In this device, Y shaped inlet channels with backflow-prevented microstructure and detection chambers were designed for an efficient immunoassay as shown in Figure 2 . The key features of microstructures in microfluidic device were successfully replicated using the microinjection molding, and they were confirmed through top and tilted scanning electron microscopy (SEM, Hitachi S4800, Ibaraki, Japan) images as shown in Figure 2(C-F) . Due to the height differences between inlet channels and main channels (Figure 2(C,D) ), this could prevent the backflow of the solutions. This device is composed of three detection chambers with 1 mm in diameter for the further immobilization of GBP-H1a fusion protein as shown in Figure 2 , and these chambers may reduce the errors during analysis by averaging signals. All protuberant microstructures near to the microchannels were specially designed as welding lines. During the ultrasonic bonding process to bond the top and bottom of the microfluidic chip, these lines would be melted by concentrating the ultrasonic energies on the top of welding lines. To investigate the sensing window of the fusion protein, different concentrations (6.25, 12.5, 25, 50, 100 and 200 μg/mL, respectively) of GBP-H1a fusion protein were immobilized onto the gold chip surface by surface plasmon resonance (SPR) microfluidics. A greater shift in resonance unit (RU) was observed by SPR analysis with increasing concentration of immobilized GBP-H1a fusion protein bound on the planar surface at various concentrations as shown in Figure 3(A) . These results suggest that the SPR sensor with GBP-fusion protein implemented on the gold surface can be an effective system for biomolecular immobilization. Furthermore, the concentration of GBP-H1a fusion protein was fixed to 100 μg/mL due to its best immobilization concentration.