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Article

Characterization of Extracellular Vesicles Labelled with a Lipophilic Dye Using Fluorescence Nanoparticle Tracking Analysis

1
Institute of Veterinary Medicine and Animal Sciences, Estonian University of Life Sciences, Kreutzwaldi 62, 51006 Tartu, Estonia
2
Department of Pathophysiology, Institute of Biomedicine and Translational Medicine, University of Tartu, Ravila St. 14b, 50411 Tartu, Estonia
3
Department of Anatomy, Faculty of Medicine, University of Peradeniya, Peradeniya 20400, Sri Lanka
4
Institute of Chemistry, University of Tartu, Ravila St. 14a, 50411 Tartu, Estonia
5
Academic Unit of Reproductive and Developmental Medicine, Department of Oncology and Metabolism, Medical School, University of Sheffield, Sheffield S10 2SF, UK
*
Author to whom correspondence should be addressed.
Academic Editor: Shiro Suetsugu
Membranes 2021, 11(10), 779; https://doi.org/10.3390/membranes11100779
Received: 8 September 2021 / Revised: 4 October 2021 / Accepted: 6 October 2021 / Published: 12 October 2021
(This article belongs to the Collection Feature Papers in Membranes in Life Sciences)
Research on extracellular vesicles (EVs) has intensified over the past decade, including fluorescent membrane labeling of EVs. An optimal fluorescent method requires the size of EVs to be preserved after labeling. Lipophilic fluorescent dyes, such as CellMask™ Green (CMG), have been widely used for this purpose. Here, we investigated conditions affecting the optimum CMG labeling of EVs derived from human choriocarcinoma cells (JAr) and different biological fluids using fluorescence NTA (fl-NTA). The effect of CMG labeling on the size, concentration and zeta potential (ZP) on JAr EVs purified with different methods were measured along with biological fluid-derived EVs. With the increase of CMG dye concentration, a significant decrease in the mean size of fluorescent nanoparticles (fl-NPs) was observed. The ZP of fl-NPs originating from JAr cells with the lowest and highest dye concentrations showed a significant shift towards more and less negative ZP values, respectively. Differences in the concentration of fl-NPs were observed for JAr EVs purified using size-exclusion chromatography (SEC) alone and SEC in combination with tangential flow filtration. The proportion of CMG labeling of NPs varied across different biological sources. CMG labeling may be a reliable technique for the detection of EVs using fl-NTA. View Full-Text
Keywords: fluorescence; NTA; lipophilic dyes; extracellular vesicles; zeta potential; detergent fluorescence; NTA; lipophilic dyes; extracellular vesicles; zeta potential; detergent
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Figure 1

  • Supplementary File 1:

    ZIP-Document (ZIP, 971 KiB)

  • Externally hosted supplementary file 1
    Link: https://zenodo.org/deposit/5495621
    Description: Video S1: NTA data of JAr EVs only in 1x PBS solution.
  • Externally hosted supplementary file 2
    Link: https://zenodo.org/deposit/5495621
    Description: Video S2: Data of JAr EVs with CMG (100ng/mL) in 1x PBS solution measured at the fluorescent mode of NTA.
  • Externally hosted supplementary file 3
    Link: https://zenodo.org/deposit/5495621
    Description: Video S3: Data of JAr EVs with CMG (100ng/mL) in 1x PBS solution measured at scatter mode of NTA.
MDPI and ACS Style

Midekessa, G.; Godakumara, K.; Dissanayake, K.; Hasan, M.M.; Reshi, Q.U.A.; Rinken, T.; Fazeli, A. Characterization of Extracellular Vesicles Labelled with a Lipophilic Dye Using Fluorescence Nanoparticle Tracking Analysis. Membranes 2021, 11, 779. https://doi.org/10.3390/membranes11100779

AMA Style

Midekessa G, Godakumara K, Dissanayake K, Hasan MM, Reshi QUA, Rinken T, Fazeli A. Characterization of Extracellular Vesicles Labelled with a Lipophilic Dye Using Fluorescence Nanoparticle Tracking Analysis. Membranes. 2021; 11(10):779. https://doi.org/10.3390/membranes11100779

Chicago/Turabian Style

Midekessa, Getnet, Kasun Godakumara, Keerthie Dissanayake, Mohammad Mehedi Hasan, Qurat Ul Ain Reshi, Toonika Rinken, and Alireza Fazeli. 2021. "Characterization of Extracellular Vesicles Labelled with a Lipophilic Dye Using Fluorescence Nanoparticle Tracking Analysis" Membranes 11, no. 10: 779. https://doi.org/10.3390/membranes11100779

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