On-chip electrical detection of parallel loop-mediated isothermal amplification with DG-BioFETs for the detection of foodborne bacterial pathogens
| Autor: | Yi Shao Liu, Bobby Reddy, Carlos Duarte-Guevara, Jui Cheng Huang, Vikhram V. Swaminathan, Rashid Bashir |
|---|---|
| Rok vydání: | 2016 |
| Předmět: |
Detection limit
Nucleic acid quantitation business.industry Chemistry General Chemical Engineering 010401 analytical chemistry Transistor Loop-mediated isothermal amplification Nanotechnology 02 engineering and technology General Chemistry 021001 nanoscience & nanotechnology 01 natural sciences Reference electrode 0104 chemical sciences law.invention law Redundancy (engineering) Optoelectronics Field-effect transistor Electronics 0210 nano-technology business |
| Zdroj: | RSC Advances. 6:103872-103887 |
| ISSN: | 2046-2069 |
| DOI: | 10.1039/c6ra19685c |
| Popis: | The use of field effect transistors (FETs) as the transduction element for the detection of DNA amplification reactions will enable portable and inexpensive nucleic acid analysis. Transistors used as biological sensors, or BioFETs, minimize the cost and size of detection platforms by leveraging fabrication methods already well developed for electronics. Here, we report a dual-gate BioFET (DG-BioFET) array platform with 1024×1024 sensors that is used for on-chip electrical detection of loop-mediated isothermal amplification (LAMP) reactions that target food borne bacterial pathogens. The DG-BioFETs of our 7 × 7 mm2 array are able to electrically detect pH changes that are triggered by nucleotide incorporation during LAMP elongation. Multiple 250 nL reactions can be simultaneously electrically monitored in our array that is divided in 30 micro-chambers with gold-coated anisotropically etched silicon wells that act both as reference electrode and confinement element. Our characterization results show that the gold-biased DG-BioFETs have a sensitivity of 32 mV pH−1 (equivalent to 2 μA pH−1) and an average resolution of 0.5 pH units. This sensitivity is high enough to detect the pH changes triggered by the amplification reaction, but to maximize our signal-to-noise ratio and improve our quantitative conclusions we use a group of data analysis techniques that are available in our high-density platform that monitors each reaction with ∼3500 independent BioFETs. We use redundancy techniques to minimize the overall standard deviation of our measurements, the Grubbs test to eliminate measurements outside the expected normal distribution, and reference micro-chambers to subtract the common noise. With these techniques we are capable of reducing the P value, of a t-test comparing positive and negative readings, from a typical 0.17 to 0.03. The platform that we present along with the analysis techniques that we developed allow the on-chip electrical detection and identification of E. coli O157 and S. typhi with parallel LAMP assays targeting eae and invA genes. The LAMP reactions are highly specific, without false positives, and our titration assays demonstrate a limit of detection of 23 CFU per reaction on chip. |
| Databáze: | OpenAIRE |
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