http://experimentationlab.berkeley.edu/sites/default/files/AFMImages/butt_AFM.pdf

Force measurements with the atomic force microscope: Technique, interpretation and applications

Solid lipid nanoparticles: Production, characterization and applications

Single-molecule force spectroscopy has emerged as a powerful tool to investigate the forces and motions associated with biological molecules and enzymatic activity. The most common force spectroscopy techniques are optical tweezers, magnetic tweezers and atomic force microscopy. Here we describe these techniques and illustrate them with examples highlighting current capabilities and limitations.

Single-molecule force spectroscopy: optical tweezers, magnetic tweezers and atomic force microscopy

The adhesion force between the tip of an atomic force microscope cantilever derivatized with avidin and agarose beads functionalized with biotin, desthiobiotin, or iminobiotin was measured Under conditions that allowed only a limited number of molecular pairs to interact, the force required to separate tip and bead was found to be quantized in integer multiples of 160 +/- 20 piconewtons for biotin and 85 +/- 15 piconewtons for iminobiotin The measured force quanta are interpreted as the unbinding forces of individual molecular pairs

https://science.sciencemag.org/content/sci/264/5157/415.full.pdf

Adhesion forces between individual ligand-receptor pairs.

Atomic force microscopy (AFM) has been used to measure the strength of bonds between biological receptor molecules and their ligands. But for weak noncovalent bonds, a dynamic spectrum of bond strengths is predicted as the loading rate is altered, with the measured strength being governed by the prominent barriers traversed in the energy landscape along the force-driven bond-dissociation pathway. In other words, the pioneering early AFM measurements represent only a single point in a continuous spectrum of bond strengths, because theory predicts that these will depend on the rate at which the load is applied. Here we report the strength spectra for the bonds between streptavidin (or avidin) and biotins-the prototype of receptor-ligand interactions used in earlier AFM studies, and which have been modelled by molecular dynamics. We have probed bond formation over six orders of magnitude in loading rate, and find that the bond survival time diminished from about 1 min to 0.001 s with increasing loading rate over this range. The bond strength, meanwhile, increased from about 5 pN to 170 pN. Thus, although they are among the strongest noncovalent linkages in biology (affinity of 10(13) to 10(15) M(-1)), these bonds in fact appear strong or weak depending on how fast they are loaded. We are also able to relate the activation barriers derived from our strength spectra to the shape of the energy landscape derived from simulations of the biotin-avidin complex.

Energy landscapes of receptor–ligand bonds explored with dynamic force spectroscopy

The atomic force microscope (AFM) can be used to image the surface of both conductors and nonconductors even if they are covered with water or aqueous solutions. An AFM was used that combines microfabricated cantilevers with a previously described optical lever system to monitor deflection. Images of mica demonstrate that atomic resolution is possible on rigid materials, thus opening the possibility of atomic-scale corrosion experiments on nonconductors. Images of polyalanine, an amino acid polymer, show the potential of the AFM for revealing the structure of molecules important in biology and medicine. Finally, a series of ten images of the polymerization of fibrin, the basic component of blood clots, illustrate the potential of the AFM for revealing subtle details of biological processes as they occur in real time.

https://science.sciencemag.org/content/sci/243/4898/1586.full.pdf

Imaging crystals, polymers, and processes in water with the atomic force microscope.

A scanning ion-conductance microscope (SICM) has been developed that can image the topography of nonconducting surfaces that are covered with electrolytes. The probe of the SICM is an electrolyte-filled micropipette. The flow of ions through the opening of the pipette is blocked at short distances between the probe and the surface, thus, limiting the ion conductance. A feedback mechanism can be used to maintain a given conductance and in turn determine the distance to the surface. The SICM can also sample and image the local ion currents above the surfaces. To illustrate its potential for imaging ion currents through channels in membranes, a topographic image of a membrane filter with 0.80-micrometer pores and an image of the ion currents flowing through such pores are presented.

https://science.sciencemag.org/content/sci/243/4891/641.full.pdf

The scanning ion-conductance microscope.

Using a new mode of scanning, the force modulation mode, surfaces are imaged by the atomic force microscope. The new contrast mechanism relies on variation in the surface elasticity. The cross section of a carbon fibre and epoxy composite is imaged, showing contrast between the two materials. Surface elasticity variations across the cross section of the fibre are revealed. A lateral modulation mode is used to highlight atomic steps in gold.

https://iopscience.iop.org/article/10.1088/0957-4484/2/2/004/pdf

Using force modulation to image surface elasticities with the atomic force microscope

Imaging cells with the atomic force microscope

An atomic force microscope (AFM) operating with the tip in contact with the surface has given molecular-resolution images of monomolecular Langmuir-Blodgett (LB) films consisting of the lipids DMPE (DL-{alpha}-dimyristoylphosphatidylethanolamine), DMPG (L-{alpha}-dimyristoylphosphatidylglycerol), DODAB (dioctadecyldimethylammonium bromide), or 1:1 DODAB/DPPG (L-{alpha}-dipalmitoylphosphatidylglycerol). All the LB films were submerged in a buffer solution during imaging. Precoated mica served as a substrate for the LB films, which were transferred onto it by vertical dipping. The molecular arrangement of the lipids in the LB films agrees with the known 3D crystal data to within the accuracy of our measurements, about 20%, and shows a clear difference from the substrate mica, whose hexagonal lattice could be seen by increasing the force applied by the AFM if the bottom monomolecular film was not covalently interlinked. The AFM also gave images of single-stranded DNA bound to LB films, thus illustrating the potential of LB films as substrates for binding and imaging macromolecules.

Scot A. C. Gould

Papers

Imaging crystals, polymers, and processes in water with the atomic force microscope.

The scanning ion-conductance microscope.

Using force modulation to image surface elasticities with the atomic force microscope

Imaging cells with the atomic force microscope

Molecular-resolution images of Langmuir-Blodgett films and DNA by atomic force microscopy