Extracellular vesicles from human plasma and serum are carriers of extravesicular cargo—Implications for biomarker discovery

Abstract

Extracellular vesicles (EVs) in human blood are a potential source of biomarkers. To which extent anticoagulation affects their concentration, cellular origin and protein composition is largely unexplored. To study this, blood from 23 healthy subjects was collected in acid citrate dextrose (ACD), citrate or EDTA, or without anticoagulation to obtain serum. EVs were isolated by ultracentrifugation or by size-exclusion chromatography (SEC) for fluorescence-SEC. EVs were analyzed by micro flow cytometry, NTA, TEM, Western blot, and protein mass spectrometry. The plasma EV concentration was unaffected by anticoagulants, but serum contained more platelet EVs. The protein composition of plasma EVs differed between anticoagulants, and between plasma and serum. Comparison to other studies further revealed that the shared EV protein composition resembles the "protein corona" of synthetic nanoparticles incubated in plasma or serum. In conclusion, we have validated a higher concentration of platelet EVs in serum than plasma by contemporary EV methods. Anticoagulation should be carefully described (i) to enable study comparison, (ii) to utilize available sample cohorts, and (iii) when preparing/selecting biobank samples. Further, the similarity of the EV protein corona and that of nanoparticles implicates that EVs carry both intravesicular and extravesicular cargo, which will expand their applicability for biomarker discovery.

Circulating Melanoma-Derived Extracellular Vesicles: Impact on Melanoma Diagnosis, Progression Monitoring, and Treatment Response

“…However, the true proportion of EVs in the circulation that is attributable to the tumour is not known. Serum is deemed to have a higher proportion of platelet-derived EVs in comparison to plasma [ 107 ]. Beyond this, even the choice of anticoagulant used for plasma preparation may alter the EV populations.…”

“…Beyond this, even the choice of anticoagulant used for plasma preparation may alter the EV populations. For example, citrate use may result in a higher concentration of platelet-derived EVs [ 107 ], but a lower yield of EVs overall [ 108 ].…”

Kinetics and Topology of DNA Associated with Circulating Extracellular Vesicles Released during Exercise

“…EVs compose of a phospholipid bilayer membrane that enables protected transport of bioactive cargo like proteins, lipids, and nucleic acids between cells and tissues inside the vesicle. Additionally, macromolecules can be attached outside of the EV as part of the vesicle corona [ 22 , 23 ].…”

Regulation of Serum Exosomal MicroRNAs in Mice Infected with Orientia tsutsugamushi

“…Some of the miRNA regulations were evident as early as day 2 post infection ( Figure 4 A), even before the appearance of bacteremia on day 4 ( Figure 1 A), suggesting their potential value as biomarkers for early diagnosis of scrub typhus. While both serum and plasma have been extensively used to study exosomal miRNAs and to identify potential biomarkers [ 35 ], compositions of extracellular vesicles in serum might be affected during blood coagulation due to involvement of platelets [ 36 , 37 ]. As serum is investigated in the present study, it remains to be determined whether exosomes isolated from platelet-free plasma will share similar miRNA regulations and functional properties.…”