Exosome-Mediated miRNA Delivery Protocol

Resuspended exosomes were diluted in the Gene Pulser electroporation buffer (Bio-Rad Laboratories, CA) in 1 : 1 ratio. 1 μmol of mouse miR-132 mimic (Ambion, NY) or inhibitor was added to 200 μl of exosome sample. The mixtures were transferred into cold 0.2 cm electroporation cuvettes and electroporated at 150 V/100 μF capacitance using a Gene Pulser II system (Bio-Rad Laboratories, CA) as described previously [18 (link)]. After removing the free-floating miRNA mimic, exosomes were reisolated using ultracentrifugation. The final pellet (exosome) was resuspended in PBS and stored at −80°C.

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Ma T., Chen Y., Chen Y., Meng Q., Sun J., Shao L., Yu Y., Huang H., Hu Y., Yang Z., Yang J, & Shen Z. (2018). MicroRNA-132, Delivered by Mesenchymal Stem Cell-Derived Exosomes, Promote Angiogenesis in Myocardial Infarction. Stem Cells International, 2018, 3290372.

Publication 2018

Gene Pulser electroporation buffer Gene Pulser II system

Buffer Cold Electroporation Exosome Gene Gene system Mirna Mouse mir 132 Ultracentrifugation

Corresponding Organization : First Affiliated Hospital of Soochow University

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Variable analysis

independent variables

Addition of 1 μmol of mouse miR-132 mimic (Ambion, NY) or inhibitor to 200 μl of exosome sample

dependent variables

Exosome characteristics after electroporation and re-isolation

control variables

Gene Pulser electroporation buffer (Bio-Rad Laboratories, CA) used for dilution of resuspended exosomes
Electroporation at 150 V/100 μF capacitance using a Gene Pulser II system (Bio-Rad Laboratories, CA)
Ultracentrifugation used for re-isolation of exosomes after removing free-floating miRNA mimic
Final exosome pellet resuspended in PBS and stored at −80°C

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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