af488 cd31  (Cell Signaling Technology Inc)

Journal: Biology

Article Title: Exosome-Mediated Transfer of X-Motif-Tagged Anti-MiR-33a-5p Antagomirs to the Medial Cells of Transduced Rabbit Carotid Arteries

doi: 10.3390/biology13120965

Figure Lengend Snippet: Figure 1. Sorting of carotid endothelial and smooth muscle cells. Three days after HDAd treatment, rabbit carotids were harvested and enzymatically digested, and cell suspensions were stained with antibodies targeting CD45 (immune cell marker) and CD31 (endothelial cell marker) or isotype controls. (A,B) Scatter plots of (A) total cells and (B) single cells were used to discard potential cell debris and doublet events. Each dot represents one event; the red color indicates a higher concentration of events, whereas blue represents less event concentration. The graph axes show SSC and FSC (arbitrary units) that represent cellular granularity and size, respectively. (C–F) Histograms show the fluorescence intensity from APC or AF488 fluorochromes (logarithmic scale, arbitrary units) on the X-axis and event count on the Y-axis. Gates for (C) CD45 and (D) CD31 were set using isotype control antibodies. (E) Histogram of CD45 staining allowed for discrimination of CD45-negative cells—primarily endothelial cells and smooth muscle cells—from CD45-positive cells (i.e., immune cells). (F) Histogram of CD31 staining using the previously gated (E) CD45-negative cell population. The CD45−CD31+ endothelial cells (shown with red background) and CD45−CD31−smooth muscle cells (green background) were sorted for downstream RNA analysis. To minimize the potential sorting of EC (with lower CD31 expression than the established CD31+ threshold) into the SMC- enriched CD45−CD31−population, the gate for CD45−CD31−cells (green background) was set at the lowest AF488 signal of the peak of CD45−CD31−cells. Gate range of EC population (AF488, arbitrary units): 2 × 103 to 2 × 105. Gate range of SMC population (AF488, arbitrary units): 5 × 101

Article Snippet: Cells were blocked with 5% normal mouse serum (Sigma, Saint Louis, MO, USA, NS03L) and then stained with 1 μg/mL Alexa Fluor (AF) 647 mouse anti-CD45 (ThermoFisher, MA5-28392), 10 μg/mL AF488 mouse anti-CD31 (SCBT, Dallas, TX, USA, sc-376764), and 1 μg/mL chicken anti-ABCA1 (Pacific Immunology, custom made) primary antibodies.

Techniques: Staining, Marker, Concentration Assay, Fluorescence, Control, Expressing

Journal: Stem Cell Research & Therapy

Article Title: Von willebrand factor increases endothelial cell adhesiveness for human mesenchymal stem cells by activating p38 mitogen-activated protein kinase

doi: 10.1186/scrt35

Figure Lengend Snippet: Image of hMSCs adherent to a confluent monolayer of HUVECs treated with vWF . Confocal image of a planar (a) and Z-axis (b) projections of hMSCs adherent to HUVECs treated with 4 μg/ml vWF for four hours. HUVECs were stained with AF488-conjugated CD31 (green). HMSCs were labeled with PE-conjugated CD90 (red). HMSCs were found on the top of endothelial monolayer within the boundaries of ECs.

Article Snippet: Cells were fixed with 5% paraformaldehyde, permeabilized with 0.1% Triton x100 in phosphate buffered saline, blocked with 5 mg/ml bovine serum albumin in PBS for one hour and stained with 1 μg/ml AF488-conjugated CD31 antibody (BD Pharmingen, Franklin Lakes, NJ, USA), the specific antigen marker of HUVECs, and 1 μg/ml PE-conjugated CD90 antibody (BD Pharmingen), the specific antigen marker of hMSCs, for four hours.

Techniques: Staining, Labeling