The mechanical properties of the cell play an important role in essential cellular functions including mechanotransduction, shape stability, adhesion, motility and gene expression. Knowledge of cell mechanics is of particular interest in lung cells given that these cells are continuously subjected to large cyclic forces owing to breathing. Nanotechnology enables manipulation of matter with nanometric and picoNewton precision, providing innovative approaches for studying the structure and function of single biomolecules and cells under physiological conditions. We use atomic force microscopy (AFM) and magnetic microbeads (MM) to study the mechanical properties of lung epithelial cells.

Atomic Force Microscopy

The AFM probe the sample with a sharp tip (radius of curvature ~ 30 nm) placed at the end of a flexible cantilever. A piezoelectric actuator drives 3D displacement of the probe with nanometric precision. The force applied by the tip to the sample is measured with a laser beam, which is focused at the end of the cantilever and reflected off onto a segmented photodetector. High-resolution images under physiological conditions can by obtained by scanning the surface of the sample keeping the tip-sample force constant. Mechanical properties of single biomolecules and cells can be probed by pressing or pulling the sample with the tip and measuring the force-displacement relationship. Ligand-receptor adhesion can be probed by coating tip with ligands to cell membrane receptors.

Atomic Force Microscope (AFM)	Triangular AFM cantilever. SEM image. The arrow indicates the tip. The bar is 20 µm.
AFM image of human bronchial epithelial cells (BEAS-2B) in culture. The bar indicates ~10 µm	Alveolar epithelial cells (A549) in culture probed with a triangular AFM cantilever. Bright field image obtained with an inverted microscope. C: triangular cantilever; T: cantilever tip L: laser beam focused onto the cantilever.

Magnetic Microbeads

Magnetic microbeads enable sample manipulation by means of external magnetic fields. Ferromagnetic microbeads (diameter ~ 5 µm) coated with ligands to membrane receptors are attached to the cell surface. The cell culture is placed on an inverted microscope and the beads are magnetized in the horizontal plane with a brief (~1 ms) and large (~100 mT) magnetic pulse. Subsequently, the beads are subjected to controlled forces and torques by means of magnetic fields and gradients produced with coils. The movement induced in the beads is tracked with nanometric resolution by signal processing of their image recorded with a CCD camera. The relationship between the magnetic force applied to the beads and the resulting movement allows us to characterize the mechanical properties of the cytoskeleton and to identify pathways of force transmission through the cell membrane.

Magnetic microbeads (arrows) coated with RGD (Arg-Gly-Asp) containing peptide attached to the surface of bronchial human epithelial cells (BEAS-2B). Image obtained by scanning electronic microscopy. The bar indicates 5 µm.	Contrast phase image of bronchial epithelial cells (BEAS-2B) in culture with coated ferromagnetic microbeads (dark dots) attached to the cell membrane.

Digitized image of a microbead recorded with a CCD camera (left). Intensity distribution of the microbead image (right).

 

 

Related links

Magnetic Twisting Cytometry

http://www.nature.com/physics/highlights/6853-4.html -- Physics in biology: soft cells. Nature magazine; Physics Portal, September, 2001.

http://focus.aps.org/v8/st16.html -- A cellular glass menagerie, Geoff Brumfiel, Physical Review Focus, American Physical Society, 24 September, 2001.