Strep fitc

Immunofluorescence staining was performed according to a previous report^{67 (link)}. Briefly, the cells were fixed in 10% buffered formalin for 30 min and washed with PBS. Non-specific binding was blocked using 10% horse serum. The cells were stained with primary antibodies at 4 °C overnight. The cells were then incubated with biotinylated secondary antibodies (Invitrogen) for 30 min and subsequently stained with Strep-FITC (Sigma). The nuclei were counterstained with DAPI (Sigma). Protein expression was visualized under a fluorescent microscope. The primary antibodies used were mouse anti-collagen I (C2456, Sigma), anti-OPN (AB1870, Merck Ltd.), and anti-RUNX2 (8486, Cell Signaling Technology).

Cells were fixed with 4% buffered formalin at room temperature for 10 min. Cell permeabilisation was performed using 0.15% Triton^®-X100 in PBS. Horse serum (2% v/v) was used to inhibit non-specific binding. Cells were stained with β-Catenin XP^® Rabbit mAb (cat. no. 8480, Cell Signaling, Danvers, MA, USA) at a 1:100 dilution at 4 °C overnight. Cells were incubated with biotinylated anti-rabbit IgG antibodies (cat. no. 2172707, Sigma-Aldrich, CA, USA) at a dilution of 1:2000 for 40 min. The targeted protein expression was visualised by staining with Strep-FITC (Sigma-Aldrich, CA, USA) at a dilution of 1:500. Nuclei were counterstained with 0.1 μg/mL 4′,6-diamidino-2-phenylindole (TOCRIS Bioscience, Minneapolis, MN, USA). Protein expression and localisation were performed using a fluorescent microscope with an ApoTome system (Carl Zeiss, Oberkochen, Germany).

Cells were fixed with 4% buffered formalin for 10 min. Horse serum (2% v/v) was used to inhibit a non-specific binding. Cells were stained with mouse anti-collagen I (C2456, Sigma) at 4 °C overnight and subsequently incubated with biotinylated anti-mouse antibodies (Invitrogen) for 40 min. The targeted protein expression was visualized by stained with Strep-FITC (Sigma). DAPI (TOCRIS bioscience) was employed for nuclei staining.