This work focuses on the development of an online programmable microfluidic bioprocessing unit (BioModule) using digital logic microelectrodes for rapid pipelined selection and transfer of deoxyribonucleic acid (DNA) molecules and other charged biopolymers. The design and construction technique for this hybrid programmable biopolymer processing device is presented along with the first proof of principle functionality. The electronically controlled collection, separation and channel transfer of the biomolecules is monitored by a sensitive fluorescence set-up. This hybrid reconfigurable architecture couples electronic and biomolecular information processing via a single module combination of fluidics and electronics and opens new fields of applications not only in DNA computing and molecular diagnostics but also in applications of combinatorial chemistry and lab-on-a-chip biotechnology to the drug discovery process. Fundamentals of the design and silicon-polydimethylsiloxane (PDMS)-based construction of these electronic microfluidic devices and their functions are described as well as the experimental results.
Spatially resolved single molecule detection in solution is a prerequisite for molecule tracking and provides new opportunities in single molecule manipulation, for example in a sample flow. A detector with fast timing properties and high spatial resolution is required for these purposes. We introduce a concept for spatially resolved optical single molecule detection in an epi‐illuminated microscope using a novel kind of detector and a new algorithm in configurable hardware for intelligent data processing. The analysis is performed on a parallel hardware interface and includes burst detection. It can be extended to on‐line spatial and temporal data analysis. This detector will be used in combination with a molecular sorter where molecules are sorted in a microstructured flow device made of silicon. In a first step towards such sorting in these microstructures, we show that a reliable detection of single molecules in silicon microstructures is possible in zero‐dimensional detection volumes. The noise structure of the data is analysed in terms of Poissonian statistics and it is shown that in the smallest structure used (depth 20 μm) a signal‐to‐noise ratio of 40 is achieved.
Objective Early diagnosis of atrial fibrillation (AFib) is a priority for stroke prevention. We sought to test four commercial pulse detection systems (CPDSs) for ability to distinguish AFib from normal sinus rhythm using a published algorithm (Zhou et al., PLoS One 2015;10:e0136544), compared with visual diagnosis by electrocardiogram inspection. Methods BAYathlon was a prospective, non-interventional, single-centre study. Adult cardiology patients with documented AFib or sinus rhythm who were due to have a routine 5-min electrocardiogram were randomized to undergo a parallel 5-min pulse assessment with a Polar V800, eMotion Faros 360, TomTom heart rate monitor, or Adidas miCoach Smart Run. Results 144 patients (73 with AFib, 71 with sinus rhythm (based on electrocardiograms); median age: 73 years; 53.5% male) were analysed. Algorithm sensitivities (primary endpoint) and specificities for AFib when applied to CPDS recordings were 93.3% and 94.1% with the Polar V800, 90.0% and 84.2% with the eMotion Faros 360, and 0% and 100% with the other CPDSs (analysis period: 127 heart rate signals + 2 min). When applied to routine electrocardiograms, the algorithm correctly detected AFib in 71/73 patients. Different analysis periods (127 heart rate signals +1 or 3 min) only slightly changed the sensitivities with the Polar V800 and eMotion Faros 360 and had no effect on the sensitivities with the other CPDSs. Conclusion AFib screening using the applied algorithm is feasible with the Polar V800 and eMotion Faros 360 (which provide RR interval data) but not with the other CPDSs (which provide pre-processed heart rate time series). ClinicalTrials.gov identifier: NCT02875106
Determining the concentration of nucleic acids in biological samples precisely and reliably still is a challenge. In particular when only very small sample quantities are available for analysis, the established fluorescence-based methods give insufficient results. Photobleaching is seen as the main reason for this. In this paper we present a method to correct for the photobleaching effect. Using confocal microscopy with single molecule sensitivity, we derived calibration curves from DNA solutions with defined fragment length. We analyzed dilution series over a wide range of concentrations (1 pg/μl-1000 pg/μl) and measured their specific diffusion coefficients employing fluorescence correlation spectroscopy. Using this information, we corrected the measured fluorescence intensity of the calibration solutions for photobleaching effects. We evaluated our method by analyzing a series of DNA mixtures of varying composition. For fragments smaller than 1000 bp, our method allows to determine sample concentrations with high precision in very small sample quantities (< 2 μl with concentrations < 20 pg/μl). Once the technical parameters are determined and remain stable in an established process, our improved calibration method will make measuring molecular biological samples of unknown sequence composition more efficient, accurate and sample-saving than previous methods.
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