Biomedical subjects

R A Mosher

Publications and source records attributed to R A Mosher.

12 recordsLinked to original sources

PubMed1995

Impact of electroosmosis on isotachophoresis in open-tubular fused-silica capillaries: analysis of the evolution of a stationary steady-state zone structure by computer simulation and experimental validation.

A dynamic computer model for simulation of open-tubular capillary electrophoresis that includes in situ calculation of electroosmosis along the fused-silica capillary column has been applied to the characterization of an anionic isotachophoretic system in presence of a cathodic electroosmotic flow. For each column segment, electroosmosis is calculated with the use of a wall mobility, the voltage gradient and the degree of dissociation of the silanol surface groups of the capillary wall. Then, the bulk capillary flow is taken to be the average of all of the segment flows and considered to represent a plug flow. This simple approach enables the combined simulation of the temporal behavior of an isotachophoretic zone structure in presence of electroosmosis. For a model anionic isotachophoretic configuration at pH 6, simulation data reveal the asymptotic formation of a stationary steady-state zone configuration in which electrophoretic and electroosmotic zone displacements are opposite and of equal magnitude. The position of the stationary boundaries are predicted to be dependent on the selected wall pK and mobility values. For two different instruments, qualitative agreement between experimental data and simulation results obtained with a wall pK between 5 and 6 is demonstrated. However, for the two experimental setups, significant differences in electroosmotic pumping (i.e. wall mobility values) are noted.

Anions↗

PubMed1990

Experimental and theoretical description of the isotachophoretic behavior of serum albumin.

The isotachophoretic behavior of serum albumin is examined for three anionic and one cationic electrolyte systems by (i) computer simulation, (ii) capillary isotachophoresis (ITP) and (iii) continuous flow ITP. The theoretical relationship between pH of the leading electrolyte and the steady state protein plateau concentration is presented for one of the anionic systems. With leading ion concentrations of the order of 10 mM, experimental protein plateau concentrations of 1.3-2.3% w/v are obtained. The computer predictions are approximately half these values.

Aminocaproic Acid↗

PubMed1990

Isotachophoretic zone formation of serum albumin in different free fluid electrophoresis instruments.

The isotachophoretic behavior of a model protein, serum albumin, was examined (i) by computer simulation, (ii) by capillary isotachophoresis in HPE 100 and Tachophor 2127, (iii) by continuous flow isotachophoresis in Elphor VaP 22 and the BIO-STREAM Separator and (iv) by recycling isotachophoresis in an apparatus of our own design. Variations in monitored zone shapes can be explained by differences in engineering aspects and fluid stabilization principles of the instruments.

Cations↗

PubMed1990

Experimental and theoretical dynamics of isoelectric focusing: IV. Cathodic, anodic and symmetrical drifts of the pH gradient.

The production of anodic, cathodic and symmetrical drifts of a pH 3.5-10 gradient formed by isoelectric focusing in polyacrylamide gels is demonstrated experimentally by manipulation of the electrolyte concentrations. Experimental behavior is reproduced by computer simulation of a model mixture of 15 hypothetical carrier ampholytes whose pIs span the pH range 3-10. The mechanism which produces the drifts is elucidated and approaches to minimize such drifts are discussed. The data suggest why most experimentally observed drifts are cathodic.

Computer Simulation↗

PubMed1990

The use of metal ion-supplemented buffers to enhance the resolution of peptides in capillary zone electrophoresis.

The potential of metal ion-containing buffers to enhance the resolution of peptides in capillary zone electrophoresis was evaluated. The impact of adding Cu(II) and Zn(II) salts to electrophoresis buffers is shown to affect the migrational behavior of several dipeptides containing histidine. Interaction with a metal ion differentially decreases the electrophoretic mobilities of peptides which comigrate in the absence of metal ions, thus causing their separation. This effect is obtained at low pH where the large net charge on the samples yields short analysis times. The dependence of the resolution on Zn(II) concentration is presented for two different samples. The influence of the background buffer is discussed.

Buffers↗

PubMed1989

Computer simulation and experimental validation of the electrophoretic behavior of proteins.

A mathematical model of the electrophoretic behavior of proteins is presented. The Debye-Hückel-Henry theory is used for the description of protein mobility, which has the important result of making net mobility a function of ionic strength. A net charge vs pH relationship and a diffusion coefficient are required to describe a specific protein. The model is employed for the computer simulation of three distinct electrophoretic modes: isoelectric focusing, isotachophoresis, and zone electrophoresis. The validity of the model is tested by comparing simulation with experimental data. Excellent qualitative agreement was found.

Computer Simulation↗

PubMed1989

The impact of boundary conditions in free flow field step electrophoresis.

The impact of the type of membrane used to separate the electrolyte compartments from the separation cell for continuous flow field step electrophoresis is examined using both experimental and computer simulation data. It is shown that the distinct transport characteristics of ion exchange membranes and dialysis membranes may have an effect on the stability of the pH and conductivity gradients created by the buffer system. These effects are interpreted using computer simulation data. The limitations of the model used to portray the transport characteristics of ion exchange membranes are discussed as are the experimental conditions which provide the greatest stability for field step electrophoresis.

