Development of a surface to increase retinal pigment epithelial cell (ARPE-19) proliferation under reduced serum conditions

Age related macular degeneration of the eye is brought about by damage to the retinal pigment epithelium (RPE) and is a major cause of adult blindness. One potential treatment method is transplantation of RPE cells grown in vitro. Maintaining RPE cell viability and physiological function in vitro is a challenge, and this must also be achieved using materials that can be subsequently used to deliver an intact cell sheet into the eye. In this paper, plasma polymerisation has been used to develop a chemically modified surface for maintaining RPE cells in vitro. Multiwell plates modified with a plasma copolymer of allylamine and octadiene maintained RPE cell growth at a level similar to that of TCPS. However, the addition of bound glycosaminoglycans (GAGs) to the plasma polymerised surface significantly enhanced RPE proliferation. Simply adding GAG to the culture media had no positive effect. It is shown that a combination of plasma polymer and GAG is a promising method for developing suitable surfaces for cell growth and delivery, that can be applied to any substrate material.     

Integrated Photoelasticity for Axisymmetric Thermal Stress Measurement Using the Stress Function

By investigating axisymmetric stress fields with integrated photoelasticity, the stress components σ_z and σ_rz can be determined directly from the experimental data. Stress components σ_r and σ_θ are usually determined using the equilibrium and compatibility equations. In this article it is shown that the stress function for an axisymmetric thermoelastic stress field can be determined on the basis of experimental data, obtained with integrated photoelasticity. Knowledge of the stress function permits one to calculate all the stress components as well as the temperature field in the test object.     

Alternative equations of magnetophotoelasticity and approximate solution of the inverse problem

In magnetophotoelasticity, photoelastic models are investigated in a magnetic field in order to initiate rotation of the plane of polarization that is due to the Faraday effect. The method has been used for the measurement of stress distributions that are in equilibrium on the wave normal and therefore cannot be measured with the traditional photoelastic technique. In this category belong bending stresses in plates and shells and residual stresses in glass plates. Two new systems of equations of magnetophotoelasticity are derived. One of them describes evolution of the polarization of light in a magnetophotoelastic medium in terms of eigenvectors, the other in terms of distinctive parameters. For the latter system, an approximate closed-form solution has been found. The integral Wertheim law has been generalized for the case of stress states in equilibrium when rotation of the plane of polarization is present.     

Identification of cross-linked peptides for protein interaction studies using mass spectrometry and 18O labeling

A new method is presented to screen proteolytic mass maps of cross-linked protein complexes for the presence of cross-linked peptides and for the verification of proposed structures. On the basis of the incorporation of 18O from isotopically enriched water into the C-termini of proteolytic peptides, cross-linked peptides are readily distinguished in mass spectra by a characteristic 8 amu shift. This is due to the incorporation of two 18O atoms in each C-terminus, so that normal and surface-labeled peptides shift 4 amu and cross-linked peptides containing two C-termini will shift 8 amu compared with their unlabeled counterparts. The method is fast, sensitive, and reliable and can be combined with any available cross-linking reagent and a wide range of proteolytic agents. As proof of principle, we successfully applied the method to a complex of two DNA repair proteins (Rad18-Rad6) and identified the interaction domain.     

A proposal to define a core of a scientific subject: A definition using concentration and fuzzy sets

Determining the core of a field"s literature, i.e. its "most important" sources, has been and still is an important problem in bibliometrics. In this article an exact definition of a core of a bibliography or a conglomerate is presented. The main ingredients for this definition are: fuzzy set theory, Lorenz curves and concentration measures. If one prefers a strict delineation, the fuzzy core can easily be defuzzified. The method we propose does not depend on the subjective notion of "importance". It is, moreover, completely reproducible. The method and the resulting core is also independent of the mathematical function (Lotka, Zipf, Bradford, etc.) that may be used to describe the relation between the set of sources and that of items.     

