Osteocalcin Antibody - #DF12303

Fig. 5 Biocompatibility assessments. a CCK-8 assay about the proliferation of MC3T3-E1 preosteoblasts cultured on Ti-OH and Ti-PAA-NCl after cells were cultured for 1, 3 and 7 days (n = 3; P = 0.673, 0.639 and 0.145 for 1, 3 and 7 days compared with Ti-OH, respectively; Student’s t test). b Morphology of MC3T3-E1 preosteoblasts cultured on Ti-OH and Ti-PAA-NCl for 1 day (green for F-actin, blue for cell nucleus, scale bar = 20 μm). c ALP activity of MC3T3-E1 preosteoblasts cultured on Ti-OH and Ti-PAA-NCl for 7 and 14 days (n = 3; P = 0.132 and 0.954 for 7 and 14 days compared with Ti-OH; Student’s t test). d Alizarin Red S staining of MC3T3-E1 preosteoblasts cultured on Ti-OH and Ti-PAA-NCl for 21 days (red for calcium nodules, scale bar = 1 mm) and semi-quantitative measurement of calcium content (n = 3; P = 0.147 compared with Ti-OH; Student’s t test). e Western blot results of osteogenic-related proteins (OCN, OPN and RUNX2) expressed in MC3T3-E1 preosteoblasts cultured on Ti-OH and Ti-PAA-NCl for 3, 7 and 14 days after osteogenic induction. f RT-qPCR results of expression levels of osteogenic-related genes (OCN, OPN and RUNX2) in MC3T3-E1 preosteoblasts cultured on Ti-OH and Ti-PAA-NCl for 3, 7 and 14 days after osteogenic induction (n = 3; P > 0.05 for all time points of these three genes between groups; Student’s t test). g In vivo biocompatibility. HE staining and CD68 immunofluorescent staining of tissues around Ti-OH and Ti-PAA-NCl embedded in the backs of nude mice for 4 weeks (fluorescent green for CD68, fluorescent blue for cell nucleus, white scale bars in HE images = 100 μm, black scale bars in HE images = 25 μm, scale bars in CD68 immunofluorescent images = 100 μm). All error bars = s.d.

Figure 1. POL stimulated BMSCs proliferation and osteogenic differentiation. (A) POL chemical structure; (B) BMSCs surface markers were detected by flow cytometry; (C) The result of CCK-8 assay (n = 5); (D-E) The result of CFU assay (n = 5); (F) Images of ALP staining (scale bar = 250 μm); (G) Quantification of ALP staining (n = 5); (H) Images of ARS staining (scale bar = 250 μm); (I) Quantification of ARS staining (n = 5); (J-K) The protein expression levels of OCN, RUNX2, ALP, and COL1A1 were evaluated by western blot (n = 3); (L-M) Immunofluorescence staining of OCN and ALP (scale bar = 200 μm). Data were presented as mean ± SEM. Compared with control group: * P < 0.05, **P < 0.01, ***P < 0.001. Compared with 1 μM group: # P < 0.05, ##P < 0.01, ###P < 0.001.