A Review of Scanning Tunneling Microscope and Atomic Force Microscope Imaging of Large Biological Structures: Problems and Prospects

The application of the scanning tunneling microscope (STM) and the atomic force microscope (AFM) to the study of small biological molecules, such as DNA and smaller molecules, has received considerable attention in the literature. This paper reviews STM and AFM studies of larger biological structures such as bacterial membranes, bacteriophages, viruses, antibodies, etc. The problems encountered in these applications are emphasized, with particular reference to the unknown conduction mechanism, tip-sample interaction forces, and tip-sample convolution artifacts in the images. The latter problem is illustrated by new results from IgG antibody complexes attached to a bacterial sheath layer. A new conduction mechanism involving a graphite film overlayer is suggested. The future prospects are discussed, with emphasis on the unique capabilities of these microscopes compared to conventional electron microscopes

A New Atomic Force Microscopy Technique for the Measurement of the Elastic Properties of Biological Materials

We developed a new technique to measure elastic properties by using the atomic force microscope (AFM) tip to press samples into grooves etched in a GaAs substrate. We measured the Young\u27s modulus of β-chitin fibres with cross-sections less than 40 nm X 20 nm to be 1-2 X 1011 N/m2. In the isotropic approximation, the Young\u27s modulus of the S-layer sheath of the archaeobacterium Methanospirillum hungatei was 1-3 X 1010 N/m2. By testing the sheath to breaking strength we estimated the bacterium can sustain an internal pressure as high as 100-200 atmospheres (1-2 x 107 N/m2)

Direct Observation by Scanning Tunneling Microscopy of the Two-Dimensional Lattice Structure of the S-Layer Sheath of the Archaeobacterium Methanospirillum hungatei GP1

Observation of the two-dimensional (2-D) 3 nm x 3 nm lattice structure of the S-layer sheath of the archaeobacterium Methanospirillum hungatei is reported for the first time by scanning tunneling microscopy. The samples consisted of sheath fragments deposited on a TaSe2 substrate and coated with a Pt/Ir film. In addition to confirming the 2-D structure, the images reveal some new information about the nano-scale details of the sheath structure. A lateral resolution of 1 nm was achieved, suggesting that the grain size of the Pt/Ir films was much less than for similar films deposited on a smooth metal surface

Gluten Induces Subtle Histological Changes in Duodenal Mu-cosa of Patients with Non-Coeliac Gluten Sensitivity: A Multi-center Study

Histological changes induced by gluten in the duodenal mucosa of patients with non-coeliac gluten sensitivity (NCGS) are poorly defined. Objectives: To evaluate the structural and inflammatory features of NCGS compared to controls and coeliac disease (CeD) with milder enteropathy (Marsh I-II). Methods: Well-oriented biopsies of 262 control cases with normal gastroscopy and histologic findings, 261 CeD, and 175 NCGS biopsies from 9 contributing countries were examined. Villus height (VH, in μm), crypt depth (CrD, in μm), villus-to-crypt ratios (VCR), IELs (intraepithelial lymphocytes/100 enterocytes), and other relevant histological, serologic, and demographic parameters were quantified. Results: The median VH in NCGS was significantly shorter (600, IQR: 400−705) than controls (900, IQR: 667−1112) (p < 0.001). NCGS patients with Marsh I-II had similar VH and VCR to CeD [465 µm (IQR: 390−620) vs. 427 µm (IQR: 348−569, p = 0·176)]. The VCR in NCGS with Marsh 0 was lower than controls (p < 0.001). The median IEL in NCGS with Marsh 0 was higher than controls (23.0 vs. 13.7, p < 0.001). To distinguish Marsh 0 NCGS from controls, an IEL cut-off of 14 showed 79% sensitivity and 55% specificity. IEL densities in Marsh I-II NCGS and CeD groups were similar. Conclusion: NCGS duodenal mucosa exhibits distinctive changes consistent with an intestinal response to luminal antigens, even at the Marsh 0 stage of villus architecture

Swimming speed of three species of Alexandrium as determined by digital in-line holography.

Digital in-line holographic (DIH) microscopy was used to track motility in several related species of the marine dinoflagellate Alexandrium in response to temperature after acclimation at selected temperatures. Numerical reconstruction of DIH holograms yielded high-contrast three-dimensional images of the trajectories of many motile cells swimming simultaneously throughout the sample volume. Swimming speed and trajectory were determined for clonal isolates of A. ostenfeldii, A. minutum and A. tamarense within the temperature range from 8 to 24\ub0C. The strains of these species revealed differences in temperature optima for growth and tolerance that were a function of both acclimation responses and genetic factors reflecting the origin of the isolates. The fastest swimming speeds were recorded at 24\ub0C for cells of A. minutum. Acclimated strains of all three species swam significantly slower at lower temperatures, although fastest swimming speeds did not always occur at temperature optima for growth. Aged cells from stationary phase cultures swam more slowly than cells in exponential growth phase. Doublets from a rapidly dividing culture swam faster than singlets from the same culture, confirming the propulsive advantage of paired cells. Holographic microscopy is a powerful tool for the acquisition of detailed observations of swimming behaviour of microalgal cells in the form of three-dimensional trajectories over the appropriate temporal (sub-second) and spatial (micrometer) scales.Peer reviewed: NoNRC publication: Ye

Theory of electrostatic effects in soft biological interfaces using atomic force microscopy.

We calculated the electrostatic force between a planar interface, such as a planar-supported lipid bilayer membrane, and the tip of a stylus on which another lipid bilayer or some other biomacromolecular system might be deposited. We considered styli with rounded tips as well as conical tips. To take into account the effect of dynamical hydrogen-bonded structures in the aqueous phase, we used a theory of nonlocal electrostatics. We used the Derjaguin approximation and identified the systems for which its use is valid. We pointed out where our approach differs from previous calculations and to what extent the latter are inadequate. We found that 1) the nonlocal interactions have significant effects over distances of 10-15 A from the polar zone and that, at the surface of this zone, the effect on the calculated force can be some orders of magnitude; 2) the lipid dipoles and charges are located a distance L from the hydrophobic layer in the aqueous medium and this can have consequences that may not be appreciated if it is ignored; 3) dipoles, located in the aqueous region, can give rise to forces even though the polar layer is unchanged, and if this is ignored the interpretation of force data can be erroneous if an attempt is made to rationalize an observed force with a knowledge of an uncharged surface; 4) the shape of the stylus tip can be very important, and a failure to take this into account can result in incorrect conclusions, a point made by other workers; and 5) when L is nonzero, the presence of charges and dipoles can yield a force that can be nonmonotonic as a function of ionic concentration