Protein Expression Analysis of hiPSC-Derived CD34+ Cells

Magnetically sorted hiPSC-derived CD34⁺ cells were washed three times with cold PBS, lysed in protein-preservative buffers, and stepwise prepared for polyacrylamide gel electrophoresis, subsequently transferred to membranes by electroblotting as previously described [30 (link)]. Afterward, membranes were incubated with the primary antibodies: p-YAP1 (phospho-Ser127, 1:500, Abcam), YAP1 (1:1000, Abcam), p-WWTR1 (phospho-Ser89, 1:500, Invitrogen), WWTR1 (1:1000, Invitrogen), and GAPDH (1:20,000, Invitrogen), overnight at 4 °C. Subsequently, membranes incubated with antibodies were washed five times with Tris-buffered saline plus Tween 20 and incubated with HRP-conjugated secondary antibodies. Targeted signals were exposed with HRP Substrate and detected by Azure 600 (Azure Biosystems). Detailed information on these antibodies and reagents is provided in Additional file 1: Table S1.