Abstract
The cell cycle is the process of tightly integrated events leading to cell growth, DNA replication and cell division. Cell cycle dysregulation is the common reason for abnormal cell growth (cancer) and therefore, scientists are interested in determining the proportion of cells in each cell cycle phase for effective targeting of cancerous cells at a specific phase. Knowledge of cells’ electrophysiological properties can provide new insights in cell cycle events studies. Dielectrophoresis (DEP) is a technique that can be used to obtain these properties for cell characterisation. DEP is a fast, label-free, high-throughput, and cost-effective micro-engineered technology that reduces the sample volume and provides automated real-time analysis. This thesis explores the application of the DEP-microwell electrode system (both the serial design and the parallel integrated system) in cell cycle analysis. It also investigates the variation of cellular electrophysiological properties (particularly the effective membrane capacitance (ceff), the effective membrane conductance (geff) and the cytoplasmic conductivity (σ_CP)) during different phases through the cell cycle, in vitro. In addition, this work assesses the relation between the intracellular DNA percentage in each phase and the acquired DEP data to find out the possible correlation between cell cycle phases and cellular electrophysiological properties. For the purpose of having the cells at different stages of the cycle, a K562 cell line was synchronised at G0/G1, late G1/S and early S-phase, using serum starvation (SS), hydroxyurea (HU) and aphidicolin (APH). DEP measurements were carried out for cells before treatment (control cells), immediately after treatment (0h-study) and at 2, 4 and 24 hours after releasing the cells back to the culture medium for each synchronisation method. Due to improved time resolution and frequency range in the parallel design of DEP-microwell electrode, acquired results of this method showed more accurate data. The obtained results demonstrated significant difference of electrophysiological properties through different phases of K562 cell cycle. Using the parallel design: σ_CP at 0h-study: σ_CP (SS) greater than σ_CP (HU) and σ_CP (APH) by ~50% and ~60%, respectively geff at 0h-study: geff (HU) greater than geff (APH) and geff (SS) by ~90% and ~65%, respectively ceff at 0h-study: ceff (HU) ≈ ceff (APH), both greater than ceff (SS) by ~22% In summary, results showed that a rising pattern for cell radii, a downward followed by an upward trend in σ_CP values, a falling followed by a rising pattern for ceff value, and a rising pattern for geff values were observed during the cell cycle. Furthermore, the change of these electrophysiological properties correlated with the proposed trends at different phases of the cell cycle. This suggests that the DEP-microwell electrode system can be used as an innovative, real-time, portable, cost-effective and high-throughput approach for cell characterisation at particular stages of the cell cycle.