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Published on: March 13, 2018
Preparation of MS2-based nanoparticles as control and standard materials for the molecular detection of dengue virus
Yu Fu1, Guojing Wang2, Qisheng Wu3
1National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology, Peking University Fifth School of Clinical Medicine, Beijing, People's Republic of China; Graduate School, Peking Union Medical College, Chinese Academy of Medical Sciences, National Center for Clinical Laboratories, Beijing Hospital, National Center of Gerontology, Beijing, People's Republic of China; Department of Nuclear Medicine, the First Affiliated Hospital of Nanjing Medical University, Nanjing, People's Republic of China.
Abstract:
To quantify dengue virus (DENV) and evaluate the performance of clinical laboratories using quantitative real-time polymerase chain reaction (qRT-PCR) assays, we constructed high-efficiency expression systems to produce DENV-1 to 4 nanoparticles and assessed their suitability as standard and control materials in 20 laboratories across China. Targeted gene sequences of DENV-1 to 4 were synthesized and inserted into pACYC-Duet 1-MS2 recombinant plasmids to generate corresponding nanoparticle expression systems. After collection, verification, and quantification by digital PCR (dPCR), DENV-1 to 4 nanoparticles were prepared as control and standard materials. Five positive and three negative samples of each DENV serotype in every panel were used for assessing the performance of the participating laboratories across China, as well as standard materials for the quantitative detection of DENV using qRT-PCR assays. The accuracy, sensitivity, and specificity of qRT-PCR used by the 20 evaluated laboratories were 89.6 (569/635), 85.1 (336/395), and 97.1% (233/240), respectively. Overall, sixteen (80.0%) laboratories were qualified in detecting DENV, among which five (25.0%) were designated as "competent", eleven (55.0%) were defined as "acceptable", and four (20%) were considered to be "improvable". The results generated from the DENV standard samples were significantly positively correlated with those generated by dPCR (r2=0.8698, P<0.001). In summary, DENV nanoparticles could potentially be used as controls for improving the performance of laboratories and as standards for the quantitative detection of DENV.
