STAT3-responsive Luciferase Assay for TrkA

Luciferase assays were performed using a dual-luciferase assay kit (Biotium, Fremont, CA, USA) and readouts were collected using the Molecular Devices iD3 plate reader (San Jose, CA, USA). Cells were co-transfected with a STAT3-responsive luciferase reporter (pGAS-luc), and either vector or TrkA (pCMV5-TrkA) plasmid for 28 h before serum starvation for 16 h. Then, cells were stimulated with 100 ng/mL β-NGF for 4 h before the addition of lysis buffer. Lysates underwent mechanical lysis by scraping followed by trituration with a pipette to ensure complete lysis. Firefly luciferase activity was measured as previously described [56 (link),63 (link)]. Results are represented as mean ± SD. Student’s t-tests were performed using GraphPad Prism 8.