Efficiency of Automated Viral RNA Purification for Pediatric Studies of Dengue and Zika in Hyperendemic Areas

Figure 4

Use of the RNA purified automatically in the serotype-specific detection assays for DENV. (a) Detection of DENV-1 (left panel, molecular weight 482 bp) and DENV-2 (right panel, molecular weight 119 bp) by conventional RT-PCR on clinical plasma samples from 12 pediatric patients with DENV infection (DENV-1 = 7 and DENV-2 = 5) and 8 negatives for DENV infection. RNA was isolated with the high-throughput protocol (upper) and standard (down). The figure shows four negative samples (for high-throughput) in the upper images and 5 positives (for standard protocol) in the lower images, as other samples were run in a separate gel. C+: supernatant mixture of Vero-76 cells infected with each of the DENV serotypes. C−: mock. (b) Amplification curves of DENV-1 (in blue) and DENV-2 (in green) by RT-qPCR on automatically purified RNA from the plasma of naturally infected patients using the high-throughput (upper) and the standard (down) protocols. The curve generated by the respective cell cultured DENV (control+) and a mock of each condition is shown. The value taken as positive corresponded to a sample with a C_t ≤ 38.

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