AFM-TIRF combination

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Atomic force microscopy allows the measurement of cellular forces on both molecular and cellular scales. Such measurements are essential for quantification and an in-depth understanding of the interaction of cells with surfaces. A current shortcoming of force microscopy data is that information on the contact area of the cells during the measurement is missing, which affects the analysis of the data. For example, higher cell-surface interaction forces can be caused by both stronger adhesion and larger cell-surface contact area. Therefore, a force measurement system based on an atomic force microscope (AFM) is used here, which is set up in combination with a fluorescence TIRF (Total Internal Reflection Fluorescence Microscopy) device. TIRF will optically detect the contact area of the cells, which will then be correlated with AFM force curves. This correlation of TIRF fluorescence data and AFM data should allow much more accurate data interpretation than with conventional systems. Since living cells are to be studied, special requirements are placed on the atomic force microscope (e.g., heatable liquid cell). To ensure the attachment of cells to the measurement system, a microfluidic cell aspiration device will be used. The combination of an AFM with a TIRF microscope applied for here is unique on campus and will allow completely novel conclusions on the forces acting during cell adhesion. Because of this specialized equipment, the requested atomic force microscope-TIRF system will be used primarily to characterize cell-surface interactions. Since the instrument is to be used by several research groups, an easy-to-use instrument that can also be used stably in liquid at different temperatures must be procured. Furthermore, the laser used in the AFM must not overlap with the excitation or emission wavelengths of the fluorescent dyes used in cells, so an infrared laser is necessary. An extremely critical instrument parameter is the range of the piezo in the z-direction (vertical to the surface). Since cells often reach diameters of 50-100 µm in the adhered state and deform significantly during the detachment process due to their viscoelasticity, a piezo with a minimum range of 100 µm is envisaged.