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Here we show an example of scanning tunneling microscopy triggering and subsequently investigating a chemical reaction on a nanometer scale. It is observed that applying a high electric field to a graphite substrate homogeneously coated with a thin film of glutaraldehyde causes the glutaraldehyde molecules to form clusters ranging from 5 to 15 molecules in size. Studies on the lateral distribution of the clusters suggest that a lateral flux of charge between the negatively biased tip and the grounded substrate causes a type of electropolymerization of these molecules. We believe that a significant current flows due to charged species which have been created by the primary tunneling electrons and which subsequently diffuse over a large distance. We therefore propose that, in general, lateral charge transfer may play an essential role in imaging low‐conductivity molecules by distributing the current between tip and substrate over a large sample area, thereby considerably reducing the current density.

The motion of gel phase domains in inhomogeneous electric fields is observed by fluorescence microscopy with lipid monolayers at the air-water interface. Quantitative data analysis yields detailed information on the field geometry and the magnitude of the resulting forces. This knowledge then serves to derive information on monolayer dynamics, in particular on the surface viscosity η. The latter is measured as η = 0.015 P for the fluid phase of dipalmitoyl phosphatidyl choline, in order-of-magnitude agreement with published values on similar systems. Hence a technique is introduced to measure monolayer viscosities directly and microscopically.

The growth of spiral solid domains in monolayers of dimyristoylphosphatidic acid containing 1 mol% cholesterol is observed by fluorescence microscopy at the air/water interface. Spiral or lamellar domains are encountered only at high pH (11) where the large head group charge induces a chain tilt. Spiral width is drastically reduced with decreasing temperature, is uniform and can be varied reversibly.—The findings can be understood considering repulsive and anisotropically attractive electrostatic forces that increase with decreasing temperature and edge tension being reduced by incorporation of cholesterol. The shape of the spirals can be numerically calculated using a crude model containing only three adjustable parameters.

La concanavaline A affect la taille des domaines lipidiques par un arrangement preferentiel a la frontiere entre les domaines ordonnes et l'environnement fluide. Les proteines agregees presentes dans les structures formees par liaison avec le dextran modifient la forme des domaines lipidiques et augmentent la viscosite de la monocouche en participant a un reseau qui interconnecte les domaines. Des resultats similaires sont observes pour les interactions anticorps antigene

The reaction centre and the light-harvesting chlorophyll protein of the photosynthesis bacterium R. sphaeroides are reconstituted into phospholipid monolayers on a water surface, and the protein/lipid monolayer is transferred on to solid supports. The systems are studied structurally by fluorescence and electron microscopy and by film balance techniques, as well as functionally by stationary and transient optical spectroscopy. A method is described to prepare homogeneous or regularly defined inhomogeneous protein distributions in the monolayer. The latter is achieved by maintaining the monolayer in co-existence between the fluid and gel phases and making use of the finding that the proteins preferentially partition into the fluid phase. In this way, protein concentrations on the surface of about 10 11 –10 12 cm −2 can be obtained while maintaining the system functional over period of days.

Etude des couches monomoleculaires mixtes a 10°C et a un pH de 11, en fonction de la concentration en cholesterol

A new experimental set-up is described to measure surface potentials at a lateral resolution of 40 μm with an accuracy better than 5 mV. It is used to investigate the heterogeneity of a phospholipid monolayer on a solid support. By comparing the lateral potential distribution with corresponding fluorescence micrographs different surface potentials can be assigned to different monolayer phases. From this one derives that the surface potential of the fluid phase of the phospholipid dimyristoyl-phosphatidylethanolamine is lower than that of the solid phase by 90±10 mV . If account is also taken of changes in molecular densities the result leads to an estimate of the decrease in tilt angle of the hydrocarbon chains with respect to the surface normal in going from the fluid to a liquid crystalline phase. We discuss the role of water during transfer and assess various support cleaning procedures and possible transfer artifacts.

