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RESEARCH PRODUCT
Flow Cytometric Analysis of Calcium Mobilization in Whole‐Blood Platelets
Maria-josé GonçalvesFilipe SansonettyMaria-do-céu MonteiroJosé-enrique O'connorsubject
Blood PlateletsPlatelet Membrane Glycoprotein IIbHistologyStimulation030204 cardiovascular system & hematologyBiochemistryFlow cytometry03 medical and health sciences0302 clinical medicineThrombinMethodsmedicineAnimalsHumansPlateletCalcium SignalingPlatelet activationFluorescent Dyes030304 developmental biologyWhole blood0303 health sciencesAniline Compoundsmedicine.diagnostic_testChemistryAntibodies MonoclonalGeneral MedicineFlow CytometryIn vitro3. Good healthKineticsMedical Laboratory TechnologyXanthenesBiochemistryBiophysicsCalciumIntracellularmedicine.drugdescription
Flow cytometry provides a convenient method to evaluate platelet activation by following the kinetics of intracellular free Ca2+, using sensitive fluorescent indicators that can be loaded into intact cells. Moreover, in the clinical setting, whole-blood techniques have obvious advantages to avoid artifactual platelet activation and allow the maintenance of near-physiological conditions. This unit describes a fast and sensitive flow cytometric procedure using the Ca2+-sensitive dye fluo-3 AM and the platelet-specific antibody CD41-PE to determine the kinetics of intracellular Ca2+ mobilization in whole-blood platelets with minimal manipulation of the samples. The technique may be applied to reveal fast and transient increases in cytosolic calcium upon platelet stimulation with the agonists ADP and thrombin. This protocol provides a simple and sensitive tool to assess in vitro the time course and intensity of signal-transduction responses to agonists under near-physiological conditions, and should be broadly applicable to studies of platelet reactivity.
| year | journal | country | edition | language |
|---|---|---|---|---|
| 2003-04-01 | Current Protocols in Cytometry |