Kinetx: A Combined Flow Cytometry Assay and Analysis Software Framework to Quantitatively Measure and Categorize Platelet Activation in Real-time

Platelets react rapidly to vascular injury and undergo activation in response to a range of stimuli to limit blood loss. Many platelet function tests measure endpoint responses after a defined time period and not the rate of platelet activation. However, the rate at which platelets convert extracell...

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Bibliographic Details
Published in:	Journal of visualized experiments no. 176
Main Authors:	Dunster, Joanne L, Mitchell, Jo L, Mohammed, Yasmin M M, Taylor, Kirk A, Gibbins, Jonathan M, Jones, Chris I
Format:	Journal Article
Language:	English
Published:	United States 07-10-2021
Subjects:	Blood Platelets - metabolism Flow Cytometry Hemostasis Platelet Activation Software
Online Access:	Get more information
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Summary:	Platelets react rapidly to vascular injury and undergo activation in response to a range of stimuli to limit blood loss. Many platelet function tests measure endpoint responses after a defined time period and not the rate of platelet activation. However, the rate at which platelets convert extracellular stimuli into a functional response is an essential factor in determining how efficiently they can respond to injury, bind to a forming thrombus, and signal to recruit other platelets. This paper describes a flow cytometry-based platelet function assay that enables simultaneous data acquisition and sample stimulation and utilizes newly developed bespoke open-source software (Kinetx) to enable quantitative kinetic measurements of platelet granule release, fibrinogen binding, and intracellular calcium flux. Kinetix was developed in R so that users can alter parameters such as degree of smoothing, identification of outlying data points, or time scales. To aid users unfamiliar with the R environment, Kinetix analysis of data can be performed by a single command. Together, this allows real-time platelet activation metrics, such as rate, acceleration, time to peak-rate, time to peak-calcium, and qualitative shape changes, to be accurately and reproducibly measured and categorized. Kinetic measurements of platelet activation give a unique insight into platelets' behavior during the first stages of activation and may provide a method of predicting the recruitment of platelets into a forming thrombus.
ISSN:	1940-087X
DOI:	10.3791/62947