Pressure-driven sample injection with quantitative liquid dispensing for on-chip electrophoresis.

A novel pressure-driven sample injection method was developed as an alternative to electrokinetic injection, and electrophoretic separation was carried out on a microfabricated device employing this method. This method enables a defined volume of liquid dispensing, followed by instantaneous injection driven by pneumatic pressure, greatly simplifying the injection procedure. A particular microstructure, called a "metering chamber", has been designed for the quantitative dispensing of an ultra-low volume of sample liquid; a "hydrophobic passive valve" equipped with an air vent channel is employed for injecting a dispensed sample into the separation channel. The reproducibility of dispensing was 3.3% (n = 15), expressed by the variation of dispensed volumes. The electrophoretic separation of DNA fragments was performed using this injection method, varying the injection volumes from 0.45 to 4.0 nL, and the separation efficiencies were compared. This precise injection method, easily variable in injection volumes, is highly suitable for quantitative as well as qualitative electrophoretic analyses.     

CE to monitor endotoxins by protein complexation

A new CE method for fast and efficient analysis of bacterial endotoxins (lipopolysaccharides) is described. It is based on the strong interaction between proteins and endotoxins. The UV absorption of the protein component in the complex is used for the detection. The electrophoretic mobility of the complex hemoglobin/endotoxin can be employed for qualitative analysis of the endotoxin. For instance, the structural differences between "smooth" and "rough" lipopolysaccharides from Salmonella minnesota (wild-type), Salmonella minnesota R595 and Shigella sonnei R562H are reflected in the electrophoretic mobilities of their hemoglobin complex.     

样品带示踪法测定毛细管电泳过程中的电渗流

电渗流在毛细管区带电泳（CZE）和胶束电动力色谱（MEKC）中具有重要作用,通过控制电渗流可以提高分辨率或加快分析速度,因而控制和测定电渗流已成为毛细管电泳领域重要的研究课题．SalomonK．［'j建立了一种电渗流近似模型,但因影响因素复杂,电渗流还无法从理论上计算,只能通过实验测定．目前常用的测定法有中性示踪剂法［'－"、称量法［'和电泳电流监测法［'j本文提出一种新的电渗流测定法--样品带示踪法,该法不必另加示踪剂,利用样品带中的基体介质即可示踪电渗流．该法适用于较宽的I。H范围及多种检测器,对于某些样品（与…     

两步电泳沉积制备TiO2光阳极用于高效染料敏化太阳能电池

采用不同沉积电压制备TiO₂光阳极,研究电压对薄膜沉积速率、厚度和形貌的影响。通过台阶仪、光学照片、扫描电子显微镜(SEM)、电化学交流阻抗谱、开路电压衰减曲线对光阳极和电池进行系统表征,并测试了染料敏化太阳能电池器件的电流密度-电压(J-V)曲线,计算其光电转换效率。结果表明,提高沉积电压时,光阳极薄膜的沉积速率加快,膜厚也增加,但是电压过高时,薄膜会有裂缝和覆盖不全的问题,这会对电池的效率造成负面影响。综合考虑低沉积电压条件下薄膜均匀无裂缝和高沉积电压条件下沉积速率快的优点,采用先30 V电压、后60 V电压的电泳沉积方式来制备光阳极,结果呈现协同效果,既降低了制备时间又得到高质量的薄膜,在无其他修饰的情况下,电池的光电转换效率可以达到7.29%。     

两步电泳沉积制备TiO2光阳极用于高效染料敏化太阳能电池

采用不同沉积电压制备TiO₂光阳极,研究电压对薄膜沉积速率、厚度和形貌的影响。通过台阶仪、光学照片、扫描电子显微镜（SEM）、电化学交流阻抗谱、开路电压衰减曲线对光阳极和电池进行系统表征,并测试了染料敏化太阳能电池器件的电流密度-电压（J-V）曲线,计算其光电转换效率。结果表明,提高沉积电压时,光阳极薄膜的沉积速率加快,膜厚也增加,但是电压过高时,薄膜会有裂缝和覆盖不全的问题,这会对电池的效率造成负面影响。综合考虑低沉积电压条件下薄膜均匀无裂缝和高沉积电压条件下沉积速率快的优点,采用先30 V电压、后60 V电压的电泳沉积方式来制备光阳极,结果呈现协同效果,既降低了制备时间又得到高质量的薄膜,在无其他修饰的情况下,电池的光电转换效率可以达到7.29%。     

Electric and hydrodynamic stretching of DNA-polymer conjugates in free-solution electrophoresis

