Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology

Article Title: Optimization of rhabdomyosarcoma disseminated disease assessment by flow cytometry.

doi: 10.1002/cyto.a.22514

Figure Lengend Snippet: Figure 1. (A) Flow cytometry analysis of PAX3 staining positivity in a panel of non-RMS cancer cell lines (gray bars) and RMS cell lines (black bars). The name of each cell line and corresponding cancer type are indicated at the base of each bar. (B) Cytometric plots in RH30 cell line illustrating the analysis made for each cell line in A. Each cell line was labeled with an isotype control (upper panel, C-) and anti- PAX3 antibody (lower panel). Plots depict the forward and reverse scattering (left), viability analysis (B Blue-A, 7-AAD, center), and PAX3 labeling (E YG-A, PE, right), respectively.

Article Snippet: Samples were then incubated with phycoerythrin (PE)-conjugated anti-PAX3 primary antibody (10 mL/5 3 105 cells, R&D Systems) for 30 min at room temperature.

Techniques: Flow Cytometry, Staining, Labeling, Control

Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology

Article Title: Optimization of rhabdomyosarcoma disseminated disease assessment by flow cytometry.

doi: 10.1002/cyto.a.22514

Figure Lengend Snippet: Figure 2. (A) PAX3 expression levels in tumors measured by quantitative PCR in eRMS (dark gray bars), aRMS (black bars), and other pediatric sarcomas (soft gray bars), including Ewing’s sarcoma (ES), synovial sarcoma (SS), and fibrosarcoma (FS). PB and BM: peripheral blood and bone marrow from healthy donors. (B) Relative PAX3 mRNA expression in RMS cell lines. In both A and B, all relative mRNA levels are referred to those of PAX3 in normal fetal muscle.

Article Snippet: Samples were then incubated with phycoerythrin (PE)-conjugated anti-PAX3 primary antibody (10 mL/5 3 105 cells, R&D Systems) for 30 min at room temperature.

Techniques: Expressing, Real-time Polymerase Chain Reaction

Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology

Article Title: Optimization of rhabdomyosarcoma disseminated disease assessment by flow cytometry.

doi: 10.1002/cyto.a.22514

Figure Lengend Snippet: Figure 3. (A) Sensitivity of quantitative PCR for PAX3, PAX3/FOXO1, PAX7/FOXO1, MYOD, and MYOGENIN. Serial dilutions of RMS cells at 10-fold increments in peripheral blood were analyzed for expression of the markers assessed by qPCR. The points not shown for each marker correspond to points under the qPCR limit of detection and therefore not detected. ND: not detected in any dilution. (B) Expression levels of MYOD, MYOGENIN, and PAX3 in the three cell lines used in spiking experiments. All determinations were made in triplicate.

Article Snippet: Samples were then incubated with phycoerythrin (PE)-conjugated anti-PAX3 primary antibody (10 mL/5 3 105 cells, R&D Systems) for 30 min at room temperature.

Techniques: Real-time Polymerase Chain Reaction, Expressing, Marker

Journal: Cytometry. Part A : the journal of the International Society for Analytical Cytology

Article Title: Optimization of rhabdomyosarcoma disseminated disease assessment by flow cytometry.

doi: 10.1002/cyto.a.22514

Figure Lengend Snippet: Figure 4. Flow cytometry plots of serial dilutions of RMS cells (RH30) in peripheral blood. Samples were labeled with PAX3 and CD45 anti- bodies. The number on the left denotes the input of RMS cells/107 blood mononuclear cells. The plots depict forward and side scatterings (left), viability and forward scattering (B Blue-A, 7-AAD, center), and double labeling PAX3 and CD45 (E-Blue-A/PE and E YG-A/FITC, respectively, right). In the last case, plots were subdivided into four quadrants and the percentage of cells in each quadrant is indicated inside the corresponding quadrant. RMS cells are shown in the lower-right quadrant (CD452/PAX31).

Article Snippet: Samples were then incubated with phycoerythrin (PE)-conjugated anti-PAX3 primary antibody (10 mL/5 3 105 cells, R&D Systems) for 30 min at room temperature.

