Ovarian cancer is one of the most lethal gynaecological diseases. One of the reasons for its poor survival rates is its typically late diagnosis at an advanced stage of tumour progression. A better understanding of why and how cancer initiates and develops is crucial for early diagnosis and the development of improved treatment methods. To this point, investigations of mutations on genes of cells to reduce mortality of ovarian cancer have not lead to the desired outcome. Recently, increased attention has been paid to the influencing factors in cancer cell progression such as the cellular microenvironment. It is known that the natural microenvironment of tumours differs greatly from that of healthy tissues and that modification of the cell’s environment causes cellular responses such as changes in morphology, protein expressions, cell division and migration behaviour. In this work, an experimental set-up was developed to trap cells in defined 3D wells to investigate how physical properties of the microenvironment influence ovarian can- cer cells. Polyacrlyamide gels were simultaneously structured and protein patterned to create a platform for cell experiments, which allows the tuning of individual physical properties of the microenvironment of cells (such as stiffness, available volume and protein compositions for cell attachment) independently of each other. The designed and optimised fabrication process begins with optical lithography to transfer a pattern onto a Silicon (Si) substrate and a dry etching step to obtain an array of pillars while transforming the Si-substrate into a Si-mould. The Si-mould serves as a stamp during a µ-contact printing approach to transfer defined patterns of protein, and simultaneously as a mould during polyacrylamide polymerisation. The resulting microstructured and protein patterned polyacrylamide gels can then be used as cell culture substrates for cell experiments.

Si-moulds and polyacrylamide gels were characterised with scanning electron microscopy, atomic force microscopy and confocal laser scanning microscopy. By investigating four differently sized circular patterns (with diameters of 20 µm, 30 µm, 40µm and 60 µm) and four different stiffnesses of the polyacrylamide gels (1 kPa, 8 kPa, 30 kPa and 100 kPa), it was shown that the fabrication process is robust and easy to adjust.

Computational analysis protocols were developed and established for traction force microscopy and brightness fluctuation analysis of cells, including the corresponding bright- ness autocorrelation functions. It was shown that microstructured and protein patterned polyacrylamide gels can be used for investigations of protein expressions, cellular traction forces and brightness fluctuations of cells. While the analyses of protein expressions and autocorrelation functions of the brightness fluctuations need further improvement, the results of the traction force experiments allow a first hypothesis to be formulated: cellular traction forces increase with decreasing volume available to the cell, as indicated by analysis of cell experiments on ovarian cancer cells of the cell line SKOV3.

The developed microstructured and protein patterned polyacrylamide gels are an important step to gain a better understanding on how mechanical properties of the microenvironment influence cellular responses. This experimental setup can be easily adapted and optimised for further investigations of cancer cells and can thus help in the development of new treatment approaches.