Dissertation

AFM Probes—Cell Imaging and Membrane Mechanics

Vaidyanathan, Priya

2004

Overview

The study of cellular response to dynamic mechanical loading is an important component of biological research into cellular function at the single-cell level. Understanding the range of cellular changes that can occur as a result of applied forces might be useful in applications such as cell repair and tissue generation. Micromanipulation techniques have been used to study the adaptation of cells to their environment in the past experiments However, accurate imaging and force response measurements as a function of position on the cell makes the use of force probes with nanometer scale dimensions very desirable, since they provide dramatically better spatial resolution and can detect very small changes in forces occurring dynamically in a cell.The main objective of this thesis is to establish protocols to study and analyze the mechanical response of single cells under mechanical loading. In order to accomplish this goal, Atomic Force Microscopy was used because this technique provides the ability to image cells in their native physiological environment, and to apply and measure response to forces in the nano-newton range at selected positions. Atomic Force Microscopy allows comparisons of the mechanical response in different regions of a cell and the observation of possible morphological changes before and after loading.Two different cell lines were studied. Mouse hybridoma and bovine aortic endothelial (BAE) cells were chosen because they have similar dimensions but different biological characteristics. The study of different cell lines also provides insight into the general response of cells to mechanical loading and the difference in responses among individual cells and cell lines. Force response measurements were carried out in a sequence of loading/unloading cycles. For each cycle, the AFM probe was moved 1 μm both towards and away from the cell. The next loading/unloading cycle would then begin after the probe had been moved to a position 0.5 μm closer to the substrate. This methodology was designed to observe changes in the force response over a small range of cell deformation. Figure a shows a typical sequence of force response curve(s) that proceed from right to left. Figure b isolates on one load-unload cycle that clearly exhibits hysteretic behavior related to the viscoelastic nature of the cell. In this case, some elastic energy stored during cell loading is being dissipated. To investigate these effects, experiments were performed to study the impact of variations in cantilever spring constants and AFM tip approach velocities. A mechanical model representing the cell / cantilever system as a one dimensional, non-linear oscillator with viscoelastic terms was developed to compare with the experimental results and good qualitative agreement was found.

• Publisher: ProQuest Dissertations & Theses
• Subject: Biomedical engineering / Biophysics / Cellular biology
• ISBN: 9798380409131