Journal: Science Advances

Article Title: Tumor collection/processing under physioxia uncovers highly relevant signaling networks and drug sensitivity

doi: 10.1126/sciadv.abh3375

Figure Lengend Snippet: Cells for flow cytometry analysis were cultured for 5 days, and only lineage-negative (CD31-PE, CD45-PE, and CD140b-PE negative) cells were gated for the analysis. ( A ) CD49f/EpCAM staining patterns ( n = 9). ( B ) The number of CD49f + /EpCAM − cells varied on the basis of O 2 tension and tumor type. ( C ) EpCAM positivity also showed tumor-specific variability under physioxia compared to ambient air. ( D ) CD24/CD44 staining patterns of cells from ascites fluid under physioxia and ambient air ( n = 9). ( E ) Percentage of CD44 + cells in each sample under physioxia and ambient air. ( F ) Percentage of CD24 + cells under physioxia and ambient air. CD24 + cells were lower under physioxia compared to ambient air. ( G ) Cells from ascites fluids collected under physioxia contained elevated levels of BRD4 protein compared to cells collected under ambient air.

Article Snippet: The antibodies used against mouse cells were CD31-PE/Cyanine7 (A14715) from Molecular Probes; CD45-PE/Cyanine7 (25-0451-82), EpCAM-APC (allophycocyanin) (17-5791-82), and CD29-FITC (fluorescein isothiocyanate) (11-0291-82) from Invitrogen; CD140a-PE/Cyanine7 (323508) from BioLegend; LGR5-PE (phycoerythrin) (FAB8240P), TSPAN8-APC (FAB6524A), and CD49f-PE (FAB13501P) from R&D Systems; and CD61-FITC (561911), CD274-PE (558091), CD24-APC (562349), and CXCR4-FITC (551967) from BD Pharmingen; isotype control antibodies used included PE/Cyanine7 (400522) and APC (400612) from BioLegend and PE (554689) and FITC (553971) from BD Pharmingen.

Techniques: Flow Cytometry, Cell Culture, Staining

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Differential expression of stem-cell-associated markers in human hair follicle epithelial cells.

doi: 10.1038/labinvest.2009.48

Figure Lengend Snippet: Figure 3 Biomarker expression in freshly isolated or cultured epidermal and follicular epithelial cells. (a) Schematic of the isolation and preparation of follicle- and epidermis-derived epithelial cells from the human scalp. (b) Representative data from a single-color FACS analysis of freshly isolated follicle- or epidermis-derived epithelial cells. Isolated cell suspensions were stained for K15, CD200, CD34, CD271, K14, K1, CD29, or CD49f. Note that a distinct expression pattern of CD200 was detected in follicle-derived cells compared to epidermis-derived cells. (c) Immunocytochemistry of cultured follicle- and epidermis-derived epithelial cells (passage 3) with stem-cell-associated surface markers and cytokeratins. Immunoreactivity was visualized with Alexa 546 in red; nuclei were counterstained with Hoechst 33342 in blue. Scale bars ¼ 20 mm. (d) Representative data from a single-color FACS analysis of cultured follicle- or epidermis-derived epithelial cells (passage 3). Isolated cell suspensions were stained for K15, CD200, or CD34. Numbers in blue indicate percentage of positive cells. Expression of these stem-cell-associated markers substantially declined with time in culture.

Article Snippet: Immunostaining against other cytokeratins and CD markers was performed with the following primary antibodies: anti-human K1 mAb (1:300, clone AE1; Progen, Heidelberg, Germany), antihuman K14 mAb (1:300, clone LL002; Abcam Inc.), anti-human CD29 mAb (1:100, clone 4B7R; Acris Antibodies, Hiddenhausen, Germany), and anti-human CD49f mAb (1:100, clone MP4F10; R&D Systems Inc., Minneapolis, MN, USA).

Techniques: Biomarker Discovery, Expressing, Isolation, Cell Culture, Derivative Assay, Staining, Immunocytochemistry