A Lab-on-a-chip system integrating dielectrophoretic detection and impedance counting units for chemotherapy guidance in leukemia

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2018
Demircan Yalçın, Yağmur
Precision medicine is defined as therapies, directed to the requirements of individual patients on the basis of genetic, biomarker, phenotypic, or psychosocial characteristics, discriminating a given patient from others with similar clinical states. Oncology is the foremost application area of precision medicine. Although the aim of precision medicine is very clear and simple, the clinical practice of it in cancer patients has some challenges, such as tumor heterogeneity. Tumor heterogeneity can be the most challenging part, causing that the targeting a single genetic abnormality cannot be optimum way of controlling the cancer. Multidrug resistant sub-clone formation is one of the heterogeneity concepts. The detection of multidrug resistance in leukemia cancer cells is the main aim of this thesis. Biological and electrical characterization, Annexin V/PI apoptosis assay and ion release-based impedance spectroscopy, respectively, have been carried out to determine the parameters of drug resistance detection, based on dielectrophoresis. The cell lines in our laboratory were classified as high-level laboratory and clinically relevant model of resistance. Next, the aim in the apoptosis assays was to determine correct time and doses for the screening of imatinib and doxorubicin effects on K562 and CCRF-CEM cells by electrical analysis. For CCRF-CEM/wt cells, time and dose were determined as 18h and 250 nM doxorubicin while the major death mode in K562/wt cells was not observed as apoptosis. Therefore, 24h and 300 nM imatinib were chosen for K562/wt cells’ electrical examinations. Cell cytoplasmic conductivity is one of the key parameters, changing according to physiological properties of cells. Ion release-based impedance spectroscopy was utilized for the first time in the literature to determine average total ion concentration of a cell. Results show that the total ion concentration of K562/imaR cells was 1.78 times higher than that of K562/wt ones while CCRF-CEM/doxR cells have 1.2 times higher ion concentration than wild type ones. Using the ratio between K562 cells, DEP medium conductivity and operation frequency were determined as 200 mS/m and 8.59 MHz. Values were identified as 160 mS/m and 6.15 MHz for CCRF-CEM cells. 3D and planar electrodes were implemented in LOC system integrated with the IS counting units for quantification of trapped cells by DEP. Although higher DEP force is necessary in high conductivity buffer-based DEP, 3D-obstacle electrodes acted like hydrodynamic traps due to the elasticity of leukemia cells and misinterpretation in trapping analysis occurred. Therefore, analyses were continued with planar electrode devices. Under normal growth conditions, 3 biological replicates of K562/wt and K562/imaR cells were analyzed with this LOC system at 8.59 MHz and 20 Vpp voltage immersing them into a DEP solution, having 200 mS/m conductivity. 57.6% (±6.7%) of resistant cells were trapped on electrodes while wild type ones were trapped with a ratio of 20.3% (±4.2%). This can be interpreted as the tumor heterogeneity can be determined in terms of MDR level. Drug exposed K562 cells during 24h were analyzed by the LOC system. Results show that the trapping ratio of wild type cells exposed to drug was decreased to half of control group, endorsing that DEP can achieve drug screening. In conclusion, DEP can achieve both the quantification of tumor heterogeneity in terms of drug resistance and drug screening. As a future work, cell adhesion to parylene, cell precipitation in inlet reservoir, and channel cleaning problems should be solved by optimizing channel coating procedure, using zero-dead-volume interfaces, and constructing a valve-tubing system at the inlet of LOC system, respectively.

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Citation Formats
Y. Demircan Yalçın, “A Lab-on-a-chip system integrating dielectrophoretic detection and impedance counting units for chemotherapy guidance in leukemia,” Ph.D. - Doctoral Program, Middle East Technical University, 2018.