Mouse anti α tubulin sc 8035

Rabbit anti-FLAG (600-401-383, 1:5000) was purchased from Rockland. For immunofluorescent microscopy, mouse anti-FLAG (F1804, 1:100) was purchased from Sigma-Aldrich. Rabbit anti-MCM4 (A300-193A, 1:5000) was purchased from Bethyl Laboratories. Mouse anti-α tubulin (sc-8035, 1:1000), rabbit anti-lamin A/C (sc-20681, 1:3000) and rabbit anti-Histone H3 (sc-10809, 1:500) were purchased from Santa Cruz. Rabbit anti-VDAC1 (ab15895, 1:5000) was purchased from Abcam. Rabbit anti-p32 (6502S, 1:1000) was purchased from Cell Signaling Technology. Rabbit anti-TWINKLE (ARP36483_P050, 1:1000) and rabbit anti-TFAM (OAAN00609, 1:1000) was purchased from Aviva Systems Biology. Mouse anti-mtSSB (1:1000) was kindly provided by Dr. Valeria Tiranti (The Foundation of the Carlo Besta Neurological Institute, IRCCS, Milan, Italy).

Lung samples were homogenized in RIPA buffer containing 20 mM Tris–HCl, pH 7.4, 150 mM NaCl, 1% Triton-X100, 0.5% deoxycholate sodium, 0.1% sodium dodecyl sulfate, 1 mM EDTA, 10 mM β-glycerophosphate, 10 mM NaF, 1 mM Na₃VO₄, and protease inhibitor cocktail. The extracts were sonicated for 10 min and centrifuged at 13,000 × g for 15 min. Western blotting analysis was performed using an ECL system [13 (link)]. The following primary antibodies were used: mouse anti–BMPR2 (#GTX60415; GeneTex, CA, USA; 1:1000 dilution), rabbit anti-RFX2 (#12622-1-AP; Proteintech, IL, USA; 1:500 dilution), and mouse anti-α Tubulin (SC-8035; Santa Cruz Biotechnology, Tx, USA; 1:1000 dilution). Signals were detected using Immunostar Zeta (Fuji film, Japan) and an Amersham Imager 600 series imager (GE Healthcare, IL, USA). Statistical analyses of protein expression levels were performed using Fiji.

Unless otherwise stated, chemicals and reagents were purchased from Sigma-Aldrich (St. Louis, MO, USA). Goat anti-GAS6 (sc-1936) and mouse anti-α-TUBULIN (sc-8035) were obtained from Santa Cruz Biotechnology (Dallas, TX, USA).