392224 Stabilities of exosomes derived from mammalian cells: nanoparticle morphology and functional RNAs

Wednesday, November 19, 2014: 9:46 AM
202 (Hilton Atlanta)
Yueting Wu1, Vanessa Cuppett1 and David J. Klinke2, (1)Department of Chemical Engineering, West Virginia University, Morgantown, WV, (2)Microbiology, Immunology and Cell Biology, West Virginia University, Morgantown, WV

Currently, it has been widely accepted that most mammalian cells release nanoscaled membrane vesicles called exosomes, transferring functional molecules of proteins and RNAs derived from the parent cells. The specific function of exosomes in regulating immunity and promoting oncogenes remains to be investigated. This study assessed the stabilities of exosomes and the functional RNA molecules enclosed. Scanning electron microscope (SEM) was used to inspect the morphology of exosomes, assessing the changes in exosomes due to different production and storage conditions. Optimal SEM sample processing method was investigated for fast and accurate analysis of intact exosome particles using SEM. As a comparison to exosomes, apoptotic vesicles were also inspected and analyzed using SEM. The quality and stability of RNAs contained within the exosomes was characterized using Agilent Bioanalyzer, showing the changes of RNA molecules enclosed in exosomes and stored for days, months, and up to 2 years under -80 degree C. Exosomal RNAs isolated from exosomes that were produced from serum-containing or serum-free conditions, respectively, and at medium incubation time 12 to 48 hours, were analyzed and showed stable and characteristic spectrum for various exosomal nucleotides. Collectively the results suggested improved methods for processing and storage of exosomes that preserve morphological and biochemical characteristics. In order to reproducibly investigate exosome functions,  it is essential to maintain a near native state of exosomes and preserve their functional molecules during sample handling, analyzing and storing.

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