Micropatterned plate preparation

The protocol employed has been reported elsewhere (Tewary et al, 2019; preprint: Kaul et al, 2020). Briefly, custom Nexterion‐D Borosilicate (Schott, D263) thin glass coverslips (dimensions 110 × 74 mm) were spin‐coated with Lipidure‐CM5206™ (Lipidure) (NOF America Corporation). Lipidure was reconstituted in 100% ethanol at 2.5 mg/ml. The coverslips were sterilized by covering their side with isopropanol alcohol and spinning them at 2,500 rpm for 30 s on the Laurell Spin Processor (Laurell Technologies Corporation, Model # WS‐650Mz‐23NPPB). This was followed by addition of 1.5 ml Lipidure solution on the sterilized side of the coverslip and spinning it at 2,500 rpm for another 30 s. To create micropatterns, the Lipidure‐coated side of the coverslip was exposed to deep UV for 20 min through a quartz photomask. The diameter of each micropattern was 1 mm. The coverslips were subsequently attached to 96‐well bottomless microtiter plates (VWR, Catalog # 82050‐714) using epoxy (Loctite, M‐31CL). Carboxyl groups on the photo‐activated regions were activated with 50 μl/well of 0.05 g/ml N‐(3‐Dimethylaminopropyl)‐N′‐ethylcarbodiimide hydrochloride (Sigma‐Aldrich, 03450) and N‐Hydrosuccinimide (Sigma‐Aldrich, 130672) solution for 20 min. The wells were then rinsed followed by the addition of 2% Geltrex. The plate was subsequently left on an orbital shaker overnight at 4°C.