We describe the fluidFM, an atomic force microscope (AFM) based on hollow cantilevers for local liquid dispensing and stimulation of single living cells under physiological conditions. A nanofluidic channel in the cantilever allows soluble molecules to be dispensed through a submicrometer aperture in the AFM tip. The sensitive AFM force feedback allows controlled approach of the tip to a sample for extremely local modification of surfaces in liquid environments. It also allows reliable discrimination between gentle contact with a cell membrane or its perforation. Using these two procedures, dyes have been introduced into individual living cells and even selected subcellular structures of these cells. The universality and versatility of the fluidFM will stimulate original experiments at the submicrometer scale not only in biology but also in physics, chemistry, and material science.

In this letter, we describe the on-demand dispensing of single liquid droplets with volumes down to a few attoliters and submicrometric spacing. This dispensing is achieved using a standard atomic force microscope probe, with a 200 nm aperture at the tip apex, opened by focused ion beam milling. The inside of the tip is used as reservoir for the liquid. This maskless dispensing, realized in ambient environment, permits the direct creation of droplet arrays. Nanoparticles, suspended in the liquid, were organized on a surface.

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Atomic force microscopy (AFM) investigations of living cells provide new information in both biology and medicine. However, slow cell dynamics and the need for statistically significant sample sizes mean that data collection can be an extremely lengthy process. We address this problem by parallelizing AFM experiments using a two-dimensional cantilever array, instead of a single cantilever. We have developed an instrument able to operate a two-dimensional cantilever array, to perform topographical and mechanical investigations in both air and liquid. Deflection readout for all cantilevers of the probe array is performed in parallel and online by interferometry. Probe arrays were microfabricated in silicon nitride. Proof-of-concept has been demonstrated by analyzing the topography of hard surfaces and fixed cells in parallel, and by performing parallel force spectroscopy on living cells. These results open new research opportunities in cell biology by measuring the adhesion and elastic properties of a large number of cells. Both properties are essential parameters for research in metastatic cancer development.

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Various 1D and 2D scanning probe arrays with integrated piezoresistive sensors have been developed in view of nanobiology and life science applications. For such applications, the experiments need usually to be performed under liquid conditions. The probe arrays have been fabricated by using silicon micromachining techniques and SOI wafers. For biological applications, when the cantilevers are immersed in electrically conductive buffer environments, the piezoresistive sensing elements have to be protected. For this purpose, the sensors were coated with a 50 nm thick silicon nitride film. The realized piezoresistive cantilevers had low spring constants between 0.1 and 0.5 N/m, well suited for biological applications. Parallel imaging using a 1 · 2 probe array was demonstrated by simultaneous imaging of a test sample in buffer solution.

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Antibody array-based technology is a powerful emerging tool in proteomics, but to enable global proteome analysis, antibody array layouts with even higher density has to be developed. To this end, we have further developed the first generation of a nanoarray platform, based on attoliter-sized vials, attovials, which we have characterized and used for the detection of complement factor C1q in human serum samples. Finally, we demonstrated proof-of-concept for individual functionalization of the attovials with a recombinant antibody.