The conjugation of an uncharged polymer to DNA fragments makes it possible to separate DNA by free-solution electrophoresis. This end-labeled free-solution electrophoresis method has been shown to successfully separate ssDNA with single monomer resolution up to about 110 bases. It is the aim of this paper to investigate in more detail the coupled hydrodynamic and electrophoretic deformation of the ssDNA-label conjugate at fields below 400 V/cm. Our model is an extension of the theoretical approach originally developed by Stigter and Bustamante [Biophys. J. 75, 1197 (1998)] to investigate the problems of a tethered chain stretching in a hydrodynamic flow and of the electrophoretic stretch of a tethered polyelectrolyte. These two separate models are now used together since the charged DNA is "tethered" to the uncharged polymer (and vice versa), and the resulting self-consistent model is used to predict the deformation and the electrophoretic velocity for the hybrid molecule. Our theoretical and experimental results are in good qualitative agreement.     

Recovery of protein by coomassie brilliant blue precipitation prior to electrophoresis.

The interaction of protein with Coomassie Brilliant Blue G-250 results in formation of an insoluble protein-dye complex which can be recovered by centrifugation and redissolved for electrophoretic analysis. The precipitated protein can be washed in acetone to remove excess dye in order to enhance resolution. The residual dye becomes dissociated from the proteins on electrophoresis and can be exploited as a "dye front". The method allows simultaneous protein assay and recovery of microgram amounts of protein from dilute solution and could be widely applied for conserving, concentrating and desalting minute amounts of valuable sample prior to electrophoretic analysis.     

Electrophoretic behaviour of calcium oxalate monohydrate in liquid mixtures

An experimental investigation on electrophoresis of calcium oxalate monohydrate in several liquid mixtures (methanol-, ethanol-, 2-propanol-, and methanol-acetonewater) is described.The electrophoretic transport is considered from the view-point of the Thermodynamics of Irreversible Processes. Linear relations have always been found between the mass flux and the applied electric field. The -potential of the interface C.O.M./liquid mixture has been estimated by using the classial theory of electrophoretic mobility of colloidal particles. The influence of the composition of the mixture on the electrophoretic coefficient has been discussed on the basis of the variation of -potential, viscosity and dielectric constant with the molar fraction of alcohol for binary mixtures and acetone for ternary mixtures. The concentration dependence of the electrophoretic coefficient is found to be linear only in the latter case.     

Gradienten-Gel-Elektrophorese mit elektrophoretischer Elution im Vergleich zur Gel-Chromatographie für die Metallspezies-Analytik in Lebensmitteln

Summary The application of electrophoresis to the separation of metal species in soy bean flour extracts was studied and compared with the routinely used gel chromatography. Different electrophoretic techniques, especially gradient gel electrophoresis and isoelectric focusing were investigated. The separated substances were electrophoretically eluted, followed by the determination of the metals zinc, nickel, and copper via flame AAS. Special attention was paid to the compatibility of the distribution patterns of the electrophoretic and chromatographic separation. The application of gradient gel electrophoresis provides a useful and necessary reference method for the separation of element species. It is a sensitive separation method with a high resolving power that can provide a lot of information on the investigated species, e.g. molecular size, charge, and stability. Isoelectric focusing is useful for further characterization (i.e. isoelectric points) of separated fractions and for checking peak purity.

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Auszugsweise vorgetragen auf der GDCh-Tagung Elemente und ihre Bindungsformen in der Umwelt am 28./29. 9. 1988 in Karlsruhe     

Cardiac muscle protein analysis by high-resolution and microscale two-dimensional gel electrophoresis

An improved method of two-dimensional gel electrophoresis of small muscle samples is described. Isoelectric focusing of cardiac whole muscle homogenate in agarose gels containing urea and detergent has a markedly increased resolution. Equilibration of the first-dimensional gels with detergent before application to the second-dimensional gels is unnecessary in this system. By applying this method to rat cardiac whole muscle, high-molecular weight proteins, such as myosin heavy chains, are focused on the first-dimensional gels and, in addition, minor components are resolved on the second-dimensional gels, without loss during equilibration with detergent. The two-dimensional electrophoretic patterns of rat cardiac whole muscle obtained with this method reveal numerous clearly separated spots. By analyzing the two-dimensional electrophoretic patterns of rat cardiac whole muscle and various rat cardiac fractions, and by staining the calcium-binding proteins with "Stains-all", we identified some cardiac muscle components, such as myosin heavy and light chains, actin, tropomyosin, and troponin C, but additional work is required to identify the remaining spots. The two-dimensional electrophoretic system described here makes possible the effective resolution of whole cardiac muscle homogenate from small samples, and looks promising as an aid to muscle research.     

