We have developed the atomic force microsocpy (AFM) to measure the complex shear modulus, G*, of a large number of cells. In the AFM technique, live cells were arranged in a micro-fabricated glass substrate under the physiological conditions, and the AFM force measurement was examined in many different cells automatically. The results shown in the previous studies revealed that the frequency-dependent G* was well fitted to the so-called structural damping model, which consists of a single power-law function with a Newtonian viscous effect. However, the detail relationship has not been understood. The aim of this study was to verify the relationship between the storage and loss moduli. As results, we found that the relation between the hysteresivity (the ratio of the storage and loss moduli) and the power-law exponent was in good agreement with the structural damping model, and the result was the same as that observed in magnetic twisting cytometry (MTC), in which cells were cultured on flat substrates. This result indicated that the AFM technique presented here becomes a useful technique for precisely measuring the statistical behavior of single cell rheology.

We present a novel experimental platform based on a combined Atomic Force Microscopy (AFM) and Micro-Electrode Array (MEA) set-up. We have used it to measure minimal changes in the morphological/mechanical properties of electrically active cell cultures as well as to measure the changes in the extracellular electrical activity when a single cell is stimulated by means of the AFM tip. In particular, we studied the dynamical changes in cell elasticity of embryonic rat cardiac myocytes along the contraction-relaxation cycle. Applying high load indentations, we also recorded the effects of mechanical stimulations on the cell electrophysiology. The dynamic elastic modulus of the cell related to the contraction-relaxation cycle reveals a temporal behavior that closely follows the changes in cell height. Observed values of dynamic elastic modulus at a maximum indentation depth of 1500nm varied between 8.93 ± 0.78 kPa during systolic (contraction) phase and 4.26 ± 0.47 kPa during diastolic (relaxation) phase. Induced electrophysiological responses were observed when applying loads in the range 40-150 nN. The probability P of recording an induced electrical response (P = 0.16 for a maximum load of 100nN) increased with the maximum applied load. Pulling-like stimulations due to the tip-cell adhesion could also evocate electrical responses.

Though atomic force microscopy (AFM) interrogates biological materials through mechanical interactions, achieving quantitative mechanical information such as modulus and adhesion at high resolution has been a challenging task. A technology for nanometer scale mechanical property mapping, peak force tapping (PFT), was developed to achieve high resolution imaging and quantitative mechanical measurements simultaneously. PFT controls instantaneous interaction force and record force spectroscopy at each pixel to calculate mechanical properties. A feedback loop maintains a constant peak force, a local maximum point in the force spectroscopy, at the level of Pico Newtons throughout the imaging process. Such high precision force controls enable application of ultra-sharp probe to image biological samples in vitro and achieve molecular resolution in protein membranes. More importantly a full suite of mechanical properties, modulus, adhesion, energy dissipation and deformation are mapped concurrent with topographic imaging. To calculate nanomechanical properties reliably cantilever spring constant and tip shape were calibrated systematically. A method to accurately determine cantilever spring constant, capable of wafer scale cantilever calibration, was developed and tested against traceable force methods. With the knowledge of tip shape, derived from morphological dilation method using a reference sample, mechanical properties measured at the nanometer scale was compared with bench mark materials ranging from 0.7 MPa to 70 GPa. The same method was also applied to OmpG membranes, Lambda DNA strings, as well as live cells. The limitation of the measurement accuracy in biology samples will be discussed.