Functional Assessment of Platelet Dense Granule ATP Release: Optimization Using Minute Blood Volumes

Objectives: This study was undertaken to explore the feasibility of assessing platelet dense granule release in response to platelet stimuli, using less than 1 mL of whole blood (WB). Methods: Optimization of the luciferin-luciferase (LL) assay for ATP release, together with additional modifications...

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Bibliographic Details
Published inAmerican journal of clinical pathology Vol. 155; no. 6; pp. 863 - 872
Main Authors Cho, Joseph H, Wool, Geoffrey D, Tjota, Melissa Y, Gutierrez, Jocelyn, Mikrut, Krzysztof, Miller, Jonathan L
Format Journal Article
LanguageEnglish
Published Chicago Oxford University Press 01.06.2021
Subjects
Analysis
Blood platelets
Collagen
Formaldehyde
Luciferase
Platelets
Product introduction
Secretion
Thrombin
Ticagrelor
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Summary:Objectives: This study was undertaken to explore the feasibility of assessing platelet dense granule release in response to platelet stimuli, using less than 1 mL of whole blood (WB). Methods: Optimization of the luciferin-luciferase (LL) assay for ATP release, together with additional modifications, was applied to 1:10 diluted WB. Results: LL assay optimization using nonstirred 1:10 diluted WB resulted in dense granule ATP release in response to thrombin receptor-activating peptide (TRAP) of similar magnitude to that observed using stirred platelet-rich plasma. Stirring of the 1:10 diluted WB restored collagen-induced dense granule secretion. Addition of lyophilized, formalin-fixed platelets, together with stirring, restored dense granule secretion responsiveness to ADP TRAP, ADP, and collagen all stimulated ATP release in 1:10 diluted WB under the optimized conditions of this study at levels close to those observed using platelet-rich plasma. Blood sample reconstitution experiments offer hope that this assay may prove robust down to WB platelet counts as low as 50 x [10.sup.3]/[micro]L. Conclusions: Platelet dense granule release in response to a number of classic stimuli, including ADP, was accomplished from less than 1 mL WB with minimal specimen processing, using widely available reagents and instrumentation. Key Words: Platelet-function testing; Platelet dense granule release; ADP; Luciferin-luciferase ATP assay; Diluted whole blood; Formalin-fixed platelets; Collagen
ISSN:0002-9173
1943-7722
DOI:10.1093/AJCP/AQAA196