Detection of changes in cell membrane structures using the Bioimprint technique

Recent advances in medical sciences have revealed the significance of cellular structures and morphology in biological function. A cell’s membrane represents the boundary between the cells and its environment. The variations in cell surface and morphologies may be used as indications of malfunction or even diseases. If abnormalities such as cancer can be detected at the molecular level this will offer an important means for early diagnosis using small numbers of cells. The use of electron and scanning probe microscopy such as atomic force microscopy (AFM) could facilitates the opportunity to study and examine the molecular processes of living cells in greater details. The difficulties faced in direct cellular analysis when using and operating the AFM in situ for morphological studies of the cells has led to the development of a novel approach called Bioimprint (Traut and Papanicolaou, 1941 [1]). Inspired by the high resolution of nanoimprint lithography processes, Bioimprint has been applied to a new area of biological cell replication for the purpose of imaging and analysis and has revealed some very important biological events when combined with AFM imaging. For this research, the structural features of endometrial cancer cells were investigated. Regulation of selected peptides were examined, especially those which are associated with angiogenic factors that promote the proliferation of nutrient-bearing blood vessels that support tumour growth. Using the Bioimprint technique which is a soft lithography process, an impression of the cell topology was created by applying a layer of monomer mixture onto the cells attached to a substrate and rapidly curing it under UV-light. Fast UV-radiation enables the imprint to lock cellular processes within minutes and replicas of the cancer cells exhibit structures down to nanometer scale. Cancer cells were cultured and incubated in accordance with standard biological culturing procedures and protocols approved by the New Zealand Human Ethics Committee. High-resolution AFM imagery provides the opportunity to examine the structure and topography of the cells closely so that any abnormalities can be identified. To study the replicated imprints, the features that resembled secretory pores on the AFM images were noted. The numbers of pores correlated well with levels of vascular endothelial growth factor (VEGF) that were secreted by the cells. Further experiments were conducted in which anti-VEGF-coated microbeads were observed in the AFM images to be attracted to the areas of the pores.