Figure 7

Assessment of DDZ-AuNP specificity. A) DENV-2 and CHIKV (1 × 106/mL each) were placed in a buffered solution containing 10 mM MgCl2, 1 × 105 DDZ-AuNP particles/mL, 1.5 M NaCl, and 0.5% (w/v) SDS. Eppendorf tubes containing these mixes were incubated at 37°C for 30 minutes and photographed. CHIKV = chikungunya virus. DENV-2 = dengue virus serotype 2, DDZ-M = anti-dengue virus DNAzyme, DDZin-M = inactive anti-dengue virus DNAzyme. B) Analysis of DENV detection by UV/Vis Spectrophotometry. Samples were assembled as was performed for in A, mixed, incubated at 37°C for 5 minutes, and spectrophotometric analysis was performed using the ND-1000 spectrophotometer. C) Detection of DENV by DDZ-M-AuNP in comparison to several other flaviviruses. The specificity of our DDZ-M-AuNP device to detect DENV and not other fellow flavivirus members YFV, JEV or ZV was determined as described earlier (see Figure 4). D) An alignment was performed on consensus sequences of each of the four DENV serotypes to determine the most optimal regions for the design of serotype specific DDZ-AuNP devices by determining the region of least conservation one serotype has with the other DENV serotypes. E) Colorimetric serotype-specific detection of DENV. Cell culture supernatants from C6/36 cells mock infected (Mock), or from cells infected with either DENV serotypes 1 through 4 or CHIKV (1 × 106/mL each) were placed in buffered solutions containing the necessary cofactors and AuNPs tethered with our all purpose DENV serotype specific DNAzyme, DDZ-M, or one of the serotype-specific DDZs (designated DDZ-1 through-4) designed to specifically target the corresponding DENV serotype (Table 1). Eppendorf tubes containing these mixes were incubated at 37°C for 5 minutes and photographed.