Collections > Electronic Theses and Dissertations > Capturing, Analyzing and Collecting Adherent Cells Using Microarray Technologies

Effective separation of a particular cell of interest from a heterogeneous cell population is crucial to many areas of biomedical research including microscopy, clinical diagnostics and stem cell studies. Examples of such studies include the analysis of single cells, isolation of transfected cells and cell transformation studies. Biological technologies can have skewed results if cells outside of the type of interest are present. Additionally, in many instances the targeted cells are of low abundance with respect to the heterogeneous population. For these reasons, it is important to have a technique capable of identifying the desired cells, separating these cells from unwanted cells and collecting the marked cells for further analysis. Two biotools, referred to as micropallets and microrafts, have recently been introduced for sorting adherent cells. These devices comprise arrays of microelements weakly attached to a substrate. Following culture of adherent cells on the elements, individual microstructures are selectively detached from the array while still carrying the cells. These technologies have shown success in sorting single cells from small heterogeneous cell populations with high post sorting viabilities. However, previous device designs employed gravity-based collection methods and small microelement arrays which substantially reduced the collection yields, purities and sample sizes. In this dissertation new approaches are described for capturing, examining and isolating individual cells by micropallet and microraft technologies. Initially a new approach was developed to isolate released microstructures from the array employing magnetism. Microstructures were embedded with uniformly dispersed magnetic nanoparticles which allowed collection by an external magnet immediately following release. Application of a magnetic field permitted microstructure collection with high yield, precision and purity. This improved collection efficiency enabled isolation of very rare cell types. Large arrays constituting over 106 micropallets were developed along with imaging analysis software to identify and sort low abundance target cells. This system was employed to isolate breast cancer stem cells from a heterogeneous cell population and circulating tumor cells directly from peripheral blood. Additionally, an array-based cell colony replication strategy was established which allowed highly efficient colony splitting and sampling.