Fluorescent labeling of platelets with polyanionic fluorescein derivatives

Area of Science:

• Hematology
• Cell Biology
• Biochemistry

Background:

• Platelet function is crucial in hemostasis and thrombosis.
• Accurate methods for labeling and quantifying platelets are essential for research.
• Fluorescein derivatives offer potential for fluorescent labeling of cells.

Purpose of the Study:

• To determine if polyanionic fluorescein derivatives can label resting and activated platelets from hamsters and humans.
• To assess the utility of these derivatives for platelet quantification and visualization.

Main Methods:

• Incubation of hamster and human platelets (resting and activated) with 5,6-carboxyfluorescein and calcein.
• Analysis using flow cytometry for quantification.
• Visualization via confocal microscopy to determine fluorophore localization.

Main Results:

• Both 5,6-carboxyfluorescein and calcein successfully labeled resting and activated platelets from both species.
• Labeling intensity and the proportion of labeled platelets increased linearly with fluorophore concentration.
• Confocal microscopy showed colocalization with the platelet glycocalyx and intracellular sequestration of the fluorophores.

Conclusions:

• 5,6-Carboxyfluorescein and calcein are effective fluorescent labels for hamster and human platelets, both resting and activated.
• The labeling mechanism involves avidity to the platelet glycocalyx and cellular sequestration.
• These findings enable reliable platelet quantification by flow cytometry and visualization by confocal microscopy.