Platelet activation was measured by whole-blood flow cytometry as previously described (43 (link)) by quantifying the platelet membrane expression of the α-granule protein P-selectin (CD62P) and the binding of fibrinogen to the activated αIIbβ3 receptor (GPIIbIIIa complex). The following antibodies were used to incubate samples from the whole-blood ex vivo assay: phycoerythrin (PE)-labeled anti-CD62P (Bio-Legend, San Diego, CA), fluorescein isothiocyanate (FITC)-labeled antifibrinogen (F0111-FITC; DAKO Ltd., High Wycombe, United Kingdom), and PC7-labeled anti-CD61 (platelet glycoprotein IIIa; Beckman Coulter, Inc., Miami, FL), the last as a platelet identification marker. The percentages of CD62P and fibrinogen in CD61-positive events were determined. Formation of PMC was measured by incubating samples with PC7-labeled anti-CD61 and PE-labeled anti-CD14 (a glycosylphosphatidylinositol [GPI]-linked membrane glycoprotein; Bio-Legend). After 20 min of incubation, OptiLyse B (Beckman Coulter, Inc., Fullerton, CA) was added to lyse erythrocytes. PMC formation was determined by quantifying the mean fluorescence intensity (MFI) of CD14+ cells that were also positive for the platelet identification marker CD61. All samples were measured using an FC500 flow cytometer (Beckman Coulter, Inc.).
Pc7 labeled anti cd61
Manufactured by Beckman Coulter
Sourced in United Kingdom, United States
The PC7-labeled anti-CD61 is a flow cytometry antibody that specifically binds to the CD61 antigen expressed on the surface of platelets. It is designed for the identification and enumeration of CD61-positive cells in biological samples.
Automatically generated - may contain errors
2 protocols using pc7 labeled anti cd61
1
Quantifying Platelet Activation and PMC Formation
2
Evaluating Platelet Activation by P-Selectin Expression
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Platelet reactivity reflected by P-selectin expression as markers of platelet degranulation was determined by flow cytometry.13 (link) Platelet reactivity was measured in unstimulated samples and after ex vivo platelet stimulation by ADP (low dose: 7.8 µM and high dose: 31.2 µM; Sigma-Aldrich, St Louis, MO, USA). Whole blood was added to a mixture of 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid-buffered saline and saturated concentrations of PE-labeled anti-CD62P (P-selectin; BioLegend, San Diego, CA, USA) and PC7-labeled anti-CD61 (platelet identification marker; Beckman Coulter, Villepinte, France). After 20 minutes of incubation at room temperature, 0.2% paraformaldehyde was added and samples were analyzed. Platelets were gated based on their forward- and sideward-scatter properties and positivity for CD61, which was defined as a median fluorescence intensity (MFI) exceeding that of its matched isotype control.
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