An IoT-based aptasensor biochip for the diagnosis of periodontal disease.

Severe periodontitis affects nearly 1 billion individuals worldwide, highlighting the need for early diagnosis. Here, an integrated system consisting of a microfluidic chip and a portable point-of-care (POC) diagnostic device is developed using a polymethyl methacrylate (PMMA) chip fabrication and a three-dimensional printing technique, which is automatically controlled by a custom-designed smartphone application to routinely assess the presence of a specific periodontitis biomarker, odontogenic ameloblast-associated protein (ODAM). A sandwich-type fluorescence aptasensor is developed on a microfluidic chip, utilizing aptamer pair (MB@OD64 and OD35@FAM) selectively binding to target ODAM. Then this microfluidic chip is integrated into an automated Internet of Things (IoT)-based POC device, where fluorescence intensity, as a signal, from the secondary aptamer binding to ODAM in a sandwich-type binding reaction on the microfluidic chip is measured by a complementary metal oxide semiconductor (CMOS) camera with a 488 nm light-emitting diode (LED) excitation source. Obtained signals are processed by a microprocessor and visualized on a wirelessly connected smartphone application. This integrated biosensor system allows the rapid and accurate detection of ODAM within 30 min with a remarkable limit of detection (LOD) of 0.011 nM under buffer conditions. Clinical application is demonstrated by successfully distinguishing between low-risk and high-risk individuals with 100 % specificity. A strong potential in the translation of this fluorescence-based microfluidic aptasensor integrated with an IoT-based POC system is expected to be employed for non-invasive, on-site, rapid, and accurate ODAM detection, facilitating periodontitis diagnosis.

Aptamer duo-based portable electrochemical biosensors for early diagnosis of periodontal disease.

Recently, point-of-care tests (POCT) have gained much attention due to their convenient, fast, simple, and easy characteristics. For POCT, portability is an essential feature. In this study, we have successfully fabricated a portable mini-potentiostat. Using chronoamperometry, electrical signals of this portable mini-potentiostat were measured, and the analytical performance of electrochemical aptasensors was compared with a benchtop potentiostat. The electrochemical signals measured by mini-potentiostat can be displayed on the screen of a smartphone. To verify the analytical performance of this portable electrochemical aptasensor platform with a mini-potentiostat, two well-known model protein biomarkers, vaspin, a type 2 diabetes biomarker, and thrombin, a biomarker for pulmonary metastasis and cardiovascular disease, were confirmed to be detected by using corresponding aptamer duo. After solid verification of this portable electrochemical aptasensor platform, we have successfully implemented this portable mini-potentiostat system to develop a portable sandwich-type binding pair of aptamers-based electrochemical biosensor, which can diagnose periodontal disease by measuring ODAM biomarker. The linear range of this ODAM biosensor was 0 to 15 nM with a detection limit of 0.02 nM and 1 nM in buffer and saliva, respectively. The sensitivity of this biosensor has been greatly enhanced, compared to previously developed surface plasmon resonance (SPR) or lateral flow assay (LFA) based aptasensors. This study showed that this new portable aptamer duo-based biosensor is expected to diagnose the early stage of periodontal diseases from real samples, such as saliva or gingival crevicular fluid in a short time as a point-of-care (POC) testing.

Specific detection of avian influenza H5N2 whole virus particles on lateral flow strips using a pair of sandwich-type aptamers.

We report a selection of a cognate pair of aptamers for whole avian influenza virus particles of H5N2 by using graphene-oxide based systemic evolution of ligands by exponential enrichment (GO-SELEX), and the application of a pair of sandwich-type binding aptamers on the lateral flow strips. The aptamers were characterized by GO-FRET assay, and Kd values of the selected aptamers were estimated to be from 6.913 × 105 to 1.27 × 106 EID50/ml (EID50/ml: 50% egg infective dose). Based on the evidence from confocal laser scanning microscope (CLSM), surface plasmon resonance (SPR), and circular dichroism (CD) spectrum analysis, the aptamers, J3APT and JH4APT, were found to be working as a cognate pair that binds to the target virus at the different sites simultaneously. This cognate pair of aptamers then was successfully applied on the lateral flow strips, clearly showing sandwich-type binding images with the presence of the certain numbers of H5N2 virus particles. On the newly developed lateral flow strips, the target virus was detectable down to 6 × 105 EID50/ml in the buffer and 1.2 × 106 EID50/ml in the duck's feces, respectively, by the naked eye. By using the ImageJ software, the LOD was found to be 1.27 × 105 EID50/ml in the buffer and 2.09 × 105 EID50/ml in the duck's feces, respectively. Interestingly, on the lateral flow strips, enhanced specificity towards the target virus (H5N2) appeared over other subtypes of H5Nx. To the best of our knowledge, this is the first report about the application of the cognate pair of aptamers for the detection of influenza virus on the lateral flow strips. This study shows the promising perspective of a cognate pair of aptamers for the on-site detection system which could be useful for rapid detection of avian influenza viruses for preventing the pandemic influenza viruses from spreading.

A new coccolith modified electrode-based biosensor using a cognate pair of aptamers with sandwich-type binding.

In this study, we report a cognate pair of the aptamer-based sandwich-type electrochemical biosensor for type 2 diabetes biomarker (Vaspin) using coccolith modified electrodeposited on the screen-printed gold electrode (CME-SPGE). The coccolith derived from E. huxleyi used in this study were known to be highly-structured microparticles with many nano-sized pores. The CME-SPGE was successfully fabricated by drop-casting coccoliths, followed by Au sputtering and electrodeposition of Au. On this CME-SPGE electrode, the sandwich-type electrochemical aptasensor was fabricated by using a cognate pair of aptamers. The morphological, electrochemical characteristics and the performances of both the CME-SPGE and the completely fabricated sandwich-type aptasensor were investigated by SEM, EDAX, cyclic voltammetry, and chronoamperometry. Due to the synergic effect of a cognate pair of aptamers on CME-SPGE, this newly developed sandwich-type electrochemical biosensor for Vaspin showed high specificity, and good sensitivity with a limit of detection (LOD) of 298 pM, along with more widen the linear range. To the best of our knowledge, this is the first report about the use of a coccolith modified electrode with a cognate pair aptamer resulting in sandwich-type binding in an electrochemical biosensor. With the advantages of using highly-structured biomineral microparticles and a cognate pair of aptamers, this new study may pave the innovative way to design a novel sandwich-type electrochemical aptasensor platform.