We studied 30 controls and 60 afflicted patients (30 confirmed by parasitologic analysis of CSF). We used a monoclonal antibody to capture circulating A. cantonensis antigens in serum samples. A DNA label generated by PCR amplification with biotinylated primer was bound by use of streptavidin to a biotinylated third antibody. Circulating antigens sandwiched by monoclonal antibody were detected by PCR amplification of the DNA label.

Results: The detection limit of the ELISA was 100–1000 times higher than that of the immuno-PCR. The concentrations of circulating antigens in patients were markedly higher than those in controls (Wilcoxon rank-sum test, P <0.001). At a cutoff of 0.1 ng/L, sensitivity and specificity for immunodiagnosis of patients with angiostrongyliasis by immuno-PCR were 98% (95% confidence interval, 91–99%) and 100% (93–100%), respectively. The test was positive in all parasitologically confirmed cases.

Conclusions: Immuno-PCR is a promising technique for diagnosis of A. cantonensis infestation.




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