Fabricated cantilever for AFM measurements and manipulations: pre-stress analysis of stress fibers.

The atomic force microscope (AFM) is a highly successful instrument for imaging of nanometer-sized samples and measurement of pico- to nano-Newton forces acting between atoms and molecules, especially in liquid. Generally, commercial AFM cantilevers, which have a sharp tip, are used for AFM experiments. In this review, we introduce micro-fabricated AFM cantilevers and show several applications for cell biology. In manipulation of samples on a cellular scale with a force of tens to hundreds of nano-Newtons, attempts have been made to secure the formation of covalent/non-covalent linkages between the AFM probe and the sample surface. However, present chemistry-based modification protocols of cantilevers do not produce strong enough bonds. To measure the tensile strength and other mechanical properties of actin-based thin filaments in both living and semi-intact fibroblast cells, we fabricated a probe with a hooking function by focused ion beam technology and used it to capture, pull and eventually break a chosen thin filament, which was made visible through fusion with fluorescent proteins. Furthermore, we fabricated a microscoop cantilever specifically designed for pulling a microbead attached to a cell. The microscoop cantilevers can realize high-throughput measurements of cell stiffness.