Analysis of electrophoretic patterns of arbitrarily primed PCR profiling

We present a mathematical algorithm for the analysis of electrophoretic patterns resulting from arbitrarily primed PCR profiling. The algorithm is based on the established mathematical procedures applied to the analysis of digital images of gel patterns. The algorithm includes (a) transformation of the image into a matrix form, (b) identification of every electrophoretic lane as a set of matrix columns that are further mathematically processed, (c) averaging of matrix columns corresponding to electrophoretic lanes that define lane representatives, (d) elimination of "smiling" bands, (e) solving the problem of a lane offset, and (f) removal of the background. Representation of individual electrophoretic lanes in the form of functions allows interlane comparisons and further mathematical analysis. Direct comparison of selected lanes was obtained by employing correlation analysis. Gel images were those obtained after arbitrarily primed PCR analysis of DNA that underwent damage induced by gamma radiation from a (60)Co source. The applied method proved to be useful for elimination of subjectivity of visual inspection. It offers the possibility to avoid overlooking important differences in case of suboptimal electrophoretic resolution. In addition, higher precision is achieved in the assessment of quantitative differences due to better insight into experimental artifacts. These simple mathematical methods offer an open-type algorithm, i.e., this algorithm enables easy implementation of different parameters that may be useful for other analytical needs.     

Analysis of ion-association reaction in aqueous solution and its utilization by capillary zone electrophoresis.

Electrophoretic migration of analytes in capillary zone electrophoresis (CZE) reflects the dissolved status of analytes in solution, and the electrophoretic mobility is controlled to develop the resolution among analytes by adding a "modifier" to the migrating solution. Such addition of modifier is essentially the utilization of molecular interactions. Precise measurement of electrophoretic mobility by CZE allows analyzing molecular interactions, and CZE apparatus is very useful for physicochemical measurements. This review focuses on the advantages on using CZE to analyze equilibrium reaction; the capillary electrophoretic method and mathematical analyses that apply acid dissociation and complex formation reactions are also validated. Ion association reactions are deeply related to analytical chemistry and separation science, and CZE has been used for the investigation of ion-ion interactions. Various types of interactions have been clarified through the CZE measurements: contributions of hydrophobicity, probability, and aromatic-aromatic interaction were quantitatively evaluated. Ion association reaction in aqueous solution also elucidates the stepwise reactions of liquid-liquid distribution of ion associates. Development and applications of ion association reaction in CZE analysis are also introduced.     

High performance electrophoresis: A new method of continuous elution of electrophoretically separated high and low molecular weight compounds

Conclusions The purification of serum proteins is a precise and suitable test for the performance of an electrophoresis apparatus. The molecular weights and the isoelectric points of the various serum proteins cover a wide range. Consequently, the protein pattern observed after gel electrophoresis is very complex. Moreover, elution of the electrophoretically purified serum proteins is particularly complicated because some proteins such as albumin and transferrin appear as highly concentrated overloaded bands. If the elution method employed does not work extremely efficiently, residual quantities of albumin and transferrin, respectively, remain in the elution chamber and contaminate the following, slower protein zones. As Fig. 2 shows, no tailing of proteins can be observed in the pure fractions of this electrophoretic run, despite their high concentrations. Even in the case of proteins of low electrophoretic mobility in the late fractions, such as haptoglobins and immunoglobins, no dilution occurred.From these results it appears that the electrophoretic methods described provide a practical and highly efficient way of separating and analyzing high and low molecular weight ampholytes. Thus, technical aspects should no longer be the factor limiting realization of High Performance Electrophoresis.

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HPE: Eine neue Methode der kontinuierlichen Elution elektrophoretisch getrennter hoch- und niedrigmolekularer Verbindungen

     

The electrophoresis of a spherical particle

The problem of predicting the electrophoretic velocity of a spherical charged particle in an infinite dielectric liquid containing impurities is re-examined. A model, somewhat different from those commonly used, is postulated, and the associated equations are formulated and are solved subject to certain approximations. The electrophoretic velocityU is found to be linear in the applied field and to vary algebraically in the parameterχ=ka.     

Electrophoretic mobility study on ion-ion interactions in an aqueous solution.

Ion-ion interactions between anions and their pairing ions in aqueous solutions were studied through the measurements of electrophoretic mobilities of analyte ions in capillary zone electrophoresis, where the electrophoretic method for the analysis of ion association reaction is shown to be more useful than the conductometric method widely used in the analysis of the reactions. The electrophoretic mobility of monovalent inorganic anions was almost identical even when the concentrations of alkali metal ions and quaternary ammonium ions in the migrating solution were varied up to 15 mM. On the other hand, the electrophoretic mobility of organic anions, such as monovalent and divalent anions, decreased with increasing concentrations of quaternary ammonium ions. Changes in the electrophoretic mobilities were analyzed by a non-linear least-squares method giving ion association constants. The results indicate that the proposed method is applicable to the analysis of such reactions to give the mobility change. The ion association constants obtained in an aqueous solution were related to the extraction constants of the ion associates, and the contributions of the association process and the distribution process were clarified.     

