To visualize the vessel networks formed by the EC, the tissues were immunofluorescently stained with mouse anti-human CD31 antibody (Dako, Carpinteria). First, the tissues were fixed in 10% formalin, and then blocked with a 2% bovine serum albumin (BSA, Sigma-Aldrich) and 0.1% Tween 20 (Sigma-Aldrich) solution. Overnight incubation with CD31 antibody diluted at 1:200 was followed by overnight incubation with Alexa Fluor 555-conjugated goat anti-mouse IgG (Invitrogen) diluted at 1:500. Washes with a 0.1% Tween 20 solution followed both incubation periods. Images of the stained tissues used for vessel quantification purposes were acquired using an inverted fluorescence microscope (Nikon Eclipse TE300). Images to confirm lumen formation and intravascular cell migration were acquired using laser scanning confocal microscopy (Zeiss LSM 510 Meta).
Mouse anti human cd31 antibody
Manufactured by Agilent Technologies
Sourced in United States
The Mouse anti-human CD31 antibody is a laboratory reagent used for the detection and identification of the CD31 antigen, also known as platelet endothelial cell adhesion molecule-1 (PECAM-1). CD31 is a transmembrane glycoprotein that is expressed on the surface of endothelial cells, platelets, and certain immune cells. This antibody can be used in various immunological techniques, such as flow cytometry, immunohistochemistry, and western blotting, to study the expression and distribution of the CD31 antigen in biological samples.
Automatically generated - may contain errors
Lab products found in correlation
Lsm 510 meta, by Zeiss (2 mentions)
Alexa fluor 555 conjugated goat anti mouse igg, by Thermo Fisher Scientific (1 mentions)
Eclipse te300, by Nikon (1 mentions)
Tween 20, by Merck Group (1 mentions)
Clone jc70a, by Agilent Technologies (1 mentions)
Fluorescent mounting media, by Agilent Technologies (1 mentions)
Millicell ez slide, by Merck Group (1 mentions)
Tryple, by Thermo Fisher Scientific (1 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (1 mentions)
Triton x 100, by Merck Group (1 mentions)
24 well cell culture plate, by Corning (1 mentions)
3 protocols using mouse anti human cd31 antibody
1
Visualization of Endothelial Cell Vessel Networks
2
Immunostaining of Endothelial Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Endothelial cells were detached with TrypLE (Life Technologies, Carlsbad,
USA) and 5000–6000 cells replated into 8-well Millicell EZ slides (Merck,
Darmstadt, Germany). Forty-eight hours later, cells were fixed with 4%
paraformaldehyde (PFA) for 30 min, (blocked with (10% (v/v) goat serum, 30
min) and overlaid with mouse anti-human CD31 antibody for 60 min (clone
JC70A, 1:100, Dako, Santa Clara, USA), or mouse anti-podoplanin (clone
D2-40, 1:100, Dako). After washing with PBS-T (PBS + 0.05% (v/v) Tween20),
secondary anti-mouse IgG-AlexaFluor488 conjugate (1:200, Life Technologies,
CA, USA) was applied for 60 min followed by DAPI (10 ng/mL, Sigma-Aldrich)
for 10 min before being washed and coverslips mounted with Fluorescent
mounting media (Dako).
USA) and 5000–6000 cells replated into 8-well Millicell EZ slides (Merck,
Darmstadt, Germany). Forty-eight hours later, cells were fixed with 4%
paraformaldehyde (PFA) for 30 min, (blocked with (10% (v/v) goat serum, 30
min) and overlaid with mouse anti-human CD31 antibody for 60 min (clone
JC70A, 1:100, Dako, Santa Clara, USA), or mouse anti-podoplanin (clone
D2-40, 1:100, Dako). After washing with PBS-T (PBS + 0.05% (v/v) Tween20),
secondary anti-mouse IgG-AlexaFluor488 conjugate (1:200, Life Technologies,
CA, USA) was applied for 60 min followed by DAPI (10 ng/mL, Sigma-Aldrich)
for 10 min before being washed and coverslips mounted with Fluorescent
mounting media (Dako).
3
Coculture of ASC and HUVEC
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
ASC were plated in 24-well cell culture plates (Corning) at 10,000 cells/cm 2 in NG-RPMI medium for 5 days. HUVEC were seeded on top of ASC monolayers or as the control on gelatin-coated wells at 10,000 cells/cm 2 in NG-ECM or HG-ECM for 7 more days after which vascular networks had formed. Cells were washed with PBS and fixed in 2% paraformaldehyde in PBS at RT for 20 min. Cells were permeabilized with 0.5% Triton X-100 in PBS (Sigma-Aldrich) for 15 min. Subsequently, samples were incubated with goat-antihuman-RGS5 antibody (1:100; Santa Cruz)/rabbit anti-human-SM22a antibody (1:100; Abcam) and mouse-anti-human-CD31 antibody (1:100; Dako) for 90 min. Samples were washed with PBS and incubated with a cyanine3-conjugated-rabbit-antibody to goat-IgG for RGS5/cyanine3-conjugated-donkey-antibody to rabbit-IgG for SM22a (1:300; Life Technologies) and fluorescein-conjugated-donkey-antibody to mouse-IgG (for CD31, 1:300; Life Technologies) for 45 min. Stained samples were mounted with PBS. Imaging was performed with a highend fully motorized Zeiss AxioObserver Z1 epifluorescent microscope (TissueGnosticsTissue Faxs). The coculture experiment was repeated with HUVEC that were lentivirally tagged to express EGFP (green) and ASC lentivirally tagged with dTomato (red) to monitor the origin cells irrespective of phenotypes changes during coculture.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!