The SOS system of Escherichia coli is coordinated by two proteins: LexA, a repressor protein of several unlinked genes, and the coprotease RecA. As known to date LexA controls 31 genes with slightly different DNA binding motifs allowing for a variable degree of repression from one gene to the other. Besides the SOS system LexA plays an important role in the regulation of transcription. The protein regulates transcription by using particular motifs to bind DNA, the helix-turn-helix motif. Here, we employed AFM-based single molecule force spectroscopy to characterize the interaction of LexA protein with two different DNA motifs: recA and yebG. We measured the dissociation rates to be 0.045 s(-1) for recA and 0.13 s(-1) for yebG, respectively, which is in accordance with the predicted higher affinity between LexA-recA compared to LexA-yebG. The widths of the binding potentials were determined to be 5.4 +/- 1 angstroms and 4.9 +/- 0.5 angstroms, respectively. This short-ranged potential is characteristic for a stiff hydrogen-bonding network between protein and DNA. The unbinding occurs in a breakup rather than a gradual sliding.

Etude des couches monomoleculaires ou bimoleculaires d'octanoylphoshatidylcholine liees de facon covalente a une surface de graphite, par microscopie electronique, spectrometrie IR a transformee de Fourier et spectrometrie photoelectronique

We report the repeated observation at molecular resolution of lipid and fatty acid mono‐ and bimolecular layers on graphite in air. The films of di‐myristoyl‐phosphatidic acid and cadmium–arachidate were deposited by the Langmuir–Blodgett technique. This confirms the possibility of gaining scanning‐tunneling microscopy (STM) images by electron transfer through layers more than 50 A thick. Sample preparation, STM technique, and results are discussed.

Studies on monomolecular layers of phospholipids containing the antenna protein B800-850 (LHCP) and in some cases additionally the reaction center of the photosynthetic bacterium Rhodopseudomonas sphaeroides are reported. Information on monolayer preparation as well as on protein /lipid and protein/protein interaction is obtained by means of fluorescence spectroscopy and microscopy at the air/water interface in combination with film balance experiments. It is shown that a homogeneous distribution of functional proteins can be achieved. This can be transformed into a regular pattern-like distribution by inducing a phospholipid phase transition. Although the LHCP preferentially partitions into the fluid lipid phase, it decreases the lateral pressure necessary to crystallize the lipid. This is probably due to an increase in order of the fluid phase. A pressure-induced conformation change of the LHCP is detected via a drastic change in fluorescence yield. A highly efficient energy transfer from LHCP to the reaction center is observed. This proves the quantitative reconstitution of both types of proteins and indicates protein aggregation also in the monolayer.

The recently developed absorbable materials demonstrate a constant and reproducible high quality when compared with the resorbable suture materials. However, based on in vitro studies by different authors the use of these materials in infected urine is not recommended. We investigated in vivo in infected urines the stability of the suture materials composed of polyglycolic acid, and compared the results obtained in vitro and in vivo. In a controlled study the stability of the absorbable suture materials polyglycolic acid, polyglactin 910 and polydioxanon 4/0 USP were tested in infected urine in vivo and in vitro. In 21 patients with infected urine the suture materials were placed into the bladder for 7 days. In another experiment these materials were incubated up to 7 days in culture flasks with infected urine. After that the suture materials were examined by densitometry and electrone microscopy. When using the results obtained in vitro in clinical work caution is recommended. However, the in vivo tests show that the stability of the newly developed absorbable materials is not influenced dramatically by the infected urine.

In lumbar disc surgery ureteral transection is a rare complication and only nine such cases have been reported so far. They are analyzed and two more cases are described. The complication in the first case was recognized immediately and the patient was treated successfully by end-to-end anastomosis of the ureter. The diagnosis in the second case was delayed and this lead to a septic course and finally required nephrectomy.

Monolayers of charged and neutral phospholipids at the air/water interface containing the cytochromes b 5 and c are studied by film balance techniques and by fluorescence microscopy. A new technique is introduced to obtain a defined and homogeneous protein distribution within the membrane. It is shown that both proteins preferentially partition into the fluid membrane phases coexisting with solid lipid domains, thus allowing formation of periodic protein distributions. Protein reconstitution in protein/lipid ratios up to 1:50 does not change the pressure, π c , corresponding to the main lipid transition but changes the slope in the pressure/area isotherms. It also affects the pressure-induced lipid crystallization, in that the monolayer can be viewed as segregated into a protein-free and a protein-enriched phase. Whereas penetration of cytochrome c into the monolayer is highly dependent on lipid head group charge, this does not hold for cytochrome b . In both cases, monolayer penetration is monotonously reduced with increasing surface pressure, pointing to the dependence of hydrophobic protein-lipid interactions on hydrocarbon chain density.

