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The study of platelet aggregation using a microtiter plate reader ‒ methodological considerations

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Abstract

Optical aggregometry by 96-well plate assay, the microplate method, is a fast, efficient, and readily available method for measuring the pharmacological effects of antiplatelet drugs. Even though recent years have witnessed growing interest in adopting the microplate method for widespread use, it remains in the shadow of the standard light transmission aggregometry (LTA). Regardless of the method used, the results of platelet aggregation depend on a variety of factors and often vary among laboratories worldwide. While several methodological papers have examined the microplate method, no standards have been established, most likely because the approach is not used as a diagnostic tool. Currently, the microplate method is recommended by researchers to be used in conjunction with LTA or as an adjunct to LTA. This raises the question of whether an optimal protocol exists for microplate aggregometry, and what are the key considerations in a good experimental protocol for obtaining reliable results? This article attempts to address these questions by summarizing the knowledge accumulated in this field over the last three decades.

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Data availability

Data presented in Fig. 1 are available from the corresponding author upon reasonable request.

Abbreviations

4PL:

4-parameter logistic model

AA:

Arachidonic acid

ADP:

Adenosine 5’-diphosphate

ATP release assay:

Adenosine 5′-triphosphate release assay

EC50 :

Half maximal effective concentration

ELISA:

Enzyme-linked immunosorbent assay

HIV:

Human immunodeficiency virus

IC50 :

Half-maximal inhibitory concentration

LTA:

Light transmission aggregometry

PAR-4 agonist:

Proteinase-activated receptor-4 agonist

PARs:

Protease-activated receptors

PBS:

Phosphate-buffered saline

PM:

Prasugrel metabolite

PRP:

Platelet-rich plasma

TP receptor:

Thromboxane A2 receptor

TRAP:

Thrombin receptor-activating peptide

TXA2/TXB2 :

Thromboxane A2/thromboxane B2

U46619:

Stable thromboxane A2 receptor agonist

VWD:

Von Willebrand disease

VWF:

Von Willebrand factor

VWF: RCo:

Ristocetin cofactor assay

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Acknowledgements

The authors thank E. Lowczowski, B.Sc., Medical University of Łódź, for proofreading the article. Funding for this project was provided by the Medical University of Łódź (503/6-020-01/503-61-001-19-00).

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  1. Department of Haemostasis and Haemostatic Disorders, Chair of Biomedical Sciences, Medical University of Łódź, ul. Mazowiecka 6/8, Łódź, 92-215, Poland

    Magdalena Boncler & Jacek Golański

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  1. Magdalena Boncler
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MB: Conceptualization, Formal analysis, Investigation, Methodology, Visualization, Writing – original draft preparation; JG: Investigation, Writing – review & editing.

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Correspondence to Magdalena Boncler.

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Boncler, M., Golański, J. The study of platelet aggregation using a microtiter plate reader ‒ methodological considerations. Pharmacol. Rep 76, 328–337 (2024). https://doi.org/10.1007/s43440-024-00576-7

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