A microfluidic biosensor based on nucleic acid sequence recognition

Abstract

The development of a generic semi-disposable microfluidic biosensor for the highly sensitive detection of pathogens via their nucleic acid sequences is presented in this paper. Disposable microchannels with defined areas for capture and detection of target pathogen RNA sequence were created in polydimethylsiloxane (PDMS) and mounted onto a reusable polymethylmethacrylate (PMMA) stand. Two different DNA probes complementary to unique sequences on the target pathogen RNA serve as the biorecognition elements. For signal generation and amplification, one probe is coupled to dye encapsulated liposomes while the second probe is coupled to superparamagnetic beads for target immobilization. The probes hybridize to target RNA and the liposome-target-bead complex is subsequently captured on a magnet. The amount of liposomes captured correlates directly to the concentration of target sequence and is quantified using a fluorescence microscope. Dengue fever virus serotype 3 sequences and probes were used as a model analyte system to test the sensor. Probe binding and target capture conditions were optimized for sensitivity resulting in a detection limit of as little as 10 amol microL(-1) (10 pmol L(-1)). Future biosensors will be designed to incorporate a mixer and substitute the fluorescence detection with an electrochemical detection technique to provide a truly portable microbiosensor system.

Microfluidic biosensor for cholera toxin detection in fecal samples

“…The microchannels with 50-μm depths were produced by standard photolithography and dry etching techniques as described previously [17,19,20]. With use of soft lithography, the microchannels were realized in PDMS covered with a glass slide and a glass slide bearing the IDUA for the fluorescence and electrochemical formats, respectively.…”

The Application of Nanotechnology in Medical Sciences: New Horizon of Treatment

“…They made up of a biological factor responsible for sampling, as well as a physical element (often called transducer) transmitting sampling outcome for moreover processing [5][6][7][8]. The biological element of a biosensor contains a biosensitive layer, which can either contain bioreceptors or be made of bioreceptors covalently attached to the transducer.Concerning the biological specificity conferring mechanism used one can distinguish 5 major categories of biosensors 1) Antibody/antigen based, 2) Enzymes based (mono-or multi enzyme systems) [9], 3) Nucleic acids based (DNA, cDNA, RNA) [10] 4) Based on cellular interactions (cellular structures/cells),utilisating the whole cells (microorganisms, such asbacteria, fungi, eukaryotic cells or yeast) or cell organellesor particles (mitochondria, cell walls, tissue slices) [11][12][13][14][15][16] 5) Employing biomimetic materials (e.g., synthetic bioreceptors). Biosensors based on nucleic acid interactions of biological element are called DNA biosensors, or genosensors, or biodetectors.…”

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“…Other applications of fluorescence biosensing are based on the recognition of target nucleic acid sequence. Dengue fever virus serotype 3 DNA probes 32 were detected in concentration of 10 pM. Photometric biosensors can also be based on enzyme-labeled antibodies.…”