Techniques: Flow Cytometry, Labeling

Journal: bioRxiv

Article Title: Genetic approaches in mice demonstrate that neuro-mesodermal progenitors express T/Brachyury but not Sox2

doi: 10.1101/503854

Figure Lengend Snippet: ( a ) Sox2 protein and mRNA levels are down-regulated in n = 5/5 (immunostaining) and n = 4/4 (WISH) T CreERT2/+ ; Sox2 f embryos. Sox2 staining: left side taken with the same exposure time as for wild-type (WT); right side taken with longer exposure time. Sox1 is not expressed at the caudal end of either WT nor in T CreERT2/+ ; Sox2 fl/fl embryos. Insets in a4 and a10 show cranial sections from the same embryo, stained and processed in parallel, as positive control for Sox1 antibody. Sox3 appears up-regulated in n = 5/5 T CreERT2/+ ; Sox2 fl/fl embryos compared to WT (n-values for WISH, IHC: WT, n = 3, 5 embryos; T CreERT2/+ ; Sox2 fl/fl , n = 4, 5 embryos). Dotted circles in a3 – 4 and a9 – 10 indicate the CNH. ( b ) Following multiple tamoxifen injections, n = 4/4 T CreERT2 +; Sox2 flfl embryos show normal neural tube morphology with normal dorso-ventral expression domains of Pax3, Pax6 and Nkx6.1 (n-values for WISH, IHC: WT, n = 3, 3 embryos; T CreERT2/+ ; Sox2 fl/fl , n = 3, 4 embryos). Dashed lines in a3 – 6, a9 – 12, b2 – 3, b5 – 6 outline the neuroepithelium. Scale bars: 1 mm for whole embryo ( b1 ), 500 μm for whole embryo and PNP ( a1 – 2 ); 100 μm for sections ( a3 – 4, b2 – 3 ).

Article Snippet: Primary antibodies: rabbit monoclonal anti-Sox2 (ab92494, Abcam, 1:500), chicken polyclonal anti-GFP (ab13970, Abcam, 1:500), mouse monoclonal anti-Sox10 (sc-365692, Santa Cruz, 1:500), mouse monoclonal anti-Pax3 (MAB2457, R&D, 1:200), mouse monoclonal anti-Cre recombinase (MAB3120, Merck Millipore, 1:1,000), rabbit monoclonal anti-Sox1 (GTX62974, Insight Biotechnology, 1:200), rabbit polyclonal anti-Sox3 (ab183606, Abcam, 1:200), rabbit polyclonal anti-Pax6 (901301, BioLegend, 1:200), mouse monoclonal anti-Nkx6.1 (F55A10, Developmental Study Hybridoma Bank, deposited by Madsen, O.D., 1:5).

Techniques: Immunostaining, Staining, Positive Control, Expressing

Journal: Stem cells (Dayton, Ohio)

Article Title: A highly enriched niche of precursor cells with neuronal and glial potential within the hair follicle dermal papilla of adult skin.

doi: 10.1634/stemcells.2007-0281

Figure Lengend Snippet: Figure 2. The dermal papilla is highly en- riched for sphere-forming cells of neural crest origin. (A–C): Immunohistochemical analysis of the end bulb of the rat vibrissal follicle (green, fibronectin; blue, Hoechst nuclear stain; red, nestin). (D): The dermal papilla was cleanly microdissected prior to dissociation to single cells (described in text). Scale bar 100 m. (E): The micro- dissected adult dermal papilla was highly enriched for sphere-forming cells; 24.4% 4.3% of dermal papilla cells generated spheres when dissociated and cultured in SKP proliferation medium. This compares with 2.8% 0.6% of dissociated whisker follicle cells and 0.029% 0.028% of dis- sociated whole facial skin. (F): Papillas- pheres express the neural crest transcription factors, Pax3 and Slug (embryonic day 12 neural tube neural crest control). (G–N): Spheres generated from the rat vibrissa der- mal papilla had an immunocytochemical profile similar to that of SKPs isolated from whole facial skin. Papillaspheres expressed nestin (red [I, M]) and fibronectin (green [J, N]). Papillaspheres and SKPs derived from the Wnt1-Cre/R26R mouse both expressed -galactosidase, detected by X-Gal staining (H, L). Abbreviations: GAPDH, glyceralde- hyde-3-phosphate dehydrogenase; SKP, skin- derived precursor cell; X-Gal, 5-bromo-4- chloro-3-indolyl--D-galactoside.

