*2.5. Cell Encapsulation Studies*

Human dermal fibroblasts (HDF, Thermo Fisher Scientific, Waltham, MA, USA) and Jurkat cells (ATCC, Manassas, VA, USA) were routinely cultured in DMEM and RMPI-1640 medium respectively, in a CO2 incubator. The medium was supplemented with 10% FBS and 1% Pen-Strep. PA66 capsules were prepared as described in Section 2.4. The capsules were saturated in complete medium overnight, and 5.0 × <sup>10</sup><sup>5</sup> cells/mL of exponentially growing cells were loaded into each capsule through the injection port. The injection port was closed tightly with a sterile nylon thread, and they were cultured in a CO2 incubator. After 3 d of incubation, the cell-laden capsules were transferred to a fresh plate with fresh medium. The presence of cells, both inside and outside the capsules, was measured using the metabolic Alamar blue assay, comparing the medium in the spent and receiving plates. In the case of HDF cells, if any cells migrate out of the capsule, they would attach to the bottom of the plate; the spent medium was exchanged directly with the fresh medium. In the case of Jurkat cells, the spent medium was centrifuged to collect cells at the bottom of the well prior to replacing the spent medium with fresh medium. Briefly, 1 mL of fresh medium together with 100μL of Alamar blue reagent was added to both the wells containing the cell-laden capsules and the wells of the spent plate. The contents were mixed well and incubated in a CO2 incubator. After 4 h of incubation, the absorbance was measured at a wavelength of 570 nm, using a 600 nm wavelength as a reference. An equal number of cells cultured in a standard tissue culture treated polystyrene (TCPS) plate, under similar conditions, were treated as the positive control. Cell-free Alamar blue reagent, under similar conditions, was treated as the negative control. The cell viability was measured using the following Equation:

$$\text{Cell viability} \%= \frac{\text{Absorbance of Test Sample}}{\text{Absorbance of Control Sample}} \times 100\tag{1}$$
