The atomic force microscope: a new tool for artificial organ research

The atomic force microscope (AFM) was developed by modifying the scanning tunneling microscope (STM). It has high resolution on the subnanometer scale (10⁻¹⁰ m), does not require troublesome preprocessing of the sample, and permits observation of living samples. With these attractive features, the AFM is expected to be a new research tool in the field of artificial organs in the near future. This review describes the history and mechanism of the AFM and some of our observations of biological samples.     

Structural aspects of the extracellular matrix of the tendon: an atomic force and scanning electron microscopy study

The mutual interactions of small proteoglycans with collagen fibrils in the extracellular matrix remain to be completely understood. The present research investigated the extracellular matrix of the rat tail tendon by atomic force microscopy (AFM) as well as by scanning electron microscopy (SEM). Observations showed simply dehydrated specimens made of large heterogeneous fibrils, tightly packed in mutual contact with no visible interfibrillar spaces. Proteoglycans usually extended onto neighboring fibrils, forming an intricate interfibrillar weaving highly sensitive to chondroitinase digestion. Pre-treatment with cupromeronic blue only affected the proteoglycans side chains, which appeared better preserved but somewhat thickened. Observation of hydrated specimens by AFM confirmed the close packing of collagen fibrils and the abundance of collagen-bound proteoglycans. Interfibrillar bridges were only occasionally observed in this tissue, whose fibrils are instead tightly bound together by proteoglycans in a structure quite consistent with its functional requirements. The molecular machinery responsible for these interactions is the subject of ongoing research.     

Probing biopolymers with the atomic force microscope: a review

This short review presents an overview of atomic force microscopy (AFM) of biopolymers and specific examples of some of the biopolymers that have been analyzed by AFM. These specific examples include extracellular polymeric substances on the surfaces of bacterial biofilms, condensed DNA, DNA constructs, and DNA-protein interactions. In addition, two examples are presented for AFM analyses of proteins: laminin flexing its arms in solution and neurofilaments entropically brushing away the space around themselves.     

Use of atomic force microscopy as a diagnostic tool to identify orthopoxvirus

Atomic force microscopy (AFM) is a versatile technique that permits the imaging of surfaces and generates topographical images from a variety of materials. Due to the fact that AFM requires minimum sample manipulation, it is a valuable tool for studying biological materials such as cells, DNA, bacteria and viruses. The aim of the present study was to standardize the AFM technique as a diagnostic tool for detection of naturally occurring orthopoxviruses. The samples analyzed were collected during natural outbreaks of Vaccinia virus (VACV) in dairy cattle in Brazil. These viruses are zoonotic infections; and therefore safe manipulation of all samples is required. The AFM technique would provide a more secure way to diagnose infection. By using the "in air" AFM technique after purification and inactivation process, relatively crude preparations of viruses were visualized rapidly. Details for efficient sample preparation and AFM imaging are described. The AFM technique provides a rapid and biosecure tool for the diagnosis of emerging orthopoxviruses and has potential as a tool for screening bioterrorism samples.     

Single-cell elastography: probing for disease with the atomic force microscope

The atomic force microscope (AFM) is emerging as a powerful tool in cell biology. Originally developed for high-resolution imaging purposes, the AFM also has unique capabilities as a nano-indenter to probe the dynamic viscoelastic material properties of living cells in culture. In particular, AFM elastography combines imaging and indentation modalities to map the spatial distribution of cell mechanical properties, which in turn reflect the structure and function of the underlying cytoskeleton. Such measurements have contributed to our understanding of cell mechanics and cell biology and appear to be sensitive to the presence of disease in individual cells. This chapter provides a background on the principles and practice of AFM elastography and reviews the literature comparing cell mechanics in normal and diseased states, making a case for the use of such measurements as disease markers. Emphasis is placed on the need for more comprehensive and detailed quantification of cell biomechanical properties beyond the current standard methods of analysis. A number of technical and practical hurdles have yet to be overcome before the method can be of clinical use. However, the future holds great promise for AFM elastography of living cells to provide novel biomechanical markers that will enhance the detection, diagnosis, and treatment of disease.     