Suppression of electroosmotic flow and its application to determination of electrophoretic mobilities in a poly(vinylpyrrolidone)-coated capillary

A hydrophilic polymer, poly(vinylpyrrolidone) (PVP), was employed for suppressing the electroosmotic flow (EOF). A capillary was filled with aqueous PVP solution for coating the capillary wall with PVP; the PVP solution was then replaced by a migration buffer solution containing no PVP. Three types of PVP with different molecular weights were examined. The EOF was suppressed more effectively as the molecular weight of PVP increased. The EOF in the coated capillary was approximately 10-fold smaller than that of a bare capillary and was constant in the pH range of 6-8. The suppressed EOF was stable even when no PVP was added to the migration buffer. However, the EOF increased significantly when sodium dodecyl sulfate was added into the migration buffer. The method was applied for determining the electrophoretic mobilities of inorganic anions that have negative electrophoretic mobilities larger than the electroosmotic mobility of the bare capillary. A novel method for determining the electrophoretic mobilities was proposed based on the linear relationship between electric current and electrophoretic mobility. The electrophoretic mobility was proportional to the electric current. Therefore, the intercept of the regression equation represents the electrophoretic mobility at room temperature. The electrophoretic mobilities were in good agreement with the absolute electrophoretic mobilities.     

Effect of the Surface Conductance on Electrokinetic Potential Measured in Nonaqueous Media

The effect of the surface conductance on the -potential of dispersed particles determined from their electrophoretic mobility in nonaqueous electrolyte solutions was considered. The conductivity of dilute quartz suspensions and the electrophoretic mobility of quartz particles in NaBr solutions in butanol-1 and dimethyl sulfoxide as well as in LiBr solutions in acetone at the salt concentration C= 10^–5–10^–2M were determined by conductometry and microelectrophoresis. The dependences of surface conductivity and -potential on the electrolyte content were calculated by formulas of the Wagner and Henry theories. It was shown that, in the region of dilute solutions, (log C) curves thus obtained significantly differ from corresponding functions calculated by the Smoluchowski equation. At the same time, these dependences agreed closely with the (log C) dependences determined for the same systems by the streaming potential method with allowances for experimental values of the surface conductivity. Using aluminum oxide suspensions in NaBr and HBr ethanol solutions as examples, it was shown that, to obtain correct values of the -potential from electrophoretic mobility of porous particles impregnated with a solution, it is necessary to allow for the bulk conductance of the particles.     

Numerical analysis of sodium dodecyl sulphate-polyacrylamide gel electrophoretic protein patterns for the classification, identification and typing of medically important bacteria

A numerical analysis of one-dimensional sodium dodecyl sulphate polyacrylamide gel electrophoretic protein patterns of three separate bacterial groups is described. Similarity between patterns was estimated using a correlation coefficient and is represented diagramatically by the unweighted pair-group with arithmetic averages method of clustering. Protein patterns were scanned using a laser densitometer interfaced directly to a microcomputer, where calculations were performed semi-automatically with specially written software. A method for the normalization of patterns from different gels is utilized. Whole cell protein patterns clearly distinguished strains representing various groups of Achromobacter. Similarly the two biovars of the Group EF-4 bacteria could be separated after analysis of partial "background patterns" as could the different species of Providencia. In addition, whole patterns could be used to type the various strains of P. rettgeri. These examples serve to illustrate the power, versatility and potential of the technique when applied to problems of classification, identification and typing of bacteria of medical interest. Its advantages and disadvantages, when compared to other available electrophoretic methodologies, are discussed, as is the future application of such technology to the hospital laboratory.     

A method for UV-bonding in the fabrication of glass electrophoretic microchips.

This paper presents an approach for the development of methodologies amenable to simple and inexpensive microchip fabrication, potentially applicable to dissimilar materials bonding and chip integration. The method involves a UV-curable glue that can be used for glass microchip fabrication bonding at room temperature. This involves nothing more than fabrication of glue "guide channels" into the microchip architecture that upon exposure to the appropriate UV light source, bonds the etched plate and cover plate together. The microchip performance was verified by capillary zone electrophoresis (CZE) of small fluorescent molecules with no microchannel surface modification carried out, as well as with a DNA fragment separation following surface modification. The performance of these UV-bonded electrophoretic microchips indicates that this method may provide an alternative to high temperature bonding.