“…Embedding TM in Lowicryl or polymer matrix had a specific advantage for AFM, since previous studies have indicated that soft biological materials can undergo deformations under the AFM tip due to nano-Newton forces applied on the samples (Hansma et al, 1997). The polymer matrix permits features to be observed, but tip-sample interactions can cause the TM structures to be observed as circular rather than square regions, as in the tapping-mode images of Figs.…”
Section: Discussionmentioning
confidence: 99%
“…Atomic force microscopy (AFM) has become a powerful tool for imaging biological structures, embedded cellular organelles, and individual biological molecules (Lal and John, 1994;Hansma et al, 1997). The advantage of AFM over conventional techniques, such as electron microscopy (EM), is that three-dimensional information regarding the materials can be obtained at the molecular level, without the heavy metal staining as in transmission EM or surface coating employed in scanning EM (Hansma et al, 1997).…”
Section: Introductionmentioning
confidence: 99%
“…The advantage of AFM over conventional techniques, such as electron microscopy (EM), is that three-dimensional information regarding the materials can be obtained at the molecular level, without the heavy metal staining as in transmission EM or surface coating employed in scanning EM (Hansma et al, 1997). The structures of unstained cellular organelles in thin sections embedded in typical polymeric materials used for EM histology, such as Lowicryl and London Resin-White (LR-White), have been characterized (Braet et al, 1997;Yamamoto and Tashiro, 1994).…”
Pulmonary surfactant stabilizes the lung by reducing surface tension at the air-water interface of the alveoli. Surfactant is present in the lung in a number of morphological forms, including tubular myelin (TM). TM is composed of unusual 40 ؋ 40 nm square elongated proteolipid tubes. Atomic force microscopy (AFM) was performed on polymerembedded Lowicryl and London Resin-White (LRWhite) unstained thin sections. AFM was used in imaging regions of the sections where TM was detected by transmission electron microscopy (EM) of corresponding stained sections. Tapping-and contact-mode AFM imaging of the unstained sections containing TM indicated a highly heterogeneous surface topography with height variations ranging from 10 to 100 nm. In tapping-mode AFM, tubular myelin was seen as hemispherical protrusions of 30-70 nm in diameter, with vertical dimensions of 5-8 nm. In contact-mode AFM and with phase imaging using a sharper (G10 nm nominal radius) probe, square open-ended tubes which resembled typical electron micrographs of such regions were observed. The cross-hatch structures observed inside the tubes using EM were not observed using AFM, although certain multilobe structures and topographic heterogeneity were detected inside some tubes. Other regions of multilamellar bodies and some regions where such bilayer lamella appear to fuse with the tubes were found in association with TM using AFM. EM of acetone-delipidated tubes in LR-White revealed rectangular tubular cores containing cross-hatched structures, presumably protein skeletons. AFM surface topography of these regions showed hollow depressions at positions at which the protein was anticipated instead of the protrusions seen in the lipid-containing sections. Gold-labeled antibody to surfactant protein A was found associated somewhat randomly within the regions containing the protein skeletons. The topography of the gold particles was observed as sharp peaks in contact-mode AFM. This study suggests a method for unambiguous detection of threedimensional nanotubes present in low abundance in a biological macromolecular complex. Only limited detection of proteins and lipids in surfaces of embedded tubular myelin was possible. EM and AFM imaging of such unusual biological structures may suggest unique lipid-protein associations and arrangements in three dimensions.