The subfibrillar arrangement of corneal and scleral collagen fibrils as revealed by scanning electron and atomic force microscopy

The present study was designed to analyze the subfibrillar structure of corneal and scleral collagen fibrils by scanning electron microscopy (SEM) and atomic force microscopy (AFM). Isolated collagen fibrils of the bovine cornea and sclera were fixed with 1% OsO4, stained with phosphotungstic acid and uranyl acetate, dehydrated in ethanol, critical point-dried, metal-coated, and observed in an in-lens type field emission SEM. Some isolated collagen fibrils were fixed with 1% OsO4, dehydrated, critical point-dried and observed without metal-coating in an AFM. Isolated collagen fibrils treated with acetic acid were also examined by SEM and AFM. SEM and AFM images revealed that corneal and scleral collagen fibrils had periodic transverse grooves and ridges on their surface; the periodicity (i.e., D-periodicity) was about 63 nm in the cornea and about 67 nm in the sclera. Both corneal and scleral collagen fibrils contained subfibrils running helicoidally in a rightward direction to the longitudinal axis of the fibril; the inclination angle was about 15 degrees in the corneal fibrils and 5 degrees in the scleral fibrils. These findings indicate that the different D-periodicity between corneal and scleral fibrils depends on the different inclinations of the subfibrils in each fibril. The present study thus showed that corneal collagen fibrils differ from scleral collagen fibrils not only in diameter but also in substructure.     

Atherosclerotic aortic aneurysm (based on observations with the scanning electron microscope)]

The inner surface of the wall of atherosclerotic aneurism represented a sort of large ulcerative atherosclerotic plaque. Its individual areas were characterised by the presence of pseudo-hypertrophy of the endothelium cell nuclei, microplaques, and desorganization of the microrelief. The inner surface of the aneurism was de-endotheliolized on a considerable area, and here there were revealed deposits of lipids in the form of millet-like formations and smooth bodies. Studies of the inner surface of the aneurism wall carried out using raster-electron-microscopy technique made it possible to identify different types of calcinosis in the form of individual and isolated plates of calcium on its inner surface. Desorganization of the microrelief and de-endotheliation of the microrelief of the aneurism wall were the leading causes of parietal thromboformation.     

Biomaterials for orthopedics: a roughness analysis by atomic force microscopy

We conducted an AFM analysis of roughness on 7 materials widely used in bone reconstruction. Roughness was evaluated by measuring Root Mean Square (RMS) values and RMS/average height (AH) ratio, in different dimensional ranges, varying from 100 microns square to a few hundreds of nanometers. The results showed that Titanium presented a lower roughness than the other materials analyzed, frequently reaching statistical significance. On the contrary, bioactive materials, such as hydroxyapatite (HA) and bioactive glasses, demonstrated an overall higher roughness. In particular, this study focuses attention on AP40 and especially RKKP, which proved to have a significant higher roughness at low dimensional ranges. This determines a large increase in surface area, which is strongly connected with osteoblast adhesion and growth and to protein absorption. Therefore, the biointegration properties of bioactive glasses can also be given as answer in terms of surface structures in which chemical composition can influence directly the biological system (e.g. with chemical exchanges and development of specific surface electrical charge) and indirectly, via the properties induced on tribological behavior that expresses itself during the smoothing of the surfaces. We also test two new bioactive glasses, RBP1 and RBP2, with a chemical composition similar to AP40, but with some significant small additions and substitutions of components, in order to make preliminary considerations on their potential role in orthopedics.     

Elasticity measurement of living cells with an atomic force microscope: data acquisition and processing

Elasticity of living cells is a parameter of increasing importance in cellular physiology, and the atomic force microscope is a suitable instrument to quantitatively measure it. The principle of an elasticity measurement is to physically indent a cell with a probe, to measure the applied force, and to process this force–indentation data using an appropriate model. It is crucial to know what extent the geometry of the indenting probe influences the result. Therefore, we indented living Chinese hamster ovary cells at 37°C with sharp tips and colloidal probes (spherical particle tips) of different sizes and materials. We furthermore developed an implementation of the Hertz model, which simplifies the data processing. Our results show (a) that the size of the colloidal probe does not influence the result over a wide range (radii 0.5–26 μm) and (b) indenting cells with sharp tips results in higher Young’s moduli (∼1,300 Pa) than using colloidal probes (∼400 Pa).     

