Glass slide

Unless otherwise stated, all materials were purchased from commercial suppliers and were used as received. Glass substrates (glass slides, 76 × 26 mm) were purchased from Marienfeld-Superior. Indium tin oxide-coated glass slides (15–25 Ω sq⁻¹, 75 × 25 mm) were purchased from Aldrich. Poly(ethylene terephthalate) films (thickness: 100 μm) for use as flexible and transparent substrates were purchased from Film Bank. Flexible graphite foils (thickness: 130 μm) were purchased from Dongbang Carbon.

Three 30-μm coronal sections at the levels of 1.7 mm, 0.7 mm, −0.3 mm, −1.3 mm, −2.3 mm, and −3.3 mm from bregma were cut and mounted on glass slides (Marienfeld). The sections were stained with 0.2% cresyl violet (Sigma-Aldrich) and imaged using a dissecting microscope equipped with a digital camera (Olympus, Tokyo, Japan). The size of the infarct area was measured using ImageJ v1.4 software (National Institutes of Health, Bethesda, MD, USA). Reductions in brain volume were calculated as follows:
Brain volume reduction = (infarct size in the ipsilateral hemisphere)/(size of the intact contralateral hemisphere).

PEMs were built-up on glass slides from Marienfeld GmbH (Lauda-Königshofen, Germany). The dimension of the slides was 10 x 10 x 1 mm. They were purified by ultrasonication (Bandelin RK 100H Sonorex, Bandelin electronic, Germany) with 2% Hellmanex solution from Hellma (Müllheim, Germany) before rinsing with ddH₂O. Air-dried slides were sterilized in a heating furnace (Binder, Germany) at 200° C for 4 h prior using in PEM films build-up procedure.

PLGA films were build-up on glass slides from Paul Marienfeld GmbH (Lauda-Königshofen, Germany). The dimension of the slides was 10 × 10 × 1 mm. They were purified by ultrasonication (Bandelin RK 100H Sonorex, Bandelin electronic, Germany) with 2% Hellmanex solution from Hellma (Müllheim, Germany) before rinsing with ddH₂O. Air-dried slides were sterilized by steam sterilization for 20 min at 121 °C (systec dx-23, systec GmbH, Linden, Germany) prior coating.

Oligonucleotides, PCR pre-mix, and DNA extraction kits were purchased from Bioneer, Daejeon, Korea. Glass slides (2.5 by 7.5 cm) were procured from Paul Marienfeld GmbH & Co. KG, Lauda-Königshofen, Germany. High-performance liquid chromatography (HPLC) grade solvents were obtained from SK Chemicals, Seoul, Korea. A Milli-Q purification system (Millipore) was used for ultrapure water (18 MΩ/cm). Qarray2 microarrayer (Genetix Technologies, Inc., New Milton, UK) was used for oligonucleotide spotting. A commercial incubator and a commercial centrifuge (1000 rpm) were used. ScanArrayLite (GSI Lumonics) and Quant Array software (Packard Bioscience) were used for fluorescence signal measurement and image analysis, respectively.

Cells were seeded on poly-l-lysine-coated cover slips (thickness: 1.5H; Marienfeld, Lauda-Königshofen, Germany) in a 24 well plate (1–2 × 10⁵ cells/well) and transfected one day after seeding as detailed earlier. 24–48 h after transfection cells were washed with PBS and then fixed with 4% FA in PBS and permeabilized [0.2% (v/v) Triton X-100 and 50 mM NH₄Cl in PBS] at RT for 20 min. Cells were blocked with PBS supplemented with 5% (v/v) normal goat serum (Thermo Fisher Scientific) and 1% (w/v) BSA (Aurion, Wageningen, Netherlands) at RT for 30 min and subsequently incubated overnight at 4 °C with primary antibodies diluted in the same buffer. The following antibodies were used: anti-V5 (1:200, mouse mAb, #46-0705, Thermo Fisher Scientific), anti-GM130 (1:2000, rabbit pAb, #12480S, Cell Signalling Technology, Denver, MA, USA) and anti-calnexin (1:500, rabbit pAb, #10427–2-AP, Proteintech). Following washing, cells were stained with the Alexa Fluor-conjugated secondary antibodies (1:500, Thermo Fisher Scientific) diluted in PBS supplemented with 5% (w/v) BSA for 1 h at RT and then mounted on glass slides (Marienfeld) using ProLong Glass Antifade Mountant (Thermo Fisher Scientific) and stored at 4 °C. Samples were imaged using a Zeiss LSM980 microscope (63 × objective) and images were processed with Image J Version 1.53c.