**4. Conclusions**

We developed an automated microfluidic assay integrated with smartphone that can obtain results in ~10 minutes, which can be beneficial in resource-limited settings. There were some shortcomings and further areas for our device to expand on. The sensitivity of our device is much higher than that of other point-of-care devices with a detection limit of 62.5 ng/mL. Further improvement in this area can result in grea<sup>t</sup> potential for this device to be clinically used in areas around the world. However, the cost effectiveness, automation, and time effectiveness compared to traditional ELISA's can be beneficial in regions that struggle with poverty and lack of resources. The utilization of smartphone video and the in-house software can be operated by low-skilled workers. This, in turn, can further the accessibility of the developed device and allow anyone to record and interpret results. Without the use of high-cost specialized instruments, the developed chip can be kept at a low cost. These results show potential that the platform would be useful for the POC diagnosis/screening of Zika virus infection patients and can be used in resource-limited settings.

**Supplementary Materials:** The following are available online at http://www.mdpi.com/2075-4418/10/1/42/s1, Table S1: Material and reagents cost of the disposable microfluidic chip, Figure S1: Reagent Loading sequence inside the microfluidic chip, Figure S2: Chip design, fabrication, and assembly, Figure S3: Sandwich ELISA assay for Dengue 2 NS1 with the capture antibody for Zika NS1 for detection in binding buffer, Figure S4: Zeta potential measurement of antibody coated beads, Video S1: Sample reagen<sup>t</sup> loading and test run.

**Author Contributions:** Conceptualization, M.A.K. and W.A.; methodology, M.A.K., H.Z., M.S, and W.A.; software, M.A.K.; validation, M.A.K., H.Z. and M.S.; formal analysis, M.A.K., H.Z. and, W.A.; investigation, W.A.; resources, M.A.K. and W.A.; data curation, M.A.K. and W.A.; writing—original draft preparation, M.A.K. and H.Z.; writing—review and editing, M.A.K., H.Z., M.S., and W.A.; visualization, M.A.K.; supervision, W.A.; project administration, W.A.; funding acquisition, W.A. All authors have read and agreed to the published version of the manuscript.

**Funding:** This research was funded by National Institute of Health (NIH) gran<sup>t</sup> number R15AI127214, and R56AI138659, and Florida Department of Health Zika gran<sup>t</sup> 7ZK10. The APC was funded by National Institute of Health (NIH) gran<sup>t</sup> number R56AI138659.

**Acknowledgments:** We acknowledge research support from NIH R15AI127214, NIH R56AI138659, Florida Department of Health ZIKA gran<sup>t</sup> (FDOH) 7ZK10, and support from the College of Engineering and Computer Science, Florida Atlantic University, Boca Raton, FL.

**Conflicts of Interest:** The authors declare no conflict of interests.
