igg1 negative control-fluorescein isothiocyanate (fitc  (Bio-Rad)

Journal: American Journal of Cancer Research

Article Title: Simultaneous inhibition of discoidin domain receptor 1 and integrin αVβ3 radiosensitizes human glioblastoma cells

doi:

Figure Lengend Snippet: Surface expression of αVβ3 and αVβ5 integrins as well as RGD peptide binding capacity remain unaltered under DDR1-IN-1 or Cilengitide administration. (A-D) Flow cytometry analysis of U-251MG and LN-229 GBM cells treated with DMSO control, DDR1-IN-1 or cilengitide under 2D adherent or 3D gliosphere conditions. Cells were surface stained with (A) FITC-conjugated non-specific IgG antibodies (control), FITC-conjugated antibodies specific for (B) integrin αVβ3 and (C) integrin αVβ5 or (D) RGD binding. Data show means ± SD (n = 3, one-way ANOVA, Dunnett post hoc test: *P<0.05).

Article Snippet: Cells were stained using FITC conjugated anti-αVβ5 (MAB1961F), FITC conjugated anti-αVβ3 (MAB1976F), FITC conjugated mouse IgG1 negative control (MABC002F) (all Merck Millipore) or a FITC conjugated RGD-containing peptide GRGDSP (AS-60619, Anaspec, Fremont, CA, USA) for 30 min on ice and protected from light.

Techniques: Expressing, Binding Assay, Flow Cytometry, Staining

Journal: International Journal of Nanomedicine

Article Title: Functionalization-dependent effects of cellulose nanofibrils on tolerogenic mechanisms of human dendritic cells

doi: 10.2147/IJN.S183510

Figure Lengend Snippet: Effects of CNF samples on viability and morphology of DC. Notes: ( A ) Viability of DC after the culture with CNF samples relative to control DC (100%). ( B ) A representative flow cytometry analysis of SSC/FSC, and the summarized data from seven experiments with different donors are shown as fold change of control DC ± SD. ( C ) MGG-stained DC collected after the cultures with CNF. ( D ) Epi-fluorescent images of CNF-treated DC after staining with phalloidin rhodamine-actin (red), anti-CD209-IgG Alexa 488 (green), calcofluor white-CNF (dark blue), and DAPI-nuclei (light blue). Scale bars represent 10 μm. ( E ) Fluorescence intensity (CD209 and actin) in the contact area with nCNF (3) against the cytoplasmic part of the cells (whole cell excluding nucleus [2]) was calculated as MGV relative to control DC, and the data for each cell measured in one experiment are shown with indicated median and range statistics. ( F ) The surface expression of CD209 on DC, as determined by flow cytometry. The results are shown as mean MFI ± SD of three independent experiments. ( C , D ) Image scale bars represent 10 μm. ( E , F ) * P <0.05 compared to control, or as indicated (RM ANOVA with Tukey’s post-test). Abbreviations: APA, 3-aminopropylphosphoric acid; APAc, APA-functionalized; c, carboxylated; CNF, cellulose nanofibrils; DC, dendritic cells; FSC, forward scatter; MFI, mean fluorescence intensity; MGG, May-Grunwald Giemsa; MGV, mean gray value; n, native; RM, repeated measures; SSC, side scatter.

Article Snippet: Phenotype analysis of DC and T cells after the cultures was carried out using flow cytometer (Sysmex Partec Cube 6) after staining the cells by using the following Abs (Clone) and reagents: immunoglobulin (Ig) G1a negative control-biotin (MCA928), IgG1 negative control-phycoerythrin (PE) (MCA928PE), IgG1 negative control-fluorescein isothiocyanate (FITC) (MCA928F), anti-CD1a-PE-Cy5 (NA1/34HLK) (all from Serotec, Oxford, UK), anti-human leukocyte antigen (HLA)-DR-biotin (LN3), IgG1a negative control-PECy5 (P.3.6.2.8.1), anti-CD86-PE (IT2.2), streptavidin-PECy5, anti-CD4-PECy5 (RPA-T4), anti-IL-4-PE (8D4-8), anti-ILT3-PE (ZM4.1), anti-ILT-4-PE (42D1), anti-TGF-β-biotin (eBio16TFB), anti-CD25-PE, anti-CD25-PECy5 (BC96), anti-forkhead box (Fox) P3-FITC (PCH101), anti-IL-10- PE (JES5-16E3), anti-CD39-FITC (A1), anti-CD8-PEcy5 (RPA-T8), anti-cytotoxic T-lymphocyte-associated protein (CTLA)-4-PE (14D3) (all from eBioscience), streptavidin-Alexa 488, anti-mouse IgG-Alexa 488, anti-CD1a-PE (HI149) (all from Biolegend, San Diego, CA, USA), anti CD40-allophycocyanin (APC) (5C3), anti-IL-12 (p40/p70)-PE (C11.5) (all from BD Pharmingen, San Diego, CA, USA), anti-CD83-FITC (HB15e), anti-IFN-γ-FITC (25723), anti-IL17-peridinin-chlorophyll-protein complex conjugate (PerCP) (41802), anti-IL-10-FITC (127107), anti-HLA-DR PerCP (L243), anti-CD4-FITC, anti-CD4-PerCP (11830), anti-IDO-1-APC (700838) (all from R&D Systems), anti-CD14-FITC (TUK4), IgG1 negative control-PerCP (IS5-21F5) (Miltenyi Biotec), anti-CD4-PE (MEM-241) (Partec Sysmex).

Techniques: Flow Cytometry, Staining, Fluorescence, Expressing

Journal: Radiation Oncology (London, England)

Article Title: Hsp70 - a biomarker for tumor detection and monitoring of outcome of radiation therapy in patients with squamous cell carcinoma of the head and neck

doi: 10.1186/1748-717X-9-131

Figure Lengend Snippet: Hsp70 membrane expression on biopsies of SCCHN patients. A . The cells stained with FITC-labeled cmHsp70.1 monoclonal antibody corrected with IgG1 isotype-matched control antibody are depicted in box plots. Data from patients’ tumor cells (n = 21) were divided into low (n = 10) and high (n = 11) mHsp70 expressing cells. Single cells from healthy donors (n = 7) derived from connective tissues were used as controls. B . Mean fluorescence intensity from the same data set as shown in A. Standard box plots are shown with boundaries indicating the 25 th and the 75 th percentile. The line inside boxes indicates the median and the whiskers indicate the 10 th and 90 th percentile, respectively. All outliers are included into the graphs. (* p < 0.05, ** p < 0.01, *** p < 0.001).

Article Snippet: 1 × 10 5 cells were washed once with 10% FCS in PBS and incubated with a FITC-conjugated mouse monoclonal antibody specific for membrane-bound Hsp70 (cmHsp70.1, IgG1, multimmune GmbH, Munich, Germany) or a FITC-labeled isotype-matched IgG1 negative control antibody (345815, BD Biosciences, Franklin Lakes, NJ, USA) on ice in the dark for 30 min. After washing, propidium iodide was added and viable cells were analyzed on a FACSCalibur flow cytometer (BD Biosciences).

Techniques: Expressing, Staining, Labeling, Derivative Assay, Fluorescence