The use of a desktop scanning electron microscope as a diagnostic tool in studying fibrin networks of thrombo-embolic ischemic stroke

Proneness to the formation of tight and rigid fibrin networks has been shown to be independently associated with thrombotic disease. These changes may be visible long before the actual event. Previous research has shown that there is a fundamental difference between fibrin network architecture of controls compared to fibrin networks of patients 48?h post-thrombo-embolic ischemic stroke. This conclusion was made using a high-tech scanning electron microscope (SEM). Here the authors investigate whether ultrastructure of these networks can be successfully analyzed when using a smaller, desktop SEM. Such a screening procedure would not only be inexpensive, but could potentially warn patients of a possible thrombotic event long before any symptoms are prevalent. Platelet-rich plasma, obtained from healthy volunteers and thrombo-embolic ischemic stroke patients (48 h poststroke), was activated by the addition of thrombin. Fibrin networks were compared using a Zeiss ULTRA plus FEG-SEM with InLens and a desktop portable ZEOL SEM (ZEOLNeoScope). This desktop version produces micrographs that may easily be analyzed, and the information gained by studying the micrographs was comparable to that of the Zeiss ULTRA plus FEG-SEM. Such a desktop machine might be used as a screening tool or to identify individuals with risk, before the actual event. In addition, it may provide valuable information in recovering stroke patients.     

两种脱敏剂封闭牙本质小管的扫描电镜观察

背景：MS Coat、极固宁脱敏剂均可以有效封闭牙本质小管,隔绝外界刺激,降低牙本质敏感。国内外学者对两者效果的对比多为描述性、定性研究。 目的：通过扫描电镜定量观察MS Coat、极固宁脱敏剂对牙本质小管的封闭效果,并做出比较。 方法：将30颗人离体新鲜第三磨牙样本制成3 mm厚的牙本质盘,以17%EDTA液体处理牙本质表面2 min,随机均分为3组,空白对照组不做任何处理,MS Coat组和极固宁组分别用MS Coat脱敏剂和极固宁脱敏剂对牙本质表面进行处理,采用扫描电镜观察各组牙本质表面和纵剖面结构。 结果与结论：①表面观察：空白对照组牙本质小管全部开放;MS Coat组绝大部分牙本质小管口被堵塞;极固宁组牙本质小管口覆盖不规则鳞状晶体物质,仍可见一小部分暴露的牙本质小管口,但直径变窄。②纵剖面观察：空白对照组牙本质小管呈条状平行排列,无任何堵塞物;MS Coat组小管口有沉淀物沉积,较致密,基本上封闭了牙本质小管口;极固宁组牙本质小管内有鳞片状晶体物质沉积。组间牙本质直径及小管数量比较：空白对照组>极固宁组> MS Coat组(P < 0.05)。表明MS Coat脱敏剂和极固宁脱敏剂均能有效封闭牙本质小管,且MS Coat脱敏剂的封闭效果优于极固宁。 中国组织工程研究杂志出版内容重点：生物材料;骨生物材料; 口腔生物材料; 纳米材料; 缓释材料; 材料相容性;组织工程     

Characteristics of the microstructure of biliary calculi based on data from scanning electron microscopy

Scanning electron microscopy was employed to study cholelithic microstructure. Irrespective of the chemical composition, the gall-stones demonstrated the presence of a cementing matrix composed of micellar-vesicular particles occurring either in aggregates or regularly spreading along the facets about the surface of crystalloid structures. There appeared to be a close relationship between the latter and the particles which are, viz., vesicles, responsible for the plane and margin epitaxy of the cryslalloid structures. Any cholesterol deposits are lipoprotein-like particles or their derivatives, appearing as micellar-vesicular particles in case of choleliths.     

Techniques for microdrop analysis of fluids (sweat, saliva, urine) with an energy-dispersive X-ray spectrometer on a scanning electron microscope

Volumes of 10(-10) liter of sweat, saliva, and urine were analyzed with an energy-dispersive spectrometer (EDS) X-ray analyzer on a standard scanning electron microscope (SEM). Results of the EDS analysis of diluted and undiluted samples were verified by comparison with results of analyses of the same sample prepared by conventional quantitative methods. Except for Cl in concentrated urines and Mg in low concentrations, good agreement was found between the methods. The analysis provides quantitation of most elements of biological interest present in concentrations of about 1 mM, while it demonstrates good linearity (r greater than 0.99) throughout a wide range of commonly encountered biological concentrations. Reproducibility of the analysis is on the order of 2% and the minimal determinable concentration is generally between 0.5 and 1.0 mM for S, P, K, and Ca and slightly more than 1.0 mM for Mg.     

Operating force information on-line acquisition of a novel slave manipulator for vascular interventional surgery

Vascular interventional surgery has its advantages compared to traditional operation. Master-slave robotic technology can further improve the operation accuracy, efficiency and safety of this complicated and high risk surgery. However, on-line acquisition of operating force information of catheter and guidewire remains to be a significant obstacle on the path to enhancing robotic surgery safety. Thus, a novel slave manipulator is proposed in this paper to realize on-line sensing of guidewire torsional operating torque and axial operation force during robotic assisted operations. A strain sensor is specially designed to detect the small scale torsional operation torque with low rotational frequency. Additionally, the axial operating force is detected via a load cell, which is incorporated into a sliding mechanism to eliminate the influence of friction. For validation, calibration and performance evaluation experiments are conducted. The results indicate that the proposed operation torque and force detection device is effective. Thus, it can provide the foundation for enabling accurate haptic feedback to the surgeon to improve surgical safety.     

