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Validation of a Diagnostic Multiplex Polymerase Chain Reaction Assay for Infectious Posterior Uveitis
Humeyra Dabil, MD;
Michelle L. Boley, BS;
Therese M. Schmitz, BS;
Russell N. Van Gelder, MD, PhD
Arch Ophthalmol. 2001;119:1315-1322.
Objective To valide a multiplex polymerase chain reaction (PCR) assay capable
of simultaneously screening vitreous biopsy specimens for a panel of common
pathogens in posterior uveitis.
Methods A multiplex PCR assay using novel primer sets for cytomegalovirus (CMV),
herpes simplex virus (HSV), varicella zoster virus (VZV), and Toxoplasma gondii was developed. The sensitivity of the assay was determined
for purified pathogen DNA. Twenty-one vitreous specimens from patients with
posterior uveitis were tested by both multiplex and monoplex PCR.
Results Fewer than 10 genomes of VZV and fewer than 100 genomes of HSV, CMV,
and T gondii could be detected using the new primer
sets. When used in multiplex, the assay lost less than 1 log of sensitivity.
Monoplex PCR detected pathogen DNA in 18 of 21 patient samples; multiplex
PCR detected pathogen DNA in 15 of the 18 samples positive by monoplex PCR.
None of 10 negative control samples were positive for pathogen DNA.
Conclusions Multiplex PCR has adequate sensitivity to simultaneously screen a substantial
differential diagnosis for posterior uveitis in a single reaction, without
loss of specificity. This assay may reduce the time and cost involved in PCR-based
molecular diagnostics of infectious pathogens.
Clinical Relevance Mutiplex PCR may allow rapid diagnosis of infectious posterior uveitis.
From the Departments of Ophthalmology and Visual Sciences (Drs Dabil
and Van Gelder and Mss Boley and Schmitz) and Molecular Biology and Pharmacology
(Dr Van Gelder), Washington University School of Medicine, St Louis, Mo. None
of the authors has a financial interest in any of the technologies discussed
in this article.
THIS ARTICLE HAS BEEN CITED BY OTHER ARTICLES
Real-Time Quantitative Polymerase Chain Reaction Diagnosis of Infectious Posterior Uveitis
Dworkin et al.
Arch Ophthalmol 2002;120:1534-1539.
ABSTRACT
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