Academic Press
“…Embedding TM in Lowicryl or polymer matrix had a specific advantage for AFM, since previous studies have indicated that soft biological materials can undergo deformations under the AFM tip due to nano-Newton forces applied on the samples (Hansma et al, 1997). The polymer matrix permits features to be observed, but tip-sample interactions can cause the TM structures to be observed as circular rather than square regions, as in the tapping-mode images of Figs.…”
Section: Discussionmentioning
confidence: 99%
“…Atomic force microscopy (AFM) has become a powerful tool for imaging biological structures, embedded cellular organelles, and individual biological molecules (Lal and John, 1994;Hansma et al, 1997). The advantage of AFM over conventional techniques, such as electron microscopy (EM), is that three-dimensional information regarding the materials can be obtained at the molecular level, without the heavy metal staining as in transmission EM or surface coating employed in scanning EM (Hansma et al, 1997).…”
Section: Introductionmentioning
confidence: 99%
“…The advantage of AFM over conventional techniques, such as electron microscopy (EM), is that three-dimensional information regarding the materials can be obtained at the molecular level, without the heavy metal staining as in transmission EM or surface coating employed in scanning EM (Hansma et al, 1997). The structures of unstained cellular organelles in thin sections embedded in typical polymeric materials used for EM histology, such as Lowicryl and London Resin-White (LR-White), have been characterized (Braet et al, 1997;Yamamoto and Tashiro, 1994).…”
Pulmonary surfactant stabilizes the lung by reducing surface tension at the air-water interface of the alveoli. Surfactant is present in the lung in a number of morphological forms, including tubular myelin (TM). TM is composed of unusual 40 ؋ 40 nm square elongated proteolipid tubes. Atomic force microscopy (AFM) was performed on polymerembedded Lowicryl and London Resin-White (LRWhite) unstained thin sections. AFM was used in imaging regions of the sections where TM was detected by transmission electron microscopy (EM) of corresponding stained sections. Tapping-and contact-mode AFM imaging of the unstained sections containing TM indicated a highly heterogeneous surface topography with height variations ranging from 10 to 100 nm. In tapping-mode AFM, tubular myelin was seen as hemispherical protrusions of 30-70 nm in diameter, with vertical dimensions of 5-8 nm. In contact-mode AFM and with phase imaging using a sharper (G10 nm nominal radius) probe, square open-ended tubes which resembled typical electron micrographs of such regions were observed. The cross-hatch structures observed inside the tubes using EM were not observed using AFM, although certain multilobe structures and topographic heterogeneity were detected inside some tubes. Other regions of multilamellar bodies and some regions where such bilayer lamella appear to fuse with the tubes were found in association with TM using AFM. EM of acetone-delipidated tubes in LR-White revealed rectangular tubular cores containing cross-hatched structures, presumably protein skeletons. AFM surface topography of these regions showed hollow depressions at positions at which the protein was anticipated instead of the protrusions seen in the lipid-containing sections. Gold-labeled antibody to surfactant protein A was found associated somewhat randomly within the regions containing the protein skeletons. The topography of the gold particles was observed as sharp peaks in contact-mode AFM. This study suggests a method for unambiguous detection of threedimensional nanotubes present in low abundance in a biological macromolecular complex. Only limited detection of proteins and lipids in surfaces of embedded tubular myelin was possible. EM and AFM imaging of such unusual biological structures may suggest unique lipid-protein associations and arrangements in three dimensions.
Academic Press
“…The use of force curves generated by single molecule sensors is potentially a useful way that information about the chemical nature, the strength of interactions, and possibly the molecular arrangement of complex biological structures can be deduced (Gad et al, 1997;Hansma et al, 1997;Li et al, 1998;Li et al, 1999;Marzalek et al, 1998;Osada et al, 1999;Okabe et al, 2000;Rief et al, 1997). To date a few attempts have been made to force probe intact plant cell walls in situ.…”
Section: Discussionmentioning
confidence: 99%
“…While imaging techniques such as transmission electron microscopy (TEM), field emission scanning electron microscopy (FESEM) and contact mode atomic force microscopy (AFM) can provide valuable information on the appearance, arrangement and structural dimensions of cell wall components (Hansma et al, 1997;Li, 1999;Thimm et al, 2002;Thimm et al, 2009), only limited chemical information can be obtained. For example, when using AFM, the contrast mechanism used to obtain an image is the force between the tip and the sample, and many samples produce the same force, thus it is difficult to reliably determine the chemical groups that are imaged.…”
Section: Imaging the Plant Cell Wall Of Higher Plantsmentioning
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