Receptor expression of typical endothelial markers was measured by flow cytometry at passage 3–5 for three matched donors. Cells were incubated with antibodies for 30 min at 4°C, washed with PBS supplemented with 0.1% bovine serum albumin and 0.1% sodium azide and then resuspended in the same buffer for flow cytometric analysis. The fluorescence was measured on a BD FACS Calibur and the results analyzed by Cell Quest software (Becton Immunocytometry Systems, Mountain View, CA, USA). PE-labelled antibodies and corresponding isotypes (mouse anti human) were used from BD Pharming unless otherwise stated: CD31 (WM59, IgG1), CD34 (581, IgG1), CD54 (HA58, IgG1), CD105 (SN6, IgG1, Invitrogen), CD106 (51-10C9, IgG1), CD144 (55-7H1, IgG1), CD309 (89106, IgG1), CD181 (5A12, IgG2b), CD182 (6C6, IgG1), CD183 (1C6, IgG1), CD184 (12G5, IgG2a, R&D systems), CD192 (48607, IgG2b, R&D systems), IgG1 (MOPC-21), IgG2a (G155-178), IgG2b (27–35), FcR Blocker (Miltenyi Biotec).
Cd144
Manufactured by R&D Systems
Sourced in United States
CD144 is a cell surface glycoprotein that functions as an endothelial cell-cell adhesion molecule. It plays a role in the formation and maintenance of tight junctions between endothelial cells.
Automatically generated - may contain errors
Lab products found in correlation
Fcr blocker, by Miltenyi Biotec (1 mentions)
Cd106, by Thermo Fisher Scientific (1 mentions)
Cd309, by R&D Systems (1 mentions)
Cd105, by Thermo Fisher Scientific (1 mentions)
Filaggrin, by Abcam (1 mentions)
Pecam 1, by Thermo Fisher Scientific (1 mentions)
Hoechst 33258, by Merck Group (1 mentions)
Ve cadherin, by R&D Systems (1 mentions)
Alexa fluor 488 conjugated ephb4, by R&D Systems (1 mentions)
Flowjotm 10, by BD (1 mentions)
Cd133, by R&D Systems (1 mentions)
Cd133, by BioLegend (1 mentions)
Trustain fcx, by BioLegend (1 mentions)
Facslyric, by BD (1 mentions)
Ficoll paque plus, by GE Healthcare (1 mentions)
3 protocols using cd144
1
Endothelial Marker Expression Analysis
2
Indirect Immunolabeling of Human Dermal Microvascular Endothelial Cells
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Indirect immunolabelings were performed on the optimal cutting temperature (OCT) compound cross-sections (5 μm and 25 μm). The primary antibodies were used as follows: vascular endothelial cadherin (VE-cadherin, CD144, R&D systems; 1:100), von Willebrand factor (vWF, DAKO; 1:200) antibody, platelet-endothelial cellular adhesion molecule-1 (PECAM-1, CD31, Invitrogen; 1:100) antibody, type IV collagen (Coll IV, Abcam; 1:400) antibody, filaggrin (Abcam; 1:500), loricrin (Cedarlane; 1:1000), keratin 10 (Monosan; 1:200), and KI67 (BD pharmingen; 1:300). Alexa 488 and Alexa 594 were used as secondary antibodies, but the secondary antibody was mixed with Hoechst 33258 (Sigma-Aldrich; 1/100) to stain the cell nuclei. The Coll IV synthesis of the ECs was analyzed under the same conditions as the SA process. HDMECs were seeded into nongelatinized flasks and cultivated in the same medium mixtures as the substitutes for the same time intervals. The monolayer ECs were fixed and stained with CD31 and Coll IV.
3
Endothelial Cell Evaluation in Kawasaki Disease
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Peripheral blood samples (2 mL each) were collected from the KD patients before IVIG and healthy children. PBMC were extracted using Ficoll-Paque PLUS (GE Healthcare Life Science) according to the manufacturer’s protocol. CECs were evaluated using a panel of antibodies: BV421-conjugated CD34 (BD Pharmingen, San Jose, CA, USA), PerCP/Cyanine5.5-conjugated CD45 (BioLegend, San Diego, CA, USA), phycoerythrin (PE)-conjugated CD133 (BioLegend), and Alexa Fluor® 647-conjugated CD144 (BioLegend). Alexa Fluor 488-conjugated EphB4 (R&D Systems, Minneapolis, MN, USA) was used to analyze the level of EphB4. CECs were identified as cells lacking CD45 expression, positive for CD34, positive for CD144, and negative for CD133 (CD45−/CD34+/CD144+/CD133−).
PBMC were blocked with TruStain FcX™ (BioLegend) for 10 min and then incubated with CD34, CD45, CD133, CD144, and Alexa Fluor 488-conjugated EphB4 (R&D Systems, MN, USA) for 15 min in the dark. Fluorescence intensity was examined by BD FACSLyric (BD) and analyzed by flow cytometry software FlowJoTM 10 (BD).
PBMC were blocked with TruStain FcX™ (BioLegend) for 10 min and then incubated with CD34, CD45, CD133, CD144, and Alexa Fluor 488-conjugated EphB4 (R&D Systems, MN, USA) for 15 min in the dark. Fluorescence intensity was examined by BD FACSLyric (BD) and analyzed by flow cytometry software FlowJoTM 10 (BD).
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