Anti caspase3

For immunohistochemistry experiments, explants were fixed with 4% PFA for 10 mins at RT, followed by permeabilization with 0.1% NP40 for 30 mins at 37°C. Next, the cells were blocked with 1% BSA in PBS for 30 mins at 37°C, prior to incubation with the primary antibody for 1h at 37°C. Primary antibodies were prepared in blocking solution, and were used in the following dilutions: mouse anti-E-cadherin antibody (BD Biosciences) (1:200), mouse anti-Paxillin antibody (BD Biosciences) (1:200), and rabbit anti-Active Yap1 (Abcam)(1:200), anti-pH3(S10) (Abcam, 1:200), anti-Caspase3 (R&D Systems, 1:200). Following primary antibody incubation, the explants were washed 4X times in PBS for 10 mins and incubated in a secondary antibody cocktail for 30 mins at 37°C. Finally, cells were washed 3X times in PBS and stained with DAPI or Phalloidin for 20 mins at RT. Post antibody staining, imaging was performed using Andor/Olympus Spinning Disk Confocal microscope at BRC facility, Cornell University.For Proximity Ligation Assay (PLA), explants were fixed and permeabilized as described above.The primary antibodies used were: Mouse Anti-YAP1 (DSHB) (1:5) and Rabbit Anti-TEAD1 (Abcam) (1:200). Following primary antibody incubation, the remaining steps of PLA were performed using reagents from the Duolink PLA detection kit (Sigma Aldrich, DUO92101) according to the manufacturer’s protocol.