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Aqueous Biphasic 3D Cell Culture Micro-Technology

Atefi, Ehsan
http://rave.ohiolink.edu/etdc/view?acc_num=akron1443112692

Abstract Details

2015, Doctor of Philosophy, University of Akron, Biomedical Engineering.
Three-dimensional (3D) cell culture technologies have gained a considerable momentum in compound screening applications to identify novel anti-cancer drugs. Increasing evidence shows substantial differences between responses of cancer cells to drug compounds in monolayer cultures (2D) traditionally used in drug discovery and in vivo during preclinical tests. 3D cell cultures more closely resemble tumors in terms of close cell-cell and cell-extracellular matrix interactions, non-uniform distribution of soluble factors, and presence of hypoxic cells. As such, they provide a relevant tumor model to elicit more realistic responses from cells treated with drugs. Screening of libraries of compounds to identify novel drugs requires high throughput 3D culture platforms that produce consistently sized cancer cell spheroids and allow convenient drug testing and analysis of cellular responses. In this study, we introduce a novel, automated technology for 3D culture of cancer cell spheroids in a high throughput format. Aqueous two-phase systems (ATPS) are used for producing spheroids with robotic tools and standard equipment. ATPS are formed by mixing appropriate mass concentrations of two biocompatible polymers such as dextran (DEX) and polyethylene glycol (PEG). A nano-liter drop of the denser aqueous DEX phase containing cancer cells is robotically dispensed into each well of a non-adherent 96-well plate containing the immersion PEG phase solution. A round drop containing cells forms at the bottom of the well while overlaid with the aqueous PEG phase. Cells remain in the DEX drop and form a spheroid, which receives nutrients from the immersion phase through diffusion into the drop. The fidelity of the ATPS spheroid culture technology depends on favorable partition of cells to the DEX drop. We investigate partition of cancer cells in ATPS and demonstrate the effect of interfacial tension between the two aqueous phases on the distribution of cells in ATPS. To facilitate this study, we determine ultralow interfacial tensions of ATPS using an axisymmetric drop shape analysis (ADSA) methodology with sessile and pendant drops and develop mathematical criteria for reliable measurements. To develop a fundamental understanding of the role of interfacial tension of ATPS in cell partition, we develop a theoretical model to predict the energy associated with displacement of a particle (cell) in ATPS. This model, which also uses our contact angle measurements with ATPS/cell systems as an input, shows that a very small interfacial tension, i.e., on the scale of ~30 µJ/m2, results in a minimum free energy when cells locate in the bottom DEX phase, corroborating with our experimental cell partition data and spheroid formation with ATPS. Finally, the utility of this new technology for compound screening is demonstrated by high throughput testing of several anti-cancer drugs against spheroids of skin and breast cancer cells. Incorporating this robotic technology in the oncology drug discovery pipeline will expedite discovery of novel anti-cancer drugs with a relevant tumor model.
Hossein Tavana (Committee Chair)
183 p.
Biomedical Engineering
3D Cell Culture, Aqueous Two Phase Systems, Anti-cancer Screening, Interfacial Tension, Surface Tension, Contact Angle, Cell Surface Property, Cancer Cell Fractionation

Recommended Citations

Citations

  • Atefi, E. (2015). Aqueous Biphasic 3D Cell Culture Micro-Technology [Doctoral dissertation, University of Akron]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=akron1443112692

    APA Style (7th edition)

  • Atefi, Ehsan. Aqueous Biphasic 3D Cell Culture Micro-Technology. 2015. University of Akron, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=akron1443112692.

    MLA Style (8th edition)

  • Atefi, Ehsan. "Aqueous Biphasic 3D Cell Culture Micro-Technology." Doctoral dissertation, University of Akron, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=akron1443112692

    Chicago Manual of Style (17th edition)

akron1443112692
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This open access ETD is published by University of Akron and OhioLINK.