Prospects and limitations of antibody-mediated clearing of lipoproteins from blood plasma prior to nanoparticle tracking analysis of extracellular vesicles

• Published in: Journal of extracellular vesicles 2017-12, Vol.6 (1), p.1308779-n/a
• Main Authors: Mørk, Morten, Handberg, Aase, Pedersen, Shona, Jørgensen, Malene M., Bæk, Rikke, Nielsen, Morten K., Kristensen, Søren R.
• Format: Article
• Language: English
• Subjects: Antibodies; Apolipoprotein B; Apolipoproteins; Cameras; Chromatography; chylomicrons; Diabetes; Enzyme-linked immunosorbent assay; extracellular vesicle array; extracellular vesicle purification; Extracellular vesicles; Fasting; Glycerol; Immunoglobulins; interference; Laboratory testing; Lipoproteins; Low density lipoprotein; low density lipoproteins; magnetic beads; Nanoparticle tracking analysis; Nanoparticles; Plasma; Preservation; Proteins; Purification; Ultracentrifugation; very low density lipoproteins; Viscosity
• ISSN: 2001-3078; 2001-3078
• DOI: 10.1080/20013078.2017.1308779

Introduction: Nanoparticle tracking analysis (NTA) enables measurement of extracellular vesicles (EVs) but lacks the ability to distinct between EVs and lipoproteins which are abundantly present in blood plasma. Limitations in ultracentrifugation and size exclusion chromatography applied for EV isolation may result in inadequate EV purification and preservation. In this proof of concept study, we aimed to evaluate the potential of antibody-mediated removal of lipoproteins from plasma prior to extracellular vesicle (EV) analysis by nanoparticle tracking analysis (NTA). Methods: Ten platelet-free plasma (PFP) samples from healthy fasting subjects were incubated with magnetic beads coated with antibodies against apolipoprotein B-48 and B-100 (ApoB). Plasma samples were analysed with NTA before and after application of the bead procedure. Four fasting PFP samples were analysed with an ELISA specific for human ApoB to estimate the degree of removal of lipoproteins and EV array analysis was used for identification of possible EV loss. Results: The magnetic bead separation procedure resulted in a median reduction of the particle concentration in plasma by 62% (interquartile range 32-72%). The mean size of the remaining particles generally increased. ApoB concentration was reduced to a level close to the background signal, whereas a median reduction of the EV content by 21% (range 8-43%) was observed. Conclusion: Anti-ApoB antibody coated magnetic beads may hold potential for removal of lipoproteins from human PFP prior to EV measurement by NTA but some artefactual effect and EV loss may have to be endured.