anti cd51 61 antibody  (Santa Cruz Biotechnology)

Journal: Cells

Article Title: A New Method to Sort Differentiating Osteoclasts into Defined Homogeneous Subgroups

doi: 10.3390/cells11243973

Figure Lengend Snippet: Human osteoclasts differentiated in 3D form a more homogeneous cell population than monolayer cells. ( A – E ) Monolayer osteoclasts : ( A ) Representative images of staining for CD51/61 (top) and visualisation of resorption tracks (bottom, scale bars = 200 µm) and ( B ) TRAP staining of osteoclasts from 4 independent donors (scale bar = 200 µm) on day 9 of differentiation showing heterogeneity of the populations; (insert) monocytes (red arrow), binuclear pre-osteoclasts (blue arrow) and mature osteoclasts (green arrow). ( C ) Rate of formation of monolayer osteoclasts (TRAP-positive cells with ≥3 nuclei) and ( D ) the size distribution of these osteoclasts; small (3–5 nuclei), medium (6–9 nuclei) or large (>10 nuclei). ( E ) Representative TRAP-stained images of the time course of osteoclast differentiation in monolayer culture (scale bar = 100 µm). ( F – H ) 3D-generated osteoclasts: Cells were fixed for analysis 4 h after reseeding onto cell culture plates ( F ) Rate of formation of 3D-generated osteoclasts (TRAP-positive cells with ≥3 nuclei) and ( G ) the size distribution of these osteoclasts; small (3–5 nuclei), medium (6–9 nuclei) or large (>10 nuclei). ( H ) Representative TRAP-stained images of the time course of osteoclast differentiation in 3D (scale bar = 100 µm). *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: CD51/61 staining: immunostaining for osteoclast-specific CD51/61 used an anti-CD51/61 antibody (clone 23C6, 1:400; BioRad, Oxford, UK) and standard DAB immunohistochemistry techniques.

Techniques: Staining, Generated, Cell Culture

Journal: Cells

Article Title: A New Method to Sort Differentiating Osteoclasts into Defined Homogeneous Subgroups

doi: 10.3390/cells11243973

Figure Lengend Snippet: Cell surface markers in monolayer and 3D osteoclasts. ( A ) FACS scatter graphs showing increasing cell size and complexity during differentiation from CD14+ monocytes to osteoclasts in 3D culture. ( B ) Representative data from cells from one donor showing variation in osteoclast surface markers over 24 days of differentiation. Black = monolayer cells, red = 3D-generated cells. ( C , D ) Comparison of expression of ( C ) CD9 (n = 3) and ( D ) CD51/61 (n = 5) during 3D differentiation of osteoclasts from different monocyte donors. ( E ) Double staining of 3D-generated osteoclasts on day 15 of differentiation for CD9 and CD51/61 distinguishes two osteoclast subpopulations. ( F ) Scatter graph of CD9+CD51/61−cells showing small early/pre-osteoclasts (left panel) with CD9+CD51/61+ cells being larger mature osteoclasts (right panel). ***, p < 0.001.

Article Snippet: CD51/61 staining: immunostaining for osteoclast-specific CD51/61 used an anti-CD51/61 antibody (clone 23C6, 1:400; BioRad, Oxford, UK) and standard DAB immunohistochemistry techniques.

Techniques: Generated, Expressing, Double Staining

Journal: Cells

Article Title: A New Method to Sort Differentiating Osteoclasts into Defined Homogeneous Subgroups

doi: 10.3390/cells11243973

Figure Lengend Snippet: Live cell sorting of primary human osteoclasts. ( A ) Flow cytometry of live 3D-generated human osteoclasts stained for CD51/61. ( B ) Following cell sorting, the population of CD51/61+ cells increased from 13.3% to 84.2%. ( C ) Cells re-seeded into monolayer culture after live cell sorting and analysed for the presence of osteoclasts (TRAP stain, 24 h post re-seed, scale bar = 100 µm) and resorption activity (resorption tracks on dentine, 7 days post re-seed, scale bar = 200 µm). Top panel: CD51/61-positive cells from 3D culture. Middle panel: CD51/61-negative cells from 3D culture. Bottom panel: CD51/61-positive cells from monolayer culture. ( D ) Flow cytometry of live 3D-generated human osteoclasts stained for CD9 and CD51/61. CD9+CD51/61- and CD9+CD51/61+ cells were re-seeded into monolayer culture after live cell sorting and analysed for the presence of osteoclasts 24 h post re-seed (TRAP stain, scale bar = 100 µm), quantified ( E ) as the number of nuclei per cell in these selected populations. ( F ) Area of bone resorbed by monolayer and 3D-generated osteoclasts selected for CD51/61. Black = monolayer cells, red = 3D-generated cells. ***, p < 0.001.

Article Snippet: CD51/61 staining: immunostaining for osteoclast-specific CD51/61 used an anti-CD51/61 antibody (clone 23C6, 1:400; BioRad, Oxford, UK) and standard DAB immunohistochemistry techniques.

Techniques: FACS, Flow Cytometry, Generated, Staining, Activity Assay