*3.1. Microfluidic Chip Embracing a Nanofiber Scaffold*

Figure 3a shows the SEM image of the electrospun nanofiber scaffold for the proposed chip. First, the electrospun fibers were randomly oriented. Moreover, the nanofiber scaffold in this study had various diameter fibers due to the combination effect of the chloroform solvent, with faster evaporation, and the process conditions [30,31]. Some fibers had diameters ranging from several to 10 μm, whereas others appeared fine with diameters in the nanometer scale. Results of detailed analysis of fiber diameter distribution are in Figure 3b. The peak frequency diameter of fine fibers (i.e., nanofibers) was between 600 and 1000 nm, and a significant number of microfibers had a diameter of 2 to 6 μm in the scaffold. This nanofiber scaffold possibly contained larger pores than the scaffold consisting of small diameter nanofibers, because the microfibers can introduce larger pores than nanofibers [32]. Figure 3c shows the differential intrusion curve with respect to pore size of the nanofiber scaffold. From the figure, the pore diameter (equivalently, size) ranged between 0.3 and 100 μm and the peak frequency pore diameter was about 3 μm. The porosity of the nanofiber scaffold was about 76%. Therefore, the fabricated nanofiber scaffold may allow the cells to infiltrate into the deep regions of the scaffold. Figure 4a shows an assembled microfluidic chip embracing a nanofiber scaffold. The developed chip looks simple and is appropriate for use in a real PDMS fabrication environment. The microchannel of the chip had a width of 307.6 ± 4.5 μm and depth of 97.2 ± 4.9 μm, as shown in Figure 4b. The microchannel had a flat and smooth bottom plane, while its side walls had a rough surface because of the cut edge of tapes. However, the rough wall surface of the microchannel did not affect the device operation quality because the microchannel engaged only in the delivery of culture medium with slow velocity.

**Figure 3.** Electrospun nanofiber scaffold: (**a**) SEM images; (**b**) diameter distribution of the electrospun fibers; (**c**) porosity analysis result.

**Figure 4.** Microfluidic chip with a nanofiber scaffold (**a**) and SEM images of the microchannel of the chip (**b**).