Process standardization for optimal virus recovery and removal of substrate DNA and bovine serum proteins in Vero cell-derived rabies vaccine

Purification of a rabies vaccine by a single zonal centrifugation run was replaced by two runs with optimal standardization of the sucrose density gradient. As a result, significant reductions in the levels of substrate DNA and bovine serum protein in the Vero cell-derived human rabies vaccine were achieved. Following many trials, for the first run, loading of the 3.2-l capacity K-3 rotor with 1800 ml of 60% sucrose solution and 1400 ml of vaccine PBS buffer solution gave a satisfactory linear gradient. However, after the first run, the substrate DNA and bovine serum contents exceeded the required levels. After protamine sulphate and Tween-80 treatment of the concentrated inactivated material, a second run using the same procedure as in the first run was tried. However, these purification procedures resulted in low virus recovery. To achieve optimal virus recovery, and removal of substrate DNA and bovine serum protein, the peak fractions from the first run as indicated by the haemagglutination, sucrose concentration, and optical density values were pooled and the sucrose concentration of the pooled fractions was increased to 60%. A second (flotation) run was then carried out. Using this method, the virus recovery rate was more than 95% that of the first run, and the levels of cellular DNA and bovine serum protein were well within the acceptable limits of less than 100 pg/dose and one part per million, respectively. The substrate DNA was quantified by both radioactive labeling and non-radioactive biotin labeling methods. For the quantification of calf serum protein, a counter-immunoelectrophoresis method was developed and effectively applied. A potency assay was performed using the National Institutes of Health (NIH) and well-standardized in vitro single radial immuno diffusion (SRD) methods. Finally, an immunogenicity study was conducted with human volunteers and the results were confirmed by a rapid fluorescent focus inhibition test (RFFIT).     

Recombinant fusion proteins for the industrial production of disulfide bridge containing peptides: purification, oxidation without concatamer formation, and selective cleavage

The relative contributions of protein tyrosine kinases (PTKs) and protein kinase C isoenzymes (PKCs), a family of serine/threonine kinases, in integrin alpha(IIb)beta3 (glycoprotein IIb/IIIa) exposure are the subject of much controversy. In the present study we measured the effect of the PTK inhibitor herbimycin A and the PKC inhibitor bisindolylmaleimide I on 125I-fibrinogen binding to alpha(IIb)beta3 and on aggregation/secretion induced by different agonists. Dose-response studies showed complete inhibition of alpha(IIb)beta3 exposure by 30 micromol/L (ADP stimulation) and 35 to 40 micromol/L (alpha-thrombin stimulation) herbimycin A. In contrast, inhibition of exposure by bisindolylmaleimide I varied from none (for ADP and epinephrine), to 30% (for platelet-activating factor), and to approximately 80% (for alpha-thrombin). Studies with a submaximal dose of herbimycin A (approximately 50% inhibition of the ADP-response) and a maximal dose of bisindolylmaleimide I showed that optical aggregation had a similar sensitivity to the inhibitors as alpha(IIb)beta3 exposure with minimal interference by secreted ADP. Thus, the relative contributions of tyrosine and serine/threonine kinases in alpha(IIb)beta3 exposure and aggregation differ among the different agonists, with an exclusive role for PTKs in ADP- and epinephrine-induced responses and a role for both PTKs and PKCs in responses induced by platelet-activating factor and alpha-thrombin.     

Microwave and far-infrared propagation in conductive microstructured composites

In a recent publication [T.E. Huber, L. Luo, Appl. Phys. Lett. 70 (1997) 2502.] we discussed a structural enhancement of the electromagnetic transmission in composites. We showed that densely packed arrays (76% volume fraction) of 10 μm diameter parallel indium wires exhibit an enhanced transmission, of 10³, relative to an indium foil of equal thickness for far-infrared (10 cm⁻¹ < k < 80 cm⁻¹) propagating along the wire length. The absorption increases as k^{0.45 ± 0.07} and is explained by the dynamic Maxwell-Gannett model, which includes eddy current dissipation. It is of considerable practical interest to extend the range of application of these composite properties to the microwave domain. Here we will discuss recent measurements of the transmission of microwaves in wire array composites in the 8–12 GHz frequency range.     

Emission characteristics of some dilute tungsten alloys

The effective work functions of nine tungsten-base alloys were measured. The second constituents were: 1) 5 pct Re, 2) 15 pct Re, 3) 2.5 pct Os, 4) 5 pct Os, 5) 1 pct Ir, 6) 2 pct Ir, 7) 5 pct Ta, 8) 10 pct Ta, and 9) 26 pct Ta. Work functions were determined from vacuum emission vehicle and thermionic emission microscope measurements. Mosaics of each surface were produced from the microscope which show the grain structure of the alloys and some anomalous emission areas. Some exceptionally high bare work functions were observed from samples with small alloy additions. For example, the work function of W1 pct Ir was 5.3 eV. DEAN LEO JACOBSON, formerly Section Manager, Electro-Optical Systems, Division of Xerox Corp., Pasadena, Calif.