CD41/PECy7 (BioLegend Cat. No. 303718) was used as an identity marker for platelets, PAC-1/FITC (BioLegend Cat. No. 362804) for glycoprotein GP IIbIIIa and CD62/PE (BioLegend Cat. No. 304906) for P-selectin were used as activation markers. IgG1 k (BioLegend Cat. No. 400125), FITC Mouse IgM k Isotype (BioLegend Cat. No. 401605) and Mouse IgG1 k Isotype (BioLegend Cat. No. 400111) were used as isotype control, respectively. The gating strategy of the cell populations was performed according to previously reported by the research group in García-Larragoiti et al. [22 (link)]. Dark conditions and minimal handling were used during the assay to avoid external activation of platelets. Adenosine Di Phosphate (ADP) (20 µM) for 20 min, collagen (20 µM) for 30 min, and epinephrine (EPI) (100 µM) for 40 min were used as positive platelet activation controls [27 (link)]. Concentrations were used following the instructions suggested by the supplier PAR/PAK II® BIO/DATA CORPORATION (Horsham, PA, USA). The acquisition was performed in a CytoFLEX, BECKMAN COULTER® (Brea, CA, USA). Results were analyzed using FlowJo v 10.8.0.
Cd62 pe
Manufactured by BioLegend
CD62/PE is a fluorescently-labeled antibody that binds to the CD62 antigen. CD62 is a cell surface glycoprotein involved in cell adhesion. The PE (Phycoerythrin) fluorescent label allows for the detection and analysis of CD62-expressing cells using flow cytometry.
Automatically generated - may contain errors
2 protocols using cd62 pe
1
Platelet Activation Markers Evaluation
2
Platelet Activation Assay by Flow Cytometry
Check if the same lab product or an alternative is used in the 5 most similar protocols
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PRP was obtained by centrifugation, diluted in Tyrodes buffer and incubated for
20 min with CD41/PECy7 (BioLegend Cat. No. 303718) as identity marker,
PAC-1/FITC (BioLegend Cat. No. 362804) and CD62/PE (BioLegend Cat. No. 304906),
were both used as activation markers (glycoprotein αIIbβIII and P-selectin,
respectively). IgG1 k (BioLegend Cat. No. 400125), FITC Mouse IgM k Isotype
(BioLegend Cat. No. 401605) and, Mouse IgG1 k Isotype (BioLegend Cat. No.
400111) were used as isotype control respectively. Subsequently, platelets were
fixed with paraformaldehyde at 4%. Dark conditions and minimal handling were
used during the assay to avoid external activation of platelets. As positive
controls of platelet activation we used known activation agonists ADP, collagen
and epinephrine and the acquisition was performed by flow cytometry as reported
before by our group.10 (link) Results were analyzed using FlowJo v 10.8.0.
20 min with CD41/PECy7 (BioLegend Cat. No. 303718) as identity marker,
PAC-1/FITC (BioLegend Cat. No. 362804) and CD62/PE (BioLegend Cat. No. 304906),
were both used as activation markers (glycoprotein αIIbβIII and P-selectin,
respectively). IgG1 k (BioLegend Cat. No. 400125), FITC Mouse IgM k Isotype
(BioLegend Cat. No. 401605) and, Mouse IgG1 k Isotype (BioLegend Cat. No.
400111) were used as isotype control respectively. Subsequently, platelets were
fixed with paraformaldehyde at 4%. Dark conditions and minimal handling were
used during the assay to avoid external activation of platelets. As positive
controls of platelet activation we used known activation agonists ADP, collagen
and epinephrine and the acquisition was performed by flow cytometry as reported
before by our group.10 (link) Results were analyzed using FlowJo v 10.8.0.
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