Tritc goat anti rabbit antibody

Immunostaining for SNAIL and αSMA were performed as described below. Paraffin sections of BM were treated with 100% xylene for 5min twice, 100% xylene and 100/% ethanol 1:1 ratio for 3 min, 100% ethanol for 5 min twice, 95% ethanol for 5min, 70% ethanol for 5min and 50% ethanol for 5 min and rinsed with cold water for 1 min followed by antigen retrieval with sodium citrate buffer at 100°C for 20 min. Blocking was performed with 10% goat serum for 2 hours. Anti-SNAIL (Rabbit, 1:100, Abclonal), anti-αSMA (rabbit, 1:300, Abclonal) and were used for staining. Secondary staining was done using TRITC goat anti-rabbit antibody (1:200 dilution) (Jackson Immunoresearch) and mounted in vectashield mounting medium with DAPI (H-1200, Vector Labs). Fluorescence was visualized using Zeiss LAM 710 confocal microscope. Data were analyzed using Image J software (Image J). Scale bars, 15μm.

Mesenchymal stromal cells (MSC) were generated from mice BM as previously described (26 (link)). For immunofluorescence staining, MSCs were grown on cover slips and incubated with TGF-β1 (50ng/ml) for 72 hours in the presence or absence of Palbociclib (0.25μM). Collagen staining was performed with un-conjugated antibodies against Collagen I or Collagen III (Abcam). Secondary staining was done using TRITC goat anti-rabbit antibody (Jackson Immunoresearch). Fluorescence was visualized using Zeiss LAM 710 Confocal microscope.