Lyophilization based isolation of exosomes

Authors

Rida E Maria Qazi, Aga Khan UniversityFollow
Zahra Sajid, Aga Khan UniversityFollow
Chunqiu Zhao, Southeast University, China
Irfan Hussain, Aga Khan UniversityFollow
Fizza Iftikhar, Aga Khan University
Muhammad Jameel, Aga Khan UniversityFollow
Fawad Ur Rehman, Aga Khan UniversityFollow
Afsar Ali Mian, Aga Khan UniversityFollow

Document Type

Article

Department

Centre for Regenerative Medicine and Stem Cell Research

Abstract

Exosomes are nanoscale extracellular vesicles which regulate intercellular communication. They have great potential for application in nanomedicine. However, techniques for their isolation are limited by requirements for advanced instruments and costly reagents. In this study, we developed a lyophilization-based method for isolating exosomes from cultured cells. The isolated exosomes were characterized for protein content using Bradford assay, and for size distribution and shape using scanning electron microscopy (SEM) and nanoparticles tracking analysis (NTA). In addition, CD63, CD9, CD81, HSP70 and TSG101 were evaluated as essential exosomal surface markers using Western blot. Drug loading and release studies were performed to confirm their drug delivery properties using an in vitro model. Exosomes were also loaded with commercial dyes (Cy5, Eosin) for the evaluation of their drug delivery properties. All these characterizations confirmed successful exosome isolation with measurements of less than 150 nm, having a typical shape, and by expressing the known exosome surface protein markers. Finally, tyrosine kinase inhibitors (dasatinib and ponatinib) were loaded on the exosomes to evaluate their anticancer effects on leukemia cells (K562 and engineered Ba/F3-BCR-ABL) using MTT and Annexin-PI assays. The expression of MUC1 protein on the exosomes isolated from MCF-7 cells also indicated that their potential diagnostic properties were intact. In conclusion, we developed a new method for exosome isolation from cultured cells. These exosomes met all the essential requirements in terms of characterization, drug loading and release ability, and inhibition of proliferation and apoptosis induction in Ph+ leukemia cells. Based on these results, we are confident in presenting the lyophilization-based exosome isolation method as an alternative to traditional techniques for exosome isolation from cultured cells.

Comments

Pagination are not provided by the author/publisher.

Publication (Name of Journal)

International Journal of Molecular Sciences

Recommended Citation

Qazi, R. M., Sajid, Z., Zhao, C., Hussain, I., Iftikhar, F., Jameel, M., Rehman, F. U., Mian, A. A. (2023). Lyophilization based isolation of exosomes. International Journal of Molecular Sciences, 24(13).
Available at: https://ecommons.aku.edu/crm/78