Please use this identifier to cite or link to this item: http://scholarbank.nus.edu.sg/handle/10635/18462
Title: Development of A Miniaturization Assay Platform and its Application to Study Scarce Biological Samples
Authors: LEE YONG YEOW
Keywords: miniaturization, high-content screening, microfluidics, cancer stem cells,
Issue Date: 31-Mar-2010
Source: LEE YONG YEOW (2010-03-31). Development of A Miniaturization Assay Platform and its Application to Study Scarce Biological Samples. ScholarBank@NUS Repository.
Abstract: Miniaturization technologies have developed rapidly over the past decade. However, the challenge in advancing miniaturization strategies largely depends on their scalability to cater to a myriad of important applications. There is an increasing demand for the accurate processing of scarce samples, such as stem cells, cancer stem cells and patients? samples. Miniaturization technologies may offer important insights into the characterization of these biologically relevant samples for research and clinical applications. This thesis presents a novel miniaturized assay technology, DropArray, for conducting heterogeneous cell-based assays. The DropArray plate consists of an array of 2-mm hydrophilic spots, insulated from each other by a hydrophobic polytetrafluoroethylene (PTFE) coating. Each spot represents a 2-ul assay. DropArray Accelerator has been designed to reproducibly automate the precise movements and fluidics during parallel rinsing so that there is negligible cross-contamination between the assay points on each plate. DropArray has been successfully employed to miniaturize a wide range of heterogeneous assays, such as enzyme-linked immunosorbent assay (ELISA) and high-content screening (HCS) cell-based assays. Besides ensuring the robustness of this technology for HCS assays, effects of miniaturization have been studied in detail. HCS assays are shown to remain robust at 2 ul, using only 500 cells per data point. Applying DropArray to HCS assays also reduces the antibody staining time significantly by ~ 60%. DropArray has also been applied to study the drug responses of various scarce cancer side population (SP) phenotypes. Interesting drug resistance phenomena that have been difficult to demonstrate have been successfully elucidated in this work. The SP phenotypes enriched from various cell lines are associated with cancer stem cell properties in the literature. Besides showing increased expressions of genes associated with drug efflux capabilities, these cells have been found to initiate an entire tumor with only 3000 cells. In vitro drug response assays with these scarce cells have been conducted effectively with the DropArray platform.
URI: http://scholarbank.nus.edu.sg/handle/10635/18462
Appears in Collections:Ph.D Theses (Open)

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