An optimized microchip electrophoresis system for mutation detection by tandem SSCP and heteroduplex analysis for p53 gene exons 5-9

With the complete sequencing of the human genome, there is a growing need for rapid, highly sensitive genetic mutation detection methods suitable for clinical implementation. DNA-based diagnostics such as single-strand conformational polymorphism (SSCP) and heteroduplex analysis (HA) are commonly used in research laboratories to screen for mutations, but the slab gel electrophoresis (SGE) format is ill-suited for routine clinical use. The translation of these assays from SGE to microfluidic chips offers significant speed, cost, and sensitivity advantages; however, numerous parameters must be optimized to provide highly sensitive mutation detection. Here we present a methodical study of system parameters including polymer matrix, wall coating, analysis temperature, and electric field strengths on the effectiveness of mutation detection by tandem SSCP/HA for DNA samples from exons 5-9 of the p53 gene. The effects of polymer matrix concentration and average molar mass were studied for linear polyacrylamide (LPA) solutions. We determined that a matrix of 8% w/v 600 kDa LPA provides the most reliable SSCP/HA mutation detection on chips. The inclusion of a small amount of the dynamic wall-coating polymer poly-N-hydroxyethylacrylamide in the matrix substantially improves the resolution of SSCP conformers and extends the coating lifetime. We investigated electrophoresis temperatures between 17 and 35 degrees C and found that the lowest temperature accessible on our chip electrophoresis system gives the best condition for high sensitivity of the tandem SSCP/HA method, especially for the SSCP conformers. Finally, the use of electrical fields between 350 and 450 V/cm provided rapid separations (<10 min) with well-resolved DNA peaks for both SSCP and HA.     

Driving matrix liquid crystal displays

Liquid crystal displays had a humble beginning with wrist watches in the seventies. Continued research and development in this multi-disciplinary field have resulted in displays with increased size and complexity. After three decades of growth in performance, LCDs now offer a formidable challenge to the cathode ray tubes (CRT). A major contribution to the growth of LCD technology has come from the developments in addressing techniques used for driving matrix LCDs. There are several approaches like passive matrix addressing, active matrix addressing and plasma addressing to drive a matrix display. Passive matrix LCD has a simple construction and uses the intrinsic non-linear characteristic of the LCD for driving. Departure from conventional line by line addressing of a passive matrix has resulted in improved performance of the display. Orthogonal functions have played a crucial role in the development of passive matrix addressing. Simple orthogonal functions that are useful for driving a matrix LCD are introduced. The basics of driving several rows simultaneously (multi-line addressing) are discussed by drawing analogies from multiplexing in communication. The impact of multi-line addressing techniques on the performance of the passive matrix LCDs in comparison with the conventional technique will be discussed.     

On the governing equations for the compressing process and its coupling with other processes

As a continuation of a recent linear analysis by Mao et al.(Acta Mech Sin,2010,26:355),in this paper we propose a general theoretical formulation for the compressing process in complex Newtonian fluid flows,which covers gas dynamics,aeroacoustics,nonlinear thermoviscous acoustics,viscous shock layer,etc.,as its special branches.The principle on which our formulation is based is the maximally natural and dynamic Helmholtz decomposition of the Navier-Stokes equation,along with the kinematic Helmholtz decompos...     

Fluorescence energy transfer-labeled primers for high-performance forensic DNA profiling

A fluorescence energy transfer (ET) dye-labeled STR typing system (ET 16-plex) is developed for the markers used in the commercial STR typing kit PowerPlex 16, and its performance assessed using a 96-lane microfabricated capillary array electrophoresis (muCAE) system. The ET 16-plex amplicons displayed 1.6-9-fold higher fluorescence intensities compared to those produced using the single-dye (SD)-labeled multiplex kits. The ET multiplex delivered full STR profiles from 62.5 pg of DNA; half the input required for the SD kits while maintaining a similar heterozygote allele balance. This increased sensitivity should improve typing of poor-quality DNA samples by making minor or imbalanced alleles more readily detectable at the low copy number (LCN) threshold. The ET 16-plex also generated complete profiles with only 28 PCR cycles; this capability should improve LCN typing by reducing the amplification time and drop-in allele incidence. To confirm the practical advantages of ET-labeled primers, six previously problematic casework samples were tested and only the ET 16-plex kit was able to capture additional allele data. The successful development and demonstration of ET primers for higher sensitivity STR typing offers a simple solution to improving current commercial multiplex typing capability. The superior spectral properties and universal compatibility with any primer sequence provided by ET cassettes will make future multiplex construction more facile and straightforward. The pairing of ET cassette technology with the muCAE system illustrates not only an enhanced STR typing platform, but a significant step toward a higher-efficiency forensic laboratory enabled by better chemistry and microfluidics.     

A bio-barcode assay for on-chip attomolar-sensitivity protein detection

Functionalized nanoparticles hold great promise in realizing highly sensitive and selective biodetection. We report a single disposable chip which is capable of carrying out a multi-step process that employs nanoparticles--a bio-barcode assay (BCA) for single protein marker detection. To illustrate the capability of the system, we tested for the presence of prostate specific antigen (PSA) in buffer solution and goat serum. Detection was accomplished at PSA concentrations as low as 500 aM. This corresponds to only 300 copies of protein analytes using 1 microL total sample volume. We established that the on-chip BCA for PSA detection offers four orders of magnitude higher sensitivity compared to commercially available ELISA-based PSA tests.