Microfluidic Device Development for Analyzing Single Glioblastoma Cells

Glioblastoma remains a deadly disease due to the diffuse infiltration of single tumorcells into the surrounding tissue. Even with the current treatment regimens of surgery,radiation, and chemotherapy, the median survival time is approximately one year. Likemany solid tumors, glioblastoma is extremely heterogeneous with respect to multiplephenotypes such as invasive capacity, therapeutic resistance, and tumorigenicity. Thisheterogeneity complicates our understanding of glioblastoma and consequently, ourability to treat this disease. Unfortunately, standard population-based assays can maskthe properties of rare subpopulations within a tumor and therefore, obscure ourunderstanding of these subpopulations. As a result, without tools that allow for singlecell analysis, we are unable to interrogate how different subpopulation phenotypes mayserve as indicators of glioblastoma tumor growth and progression.In this dissertation, we sought to develop and optimize new microfluidic tools to analyzesingle glioblastoma cells for a range of phenotypes: viscoelasticity, motility, andinvasion. First, we developed a cross-slot based platform and a correspondinganalytical model that enables the determination of cellular viscoelastic properties(stiffness and fluidity) in a high-throughput manner. Using this platform, we quantifiedthe viscoelastic properties of 3T3 fibroblasts and glioblastoma tumor initiating cells(TICs) and observed the expected changes in the cellular elastic modulus in responseto agents that soften or stiffen the cytoskeleton. Second, we developed a microfluidicdevice and workflows that integrates measurements of invasive motility and targetedprotein expression with single cell resolution, which we named SCAMPR (Single CellAnalysis of Motility and Proteotype). Using this platform, we identified two proteins,Nestin and EphA2, which positively correlates with TIC invasive motility.In summary, this dissertation focuses on the development of microfluidic platforms forsingle cell analysis. Our developed platforms provide a method in which to interrogatesingle cells in a high throughput manner and to identify novel relationships betweenvaried cellular phenotypes. These insights are crucial for the identification of both raresubpopulations within a given tumor and patient-specific protein markers that describethese subpopulations