Buffers↗

PubMed1985

Characterization of synthetic carrier ampholytes by repetitive scanning of the electric field during focusing.

This paper reports the utilization of a potential gradient array detector for monitoring the dynamics of the electric field during isoelectric focusing. Transient and steady state electric field profiles are presented for synthetic carrier ampholyte mixtures with a wide (approximately 3-10) pH range. Two available commercial products (Ampholine and Pharmalyte) and a laboratory synthesized mixture (PEHA ampholytes) are compared. The formation of conductivity gaps and their migration toward the cathode in extended experiments (cathodic drift) can be visualized with this system.

Ampholyte Mixtures↗

PubMed1983

Electrophoresis: mathematical modeling and computer simulation.

A mathematical model of electrophoretic separation processes has been developed and adapted for computer simulations. The model is used to predict the characteristic behavior of a variety of electrophoretic techniques from a knowledge of chemical equilibria and physical transport phenomena. The model provides a unifying basis for a rational classification of all electrophoretic processes.

Computers↗

PubMed1981

Mathematical modeling and computer simulation of isoelectric focusing with electrochemically defined ampholytes.

A mathematical model of isoelectric focusing at the steady state has been developed for an M-component system of electrochemically defined ampholytes. The model is formulated from fundamental principles describing the components' chemical equilibria, mass transfer resulting from diffusion and electromigration, and electroneutrality. The model consists of ordinary differential equations coupled with a system of algebraic equations. The model is implemented on a digital computer using FORTRAN-based simulation software. Computer simulation data are presented for several two-component systems showing the effects of varying the isoelectric points and dissociation constants of the constituents.

Journal Article↗

PubMed1979

Bacteriophage T4-coded dihydrofolate reductase: synthesis, turnover, and location of the virion protein.

Dihydrofolate reductase plays a dual role in bacteriophage T4, first, as an enzyme of thymidylate metabolism, and second, as a protein component of the tail baseplate. Antibody to the purified enzyme has been used to study its synthesis and intracellular turnover. The antibody specifically precipitates one protein from T4D-infected cell extracts. This has been identified as dihydrofolate reductase, although the polypeptide molecular weight (22,000) is lower than that earlier determined for this enzyme. The protein comigrates on gels with pY, a genetically undefined protein component of the baseplate. However, it is not pY, for pY is synthesized late in infection, whereas virtually no dihydrofolate reductase synthesis occurs later than 10 min after infection at 37 degrees C. Dihydrofolate reductase, once formed, is neither degraded nor converted to proteins of higher or lower molecular weight. Thus, it is probably incorporated into virions at the same molecular weight as that of the soluble enzyme. 125I-radiolabeled antibody binds to the wedge substructure of the baseplate, and this binding is blocked by preincubation with purified T4 dihydrofolate reductase. Thus, the enzyme protein seems to be a component of the wedge.

Immunodiffusion↗

PubMed1977

Bacteriophage T4 virion dihydrofolate reductase: approaches to quantitation and assessment of function.

This paper is concerned with the physiological role(s) of T4 phage-coded dihydrofolate reductase, which functions both in DNA precursor metabolism and as a virion protein. (i) We have detected enzyme activity in noninfectious particles produced under restrictive conditions by gene 11 mutants. This supports the conclusion of Kozloff et al. (J. Virol. 16:1401-1408, 1975) that the protein lies in the baseplate, covered by the gene 11 protein. (ii) We have obtained further evidence for virion dihydrofolate reductase as the target for neutralizing activity of T4 dihydrofolate reductase antiserum and as a determinant of the heat lability of the virion. This derives from our observation that the reductases specified by T4B and T4D differ in several properties. (iii) We have investigated several anomalous properties of T4 mutants bearing deletions that reportedly extend into or through the frd gene, which codes for dihydrofolate reductase. Evidence is presented that the deletions in fact do not extend through frd. These strains direct the synthesis of material that cross-reacts with antiserum to homogeneous dihydrofolate reductase. Moreover, they are all quite sensitive to the phage-neutralizing effects of this antiserum. In addition, they are restricted by several of the hospital strains, wild-type strains of Escherichia coli supplied by the California Institute of Technology group. (iv) We have attempted to detect dihydrofolate reductase among early-synthesized proteins present in T4 tails. Two such proteins are seen, one of which is evidently the gene 25 product and one that is a bacterial protein. Quantitation of our electrophoretic technique has allowed determination of the number of molecules of some T4 tail components present per virion. (v) Finally, we have compared the T4 dihydrofolate reductase with the corresponding enzyme specified by two plasmids conferring resistance to trimethoprim (Skold and Widh, J. Biol. Chem. 249:4324-4325, 1974). Although the enzymes are similar in some properties, they differ in several important respects, including immunological activity.

Coliphages↗