Quantitative electrolateral flow immunosensor (ELLI^™): Opening new diagnostic frontiers such as dengue triage and secondary stroke prognosis

The development of a rapid, affordable, and sensitive diagnostic kit for point-of-care is important in most healthcare settings; however, only a few exhibit the needed quantitative capability. Our technology, ELLI^™, is a patented diagnostic device that aims to provide on-site quantitation by utilizing proprietary electroactive immunonanoparticles that bind to the target biomarker and subsequently move along toward the biofunctionalized screen-printed gold electrodes (SPGE) to generate an amperometric signal. The SPGE functions simultaneously as a signal transducer and a solid-state support for a sandwich immunoassay. The successful immunocomplex formation is then recorded electrochemically using a potentiostat, whereby the signal was contributed by the presence of ferrocene or a more hydrophilic redox label, radical TEMPO, immobilized on the formulated nanoparticles. A bifunctional ligand, thiolated polyethylene glycol (PEG-thiol), was used to stabilize 20 nm gold colloidal nanoparticles (AuNPs). The ligand was incorporated to not only prevent the salt-mediated AuNPs aggregations but also provide an anchor for antibody and redox species conjugation. To-date, we have miniaturized a 3D-printed prototype device able to sensitively detect and quantify dengue NS1 protein with only 0.6 u L human clinical serum samples diluted in a volume ratio of 1:100 in PBS diluent in less than 30 min with a simple cyclic voltammetry analysis. Other applications are being developed, including stroke prognosis using NT-proBNP biomarker, that is significantly associated with cardioembolic stroke and secondary stroke reoccurrence, with sensitivity >90% and specificity >80%. Initial results of this research in mock porcine plasma have shown that the label-free impedimetric electrochemical biosensor was capable of differentiating such concentrations in a low concentration range, especially among 0, 0.1, 0.5, 1, and 3 ng mL^-1 within 25 min. This range is valuable not only for classifying cardioembolic stroke (>=0.5 ng mL^-1) but also predicting the risk of secondary stroke reoccurrence (>0.255 ng mL^-1). We are now adapting these results to the ELLI^™ platform. The applications above need quantitation as a measure of severity and a classification means. In dengue, for example, a simple, 1-2 step, point-of-careformat is recommended for disease surveillance in communities as a first line of defense, especially when there is no specific treatment for dengue/severe dengue. Meanwhile, in stroke prognosis, a quantitative point-of-care device that reduces the delays in IV-tPA administration and predicts risks as well as secondary reoccurrence will be highly favored. ELLI^™ promises to deliver sensitive measurements of the target biomarker and acts as a diagnostic intervention for faster therapeutics administration that eventually will contribute to a better disease emergency preparedness, such as in trauma triage (internal bleeding) for soldiers. ELLI^™ is the answer to whichever applications that would require quantitation of biomarkers or pathogens at the point of care such as for HAZMAT teams, the military, or home use.