Platelets confound the measurement of extracellular miRNA in archived plasma.
Extracellular miRNAs are detectable in biofluids and represent a novel class of disease biomarker. Although many studies have utilized archived plasma for miRNA biomarker discovery, the effects of processing and storage have not been rigorously studied. Previous reports have suggested plasma samples are commonly contaminated by platelets, significantly confounding the measurement of extracellular miRNA, which was thought to be easily addressed by additional post-thaw plasma processing. In a case-control study of archived plasma, we noted a significant correlation between miRNA levels and platelet counts despite post-thaw processing. We thus examined the effects of a single freeze/thaw cycle on microparticles (MPs) and miRNA levels, and show that a single freeze/thaw cycle of plasma dramatically increases the number of platelet-derived MPs, contaminates the extracellular miRNA pool, and profoundly affects the levels of miRNAs detected. The measurement of extracellular miRNAs in archived samples is critically dependent on the removal of residual platelets prior to freezing plasma samples. Many previous clinical studies of extracellular miRNA in archived plasma should be interpreted with caution and future studies should avoid the effects of platelet contamination.
Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.
Adam J
AJ
Mitchell
Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.
Warren D
WD
Gray
Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.
Salim S
SS
Hayek
Department of Biostatistics, Rollins School of Public Health, Emory University, Atlanta, GA, USA.
Yi-An
YA
Ko
Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.
Sheena
S
Thomas
Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.
Kim
K
Rooney
Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.
Mosaab
M
Awad
Department of Pathology, Emory University, Atlanta, GA, USA.
John D
JD
Roback
Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.
Arshed
A
Quyyumi
Division of Cardiology, Department of Medicine, Emory University, Atlanta, GA, USA.|Section of Cardiology, Atlanta VA Medical Center, Decatur, GA, USA.
Charles D
CD
Searles
D001792:Blood Platelets; D055252:Cell-Derived Microparticles / Q000235:genetics; D005434:Flow Cytometry; D005615:Freezing; D006801:Humans; D035683:MicroRNAs / Q000097:blood* / Q000302:isolation & purification; D010976:Platelet Count; D013048:Specimen Handling / Q000379:methods*
D035683:MicroRNAs