A disposable microfluidic amperometric dual-sensor was developed for the detection of glycated hemoglobin (HbAic) and total hemoglobin (Hb), separately, in a finger prick blood sample. The accurate level of total Hb was determined through the measurements of the cathodic currents of total Hb catalyzed by a toluidine blue O (TBO)-modified working electrode. Subsequently, after washing unbound Hb in the fluidic channel of dual sensor with PBS, the cathodic current by only HbA(1c) captured on aptamer was monitored using another aptamer/TBO-modified working electrode in the channel. To modify the sensor probe, poly(2,2' :5 ',5"-terthiophene-3 -p-benzoic acid) and a multi-wall carbon nanotube (MWCNT) composite layer (pTBA@ MWCNT) was electropolymerized on a screen printed carbon electrode (SPCE), followed by immobilization of TBO for the total Hb probe and aptamer/TBO for the HbAic probe, respectively. The characterization of each sensor surface was performed using cyclic voltammetry (CV), electrochemical impedance spectroscopy (EIS), Xray photoelectron spectroscopy (XPS), quartz crystal microbalance (QCM), field-emission scanning electron microscopy (FE-SEM), and transmission electron microscopy (TEM). The experimental conditions affecting the analytical signal were optimized in terms of the amount of TBO, pH, temperature, binding time, applied potential, and the content ratio of monomer and MWCNT. The dynamic ranges of Hb and HbAlc were from 0.1 to 10 M and from 0.006 to 0.74 mu M, with detection limits of 82(+/- 4.2) nM and 3.7(+/- 0.8) nM, respectively. The reliability of the proposed microfluidic dual-sensor for a finger prick blood sample (1 mu L) was evaluated in parallel with a conventional method (HPLC) for point-of-care analysis.