Uncombable hair (pili trianguli et canaliculi): evidence for dominant inheritance with complete penetrance based on scanning electron microscopy

Uncombable or spun-glass hair (pili trianguli et canaliculi) is an uncommon condition in which the hair is "unmanageable" and has a distinct appearance on scanning electron microscopy. The hair is usually grossly abnormal in infancy and childhood, but may become normal later in life. Although dominant inheritance has been observed, most cases have been sporadic. Both recessive and dominant transmission with incomplete penetrance have been suggested as modes of inheritance. We report the occurrence of this condition in a young girl, her brother, and her father. Although the proposita and her brother had characteristically uncombable hair, their father appeared normal and denied any history of hair abnormality. However, the characteristic hair morphology was observed on scanning electron microscopy in all 3 relatives, documenting dominant transmission and complete penetrance of the gene in this family.     

Morphology of the antenna of Dermatobia hominis (Diptera: Cuterebridae) based on scanning electron microscopy

Sensilla of the antennae of male and female Dermatobia hominis were studied by scanning electron microscopy. The images obtained were interpreted according to the scientific literature referring to the sensory structures of insects. Sensilla of the "long bristle" type and smaller spines of the "microtrichia" type were found in different numbers and patterns of distribution on the scape and pedicel. Coeloconic sensilla with longitudinal cuticular furrows were observed on the female flagellum, as well as two varieties of basiconic sensilla: a large one surrounded by pointed foliaceous structures and a smaller form implanted on a raised cuticular process. The larger type of basiconic sensilla was also observed on the flagellum of the male antenna, as well as a variety of coeloconic sensilla with apical dilatations. Trichoid and chaetic sensilla were encountered in greater numbers on the arista of females, with the former type predominating. Coeloconic sensilla were observed on the arista of both sexes, as well as sensilla of the "long bristle" and styloconic types exclusively in males. Adult D. hominis were observed to possess sensory structures with chemoreceptory, thermo-hygroreceptory and mechanoreceptory functions on their antennae. These results could facilitate the identification of the chemoreceptors by electrophysiological techniques. The sexual dimorphism noted for the antennae constitutes a new criterion for distinguishing between the sexes of D. hominis     

A novel tool for high-resolution transmission electron microscopy of intact interfaces between bone and metallic implants

A key feature in the understanding of the mechanisms of integration versus rejection of implanted materials is a deepened understanding of the elemental and molecular compositions of the interface zone between the surface of the synthetic man-made material and the biological components of tissue. Intact interfaces between metallic implants and tissues have not been able to image and analyse on the ultrastructural level with the common transmission electron microscopy (TEM) sample preparation techniques. By using focused ion beam microscopy for site-specific preparation of TEM samples, intact interfaces between metal implants and calcified tissue were imaged for the first time. The interface's elemental and crystallographic compositions were determined using energy dispersive X-ray mapping and electron diffraction. The developed technique fulfills a long-sought-for demand to correlate the surface properties of implanted metal prostheses with the fine structure and composition of preserved interfaces with tissues.     

Morphology of mouse epidermal cells in vitro: a scanning electron microscopy study

Scanning electron microscopy of isolated epidermal cells from newborn mice grown in vitro showed that the cultures consisted of several morphologically different types of cells. Young cultures had many smooth, round cells while older cultures contained more cells with rough or ruffled surface and a varying number of flat, irregular cells. Probably, the various cell types corresponded to different stages of differentation (keratinization). Cultures that were grown in the presence of retinyl acetate (12.5 microgram/ml) had more round and smooth cells after several days in vitro than the controls. This could indicate that retinyl acetate delayed or altered cell differentiation. Scanning electron microscopy of the cultures consistently showed that the cells were situated at different layers. The apparent monolayer seen by phase contrast microscopy therefore seems to be an optical phenomenon due to projection of the cells onto the same plane.     

Studies on a canine intestinal spirochete: scanning electron microscopy of canine colonic mucosa.

A large homogeneous population of spirochetes was observed attached to the colonic mucosa of clinically normal dogs. Their association with the colonic mucosa and their mode of attachment resembled those of a spirochete reported in primates.