Article Snippet: Primary antibodies used were as follows: fibronectin (rabbit polyclonal, 1:800; Sigma-Aldrich), vimentin (mouse monoclonal IgG1, MAB1681; Chemicon, Temecula, CA, http://www.chemicon.com), versican (mouse IgG1, 1:4 supernatant; kind gift Dr. Richard Asher), -smooth muscle actin (mouse IgG2a, 1:200, clone 1A4; Sigma-Aldrich), nestin (rat: mouse IgG1, 1:200, MAB353; human: mouse IgG, 1:500; Chemicon), musashi (rabbit polyclonal, 1:200, AB5977; Chemicon), III-tubulin (mouse IgG2b, 1:500; Sigma-Aldrich), microtubule-associated protein (MAP2ab) (1:200; Sigma-Aldrich), glial fibrillary acidic protein (GFAP; 1:250; Dako, Glostrup, Denmark, http://www.dako.com), S100 (rabbit polyclonal, 1:200; Dako), p75 (mouse IgG1monoclonal, 1:50, MAB365R; Chemicon), CNPase (mouse IgG1 monoclonal, 1:100, C5922; Sigma-Aldrich), fatty acid-binding protein (FABP; goat, 1:10, SC006; R&D Systems), -galactosidase (rabbit polyclonal, 1:100, AB1211; Chemicon), Sox10 (monoclonal mouse IgG1, 1:50, MAB2864; R&D Systems), and Pax3 (goat IgG, 1:100, AF2457; R&D Systems).

Techniques: Immunohistochemical staining, Staining, Generated, Cell Culture, Whisker Assay, Control, Isolation, Derivative Assay

Journal: Stem cells (Dayton, Ohio)

Article Title: A highly enriched niche of precursor cells with neuronal and glial potential within the hair follicle dermal papilla of adult skin.

doi: 10.1634/stemcells.2007-0281

Figure Lengend Snippet: Figure 4. Generation of human papillas- pheres by microdissection of facial hair fol- licles. (A): H&E stain of human facial skin showing epidermis, dermis, and location of the end bulb of the hair follicle within the skin (boxed area). (B): Higher-power image of the same section showing the outline of the dp, which can be microdissected with minimal contamination using a method sim- ilar to that outlined in Figure 1. Scale bar 50 m. (C): Cells generated from a single human hair follicle dp grew as spheres in SKP proliferation medium. (D–H): The ma- jority (78.6% 12.4%) of plated human papillasphere cells expressed the neural crest transcription factor Sox10. A subpopulation of cells coexpressed Pax3 (white arrow). Ab- breviation: dp, dermal papilla.

Article Snippet: Primary antibodies used were as follows: fibronectin (rabbit polyclonal, 1:800; Sigma-Aldrich), vimentin (mouse monoclonal IgG1, MAB1681; Chemicon, Temecula, CA, http://www.chemicon.com), versican (mouse IgG1, 1:4 supernatant; kind gift Dr. Richard Asher), -smooth muscle actin (mouse IgG2a, 1:200, clone 1A4; Sigma-Aldrich), nestin (rat: mouse IgG1, 1:200, MAB353; human: mouse IgG, 1:500; Chemicon), musashi (rabbit polyclonal, 1:200, AB5977; Chemicon), III-tubulin (mouse IgG2b, 1:500; Sigma-Aldrich), microtubule-associated protein (MAP2ab) (1:200; Sigma-Aldrich), glial fibrillary acidic protein (GFAP; 1:250; Dako, Glostrup, Denmark, http://www.dako.com), S100 (rabbit polyclonal, 1:200; Dako), p75 (mouse IgG1monoclonal, 1:50, MAB365R; Chemicon), CNPase (mouse IgG1 monoclonal, 1:100, C5922; Sigma-Aldrich), fatty acid-binding protein (FABP; goat, 1:10, SC006; R&D Systems), -galactosidase (rabbit polyclonal, 1:100, AB1211; Chemicon), Sox10 (monoclonal mouse IgG1, 1:50, MAB2864; R&D Systems), and Pax3 (goat IgG, 1:100, AF2457; R&D Systems).

Techniques: Laser Capture Microdissection, Staining, Generated