Precise cell manipulations and imaging of cellular responses : Methods developed using microfluidic, 3D-printing and microfabrication technologies

It is at the heart of biological and medical research to try and understand how cells communicate with each other, and how cells respond to alterations in their environment, including treatment with different drugs. There is in this context a continued need for better methods that allow researchers to precisely manipulate cells and their microenvironment and to study the resulting responses using high-resolution live microscopy. This thesis presents the development and implementation of several devices that addresses these needs. A novel microfluidic device called the cell assembly generator (CAGE) was created to generate precisely composed cell clusters of different cell types; the first of its kind. Experimental evidence demonstrated that the CAGE chip can be used to study paracrine signalling in tailor-made cancer cell clusters composed of up to five cells. A high-throughput microfluidic chip for rapid phenotypic antibiotic susceptibility testing was developed and tested using 21 clinical isolates of Klebsiella pneumoniae, Staphylococcus aureus and Escherichia coli against a panel of antibiotics. Stable minimum inhibitory concentration values were obtained from this system within 2-4 hours with high accuracy to the standard method. 3D-printing was used to create a modular and affordable time-lapse imaging and incubation system, called ATLIS. This system enables researchers to convert simple inverted microscopes into live cell imaging systems, where images and movies of living cells can be recorded using a regular smartphone. Finally, a strategy was developed for the generation of modular microfluidic systems using 3D-printed moulds for PDMS casting, to enable studies of leukocyte adherence to differentially treated endothelial cell populations in the same field of view and under the same conditions