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• W3015862397 abstract "The number of human cell types used to be estimated as approximately 300, but recent research suggests that this is closer to 10,000 because it is becoming evident that diverse cell populations exhibit a significant extent of functional heterogeneity. Characterisation of cell heterogeneity requires monitoring of a large number of single cells. A commonly used method in this respect is flow cytometry, which has a very high throughput of millions of cells but in its conventional implementation it is not capable of subcellular imaging. More detailed characterisation of cell populations requires the development of platforms capable of high-resolution microscopy of large numbers of single cells. The work in this thesis concerns the application of optically accessible microfluidic cell arrays to a number of cell populations, with the aim to assess the potential of this method for the characterisation and quantification of cell heterogeneity in general, and nanoparticle drug delivery for regenerative medicine applications in particular. Microfluidic devices with ~500-2,000 cell array sites were fabricated by soft lithography. Array chambers consist of a serpentine microfluidic channel that is lined by trap pockets, where cells are immobilized by a hydrodynamic cross-flow, presenting a well-defined array for optical imaging with oil-immersion objectives. The fabricated device was able to trap up to 20,000 cells and enumerate large populations by fluorescence tags while still getting subcellular details in single cells. The interface with the syringe pump and the fluorescence microscope was optimised. The microfluidic chip was shown to be able to array fixed freshly isolated bone marrow mononuclear cells, and the sub-population of skeletal stem cells could be identified by plasma membrane labelling with two fluorescent antibodies anti-Stro-1 and anti-GPA. A post-processing workflow for image analysis was developed to analyse single cells and obtain properties (e.g. size, fluorescence intensity, circularity) and identify the array position of cells of interest. Comparison with flow cytometry suggested enrichment of large sub-populations. Polymeric nanoparticles were prepared with different fluorescent stains as cargo, and nanoparticle-facilitated dye delivery was subsequently demonstrated for arrayed osteosarcoma MG63 cells, with subcellular imaging indicating lysosomal uptake. Nuclear cell signalling in bone marrow stem cells was also characterised. Overall, this work provides detailed methodologies and procedures to develop customisable microfluidic cell arrays for single-cell imaging which are comparable with more complex analytical equipment such as flow cytometry and imaging flow cytometry. Additionally, the project indicated that microfluidic cell arrays are suitable for characterisation of heterogeneous cell populations by image analysis, and subcellular 3D imaging in non-adherent cells." @default.
• W3015862397 created "2020-04-17" @default.
• W3015862397 creator A5018267127 @default.
• W3015862397 date "2019-10-01" @default.
• W3015862397 modified "2023-09-25" @default.
• W3015862397 title "Microfluidic stem cell arrays to image heterogeneous cell populations and intracellular drug delivery" @default.
• W3015862397 hasPublicationYear "2019" @default.
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