Collagen XIII alpha 1 Antibody - #AF0598

Figure 2 Merlin deficiency impairs chondrocyte proliferation and hypertrophy (A) EdU incorporation and staining in E18.5 Nf2f/f and Nf2f/f;Prx1-Cre mouse tibial growth plates to assess chondrocyte proliferation. (B and C) Quantification of the percentage of EdU+ (proliferating) cells in the resting zone (B) and proliferative zone (C) in (A). (D and E) Ki67 IF in primary chondrocytes (D) and quantification (E). (F) CCK8 assay for cell proliferation. (G–I) IHC for COL10 and MMP13 (G) and quantification of positive area/intensity (H–I). (J and K) Alkaline phosphatase (ALP) and Alizarin red staining of differentiated chondrocytes (J) and quantification (K). (L) RT-qPCR for Mmp13, Col10, Runx2, and Opn, normalized to Hprt1. (M) Western blot for OPN, RUNX2, and MMP13; β-actin was the loading control. (N and O) TUNEL staining in hypertrophic zones (N) and quantification of apoptotic chondrocytes (O). EdU, 5-ethynyl-2′-deoxyuridine; TUNEL, terminal deoxynucleotidyl transferase (TdT) dUTP nick-end labeling. Scale bars, 20 μm (D), 50 μm (N), 100 μm (A and G), and 1 mm (J). Quantification data are presented as mean ± SD; p values (t test) are indicated between the bars. n≥ 4 (A–C), n = 7 (D and E), n = 3 (F), n≥ 3 (G–I), n = 8–13 (J–L), n ≥ 4 (M), and n = 8 (N and O) per group.

Figure 1 Merlin is expressed during limb development, and its deficiency causes limb dysplasia (A and B) In situ hybridization (ISH) showing Nf2/Merlin expression in limb buds at embryonic day 10.5 (E10.5)–E14.5. (C) Immunohistochemistry (IHC) of Merlin in E18.5 tibial growth plates, with enlarged views of proliferative (top) and hypertrophic (bottom) zones. Scale bars, 100 μm. (D) Western blot of COL10, MMP13, OPN, and Merlin in primary chondrocytes during hypertrophic differentiation; β-actin was the loading control. (E) Representative images of Nf2f/f and Nf2f/f;Prx1-Cre mice. (F and G) RT-qPCR of Nf2 expression in cartilage (F) and primary chondrocytes (G) from Nf2f/f;Prx1-Cre versus controls. Nf2 mRNA levels were normalized to Hprt1. (H–J) Whole-mount alizarin red-Alcian blue skeletal staining of E18.5 embryos, showing the whole body (H), limbs (I), and autopods (J). Quantification data are presented as mean ± SD; p values (t test) are indicated between bars. Data represent n = 3 (A-E), n ≥ 4 (H-J), n = 6 (F) and n = 4 (G) per group.