Multiple DNAs And α-amylase Detection Using Portable Glucose Meter

Traditional clinical assay of many analytes often need to be converted into thedetection of few indicators， such as hydrogen peroxide (H2O2) or nicotinamideadenine dinucleotide (NADH). These traditional methods often share the drawbacksof expensive instrument, tedious procedures and high assay costs. In order to developsimple and low-cost methods, new signal transduction means and portable deviceswere developed for clinical diagnosis and self monitoring. Recently, the portablepersonal glucose meter could detect a series of targets by using glucose as indicator,which provided a new means for traditional clinical assay. However, all of the worksbased on personal glucose meter can only detect single target and take long time fordetermination. In this paper, we construct rapid and sensitive methods to detectmultiple DNAs and α-amylase by using personal glucose meter. The main researchesare as followed:1. A new method for multiplex detection of nucleic acids was developed based ona personal glucose meter and a microfluidic chip. In this system, a microfluidic chipwith multiple channels was fabricated firstly. Then capture probes of differentgenotypes of Hepatitis B virus (HBV) DNA were respectively immobilized on thesurface of each branch channels. When sample solution was added into the chip, itwas seperated into four branch channels automatically. In the presence of HBV DNA,it hybridized with capture probes. After high-stringency washing buffer wasintroduced, mismatched hybrids between HBV DNA and noncomplementary captureprobes were removed. Then, invertase-DNA conjugate was added and subsequentlycaptured by the HBV DNA, forming a “sandwich” hybridization structure. Thus,invertase was immobilized on the surface of branch channel. Upon addition of sucrose,the bound invertase can catalyze the hydrolysis of sucrose into glucose. Finally, byanalysing the different PGM signal values obtained from different channels, theconcentration of targets can be determined simultaneously. Furthermore, this protocolcould be altered for the detection of protein, small molecule and other targets.2. A one-step method for α-amylase detection using personal glucose meter. In thissystem, maltopentaose was used as substrate for sensitive detection of α-amylase bycoupling with α-glucosidase. In the presence of α-amylase, maltopentaose washydrolyzed into maltotriose and maltose. Then α-glucosidase catalyze hydrolysis ofmaltotriose and maltose into glucose subsequently. The released glucose can be quantitatively monitored using a portable personal glucose meter. Under optimalconditions, the proposed method can detect α-amylase in15minutes. For highconcentration α-amylase (700U/L), signal can be detected in5minutes. In addition,this method also exhibits excellent selectivity and could applied to the determinationof α-amylase in human serum and urine. Considering the compact size, simpleoperation, our approach holds great promise for further applications in the field ofPOCT clinical diagnosis.