The development of a spheroid-based migration assay on tissue-specific extracellular matrix

Cell dissemination during tumour development is a characteristic of cancer metastasis. Dissemination from three-dimensional spheroid models on extracellular matrices designed to mimic tissue-specific physiological microenvironments may allow us to better elucidate the mechanism behind cancer metastasis and the response to therapeutic agents. The orientation of fibrillar collagen plays a key role in cellular processes and mediates metastasis through contact-guidance. Understanding how cells migrate on aligned collagen fibrils requires in vitro assays with reproducible and standardised orientation of collagen fibrils on the macro-to-nanoscale. Herein, we implement a spheroid-based migration assay, integrated with a fibrillar type I collagen matrix, in a manner compatible with high throughput image acquisition and quantitative analysis. The migration of highly proliferating U2OS osteosarcoma cell spheroids onto an aligned fibrillar type I collagen matrix were quantified. The assay developed here can be applied to a fully automated imaging and analysis pipeline for the assessment of tumor cell migration with high throughput screening. To develop the three-dimensional (3D) migration assay, various biomimetic and aligned fibrillar collagen scaffolds were characterised in Chapter 4 using Atomic force microscopy revealing interconnected, aligned and crimped collagen fibrils. In addition, AFM indentations on aligned fibrillar collagen scaffold in phosphate buffer saline (PBS) showed a mean Young’s modulus of 31 ± 12 kPa using the Hertz model. Furthermore, in chapter 5, methods for reproducible spheroid transfer and positioning control in the well plate are explored using additive manufacturing for quick prototyping of a spheroid transfer adapter. Finally, a complete spheroid-based migration assay on fibrillar collagen that is amenable to high-throughput (HT) image acquisition and analysis is demonstrated in chapter 6.