Automated microdroplet platform for sample manipulation and polymerase chain reaction

We present a fully automated system performing continuous sampling, reagent mixing, and polymerase chain reaction (PCR) in microdroplets transported in immiscible oil. Sample preparation and analysis are totally automated, using an original injection method from a modified 96-well plate layered with three superimposed liquid layers and in-capillary laser-induced fluorescence endpoint detection. The process is continuous, allowing sample droplets to be carried uninterruptedly into the reaction zone while new drops are aspirated from the sample plate. Reproducible amplification, negligible cross-contamination, and detection of low sample concentrations were demonstrated on numerous consecutive sample drops. The system, which opens the route to strong reagents and labor savings in high-throughput applications, was validated on the clinically relevant quantification of progesterone receptor gene expression in human breast cancer cell lines.     

The Desorption of a Volatile Corrosion Inhibitor from Polyethylene

The kinetics of the desorption of a volatile inhibitor of atmospheric corrosion, hexamethyleneimine m-nitrobenzoate, from polyethylene film at 298–333 K was examined. The dependence of the desorption rate on concentration and the character of the inhibitor's distribution in the film was revealed. Effective diffusion coefficients of the inhibitor in polyethylene, as well as activation energy of the diffusion process, were calculated. Recommendations on the practical application of the inhibited film were offered.     

Measurement of the surface concentration for bioassay kinetics in microchannels

We present a simple and versatile method, based on fluorescence microscopy, to reliably measure the concentration of advected molecules in the vicinity of surfaces in microchannels. This tool is relevant to many microfluidic applications such as immunoassays and single-molecule experiments, where one probes the kinetics of a reaction between an immobilized target and a reactant carried by the bulk flow. The characterization of the surface concentration highlights the dominant role of transverse diffusion, which generates an apparent diffusivity at the surface 3-4 orders of magnitude greater than molecular diffusion alone, even close to the point of injection. We directly measure the effects of the longitudinal position along the channel and of the flow rate on the concentration front and develop a simple analytical model that compares well with the data. Finally, we propose a method to properly account for concentration fronts in single-molecule measurements and use it to directly access the kinetics parameters of protamine-induced condensation of DNA.     

Degeneracy and discrete receiver operating characteristic rating data

RATIONALE AND OBJECTIVES: Observer performance studies sometimes use too few cases for estimating diagnostic accuracy from binormal receiver operating characteristic (ROC) curves. One important problem is degenerate data sets. We compared a new algorithm, RSCORE4, with the exact-solution approach to degeneracy in ROCFIT and with the Wilcoxon statistic. METHODS: Degenerate ROC solutions result from empty cells in the data matrix. We addressed this problem by adding a small constant to empty cells in a maximum-likelihood program, RSCORE4. When this method failed, the program branched to a pattern-search algorithm. We tested the program in a series of Monte Carlo studies. RESULTS: RSCORE4 converged to nondegenerate solutions in every case and gave results closer to population values than ROCFIT or Wilcoxon. ROCFIT converged to exact-fit degenerate solutions, those with zero or infinite parameter values, in more than 40% of the samples. The Wilcoxon statistic was biased. CONCLUSION: RSCORE4 seems to outperform other currently recommended methods for dealing with degeneracy.     

The exergy method of calculating the optimum parameters of multistage evaporators

Translated from Khimicheskoe i Neftyanoe Mashinostroenie, No. 5, pp. 1–2, May, 1991.     

Automatic Decomposition of the Clinical Electromyogram

We describe a new, automatic signal-processing method (ADEMG) for extracting motor-unit action potentials (MUAP's) from the electromyographic interference pattern for clinical diagnostic purposes. The method employs digital filtering to select the spike components of the MUAP's from the background activity, identifies the spikes by template matching, averages the MUAP waveforms from the raw signal using the identified spikes as triggers, and measures their amplitudes, durations, rise rates, numbers of phases, and firing rates. Efficient new algorithms are used to align and compare spikes and to eliminate interference from the MUAP averages. In a typical 10-s signal recorded from the biceps brachii muscle using a needle electrode during a 20 percent-maximal isometric contraction, the method identifies 8-15 simultaneously active MUAP's and detects 30-70 percent of their occurrences. The analysis time is 90 s on a PDP-11/34A.     

Coupled flow and reaction during natural convection PCR

Polymerase chain reaction (PCR) in a microfluidic Rayleigh–Benard convection cell represents a promising route towards portable PCR for point-of-care uses. In the present contribution, the coupled fluid mechanics and heat transport processes are solved numerically for a 2-D flow cell. The resultant velocity and temperature fields serve as the inputs to a convection-diffusion-reaction model for the DNA amplification, wherein the reaction kinetics are modeled by Gaussian distributions around the conventional bulk PCR reaction temperatures. These evolution equations are integrated to determine the exponential growth rate of the double-stranded DNA concentration. The predicted doubling time is approximately 10–25 s, increasing with the Péclet number. This effect is attributed to low velocity, slow kinetics “dead zones” located at the center of the reactor. The latter observation provides an alternative rationalization for the use of loop-based natural convection PCR systems.