Being a membrane-bound process photosynthesis depends on the functional interaction of membrane-bound or associated proteins [1] . For molecules of the photosynthetic membrane carrying out the primary process of energy and electron- transfer it is therefore essential to study protein/lipid and lipid mediated protein/protein interaction.

A correlative morphologic analysis was carried out on isolated metaphase chromosomes by means of field emission in-lens scanning electron microscopy (FEISEM) and atomic force microscopy (AFM). Whereas FEISEM provides ultra-high resolution power and allows the surface analysis of biological structures free of any conductive coating, the AFM allows imaging of biological specimens in ambient as well as in physiologic conditions. The analysis of the same samples was made possible by the use of electrical conductive and light transparent ITO glass as specimen holder. Further preparation of the specimen specific for the instrumentation was not required. Both techniques show a high correlation of the respective morphologic information, improving their reciprocal biological significance. In particular, the biological coat represents a barrier for surface morphologic analysis of chromosome spreads and it is sensitive to protease treatment. The chemical removal of this layer permits high-resolution imaging of the chromatid fibers but at the same time alters the chromosomal dimension after rehydration. The high-resolution level, necessary to obtain a precise physical mapping of the genome that the new instruments such as FEISEM and AFM could offer, requires homogeneously cleaned samples with a high grade of reproducibility. A correlative microscopical approach that utilizes completely different physical probes provides complementary useful information for the understanding of the biological, chemical, and physical characteristics of the samples and can be applied to optimize the chromosome preparations for further improvement of the knowledge about spatial genome organization.

The formation of crystalline domains of the phospholipid L-alpha-dimyristoyl-phosphatidic acid containing 1 mol% cholesterol, was studied as a function of head group charge by fluorescence microscopy with monolayers at the air/water interface. It is shown that the usual dendritic growth occurs at low pH (8), whereas spiral domains are formed at high pH (11), where the head group contains two negative charges. The findings are ascribed to an electrostatically induced chain tilt that, in conjunction with an in-plane dipole moment, causes a ferroelectric state. This allows for domain aggregation and orientation originating in elongated domains that, additionally, are bent because of the chirality of the molecules. The structure is stabilized and further elongated due to the anisotropic edge activity of cholesterol.

Chromosome-specific paint probes provide a powerful tool with wide applications in cytogenetic analysis. Here, we present a new approach using UV-laser microbeam microdissection in combination with laserpressure catapulting, which allows the fast isolation of single chromosomes for the generation of chromosome-specific paint probes. To demonstrate the feasibility of this approach, single chromosomes were collected and amplified with degenerate oligonucleotide-primed PCR, hapten-labeled and hybridized onto normal metaphase spreads. Fluorescence in situ hybridization signals revealed specific painting of the respective chromosomes.

An atomic force microscope (AFM) based system has been built for the manipulation of materials at the nanometer scale. The AFM is combined with an inverse optical microscope and an UV-laser microbeam system for photoablation. The actuators of the AFM are controlled using a digital signal processor. Real-time routines and a graphical user interface have been programmed for high resolution imaging and nanomanipulation. The nanomanipulation can be pre-programmed offline or directly performed using a low-cost haptic interface. In this paper, we discuss the whole system and the different methods for manipulation.

We compute the correlation dimension, the bifurcation diagram, and a Poincare map for experimental data obtained by dynamic atomic force microscope. From the analysis, we infer a period doubling mode and chaotic modes of the system.

Laser microbeam microdissection and laser pressure catapulting offer the possibility of separating cell compartments, thus allowing for contamination-free analysis. Using these methods, we were able to select single chloroplasts of Nicotiana tabacum. Starting from homogenized leaf material, chloroplasts were purified by differential centrifugation and applied directly onto a poly-ethylene-naphthalate membrane that was mounted on a microscope slide. Single chloroplasts were dissected under microscopic control and catapulted into a PCR tube. Subsequent PCR of a spacer region between the trnT and trnF genes verified the successful amplification of DNA from a single chloroplast. The advantage of this method compared to the use of capillaries or optical tweezers is that one is able to prepare high numbers of samples in a short time.