Abstract
A low cost thin-film transistor (TFT) nanoribbon (NR) sensor has been developed for rapid real-time detection of DNA amplification using an isothermal Recombinase Polymerase Amplification (RPA) method. The semiconductor chip measures DNA amplification through a pH change, rather than via fluorescence. The utility of the method was demonstrated by amplifying CTX-M and NDM, two genes that confer bacterial resistance to cephalosporins and carbapenems, respectively. It is shown that this approach provides extremely fast and sensitive detection. It can detect <10 copies of the gene in genomic DNA extracted from E. coli or K. pneumoniae clinical isolates within a few minutes. A differential readout system was developed to minimize the effect of primer-dimer amplification on the assay. The simple device has the potential for low cost, portable and real-time nucleic acid analysis as a Point of Care device.
Highlights
There is a drive to develop fast and simple diagnostic tools to aid in the identification and diagnosis of a range of diseases
Amplification is usually monitored in real time using fluorescent exo-probes that are complementary to the amplified DNA fragment, see Fig. 2b
This work demonstrates a novel method for real-time on-chip isothermal DNA amplification using a low cost thin-film transistor (TFT) NR sensor with high speed and high sensitivity
Summary
There is a drive to develop fast and simple diagnostic tools to aid in the identification and diagnosis of a range of diseases. Antimicrobial resistance (AMR) is rapidly becoming a global threat as pathogens evolve new resistance strategies, potentially making even simple infections difficult to treat (O'Neill, 2016). NDM-1 is a gene which mediates resistance to carbapenems, one of the last drug types which is effective against multidrug resistant Gram-negative bacteria. This resistance gene was originally identified in 2008 and has spread widely around the world, facilitated by its presence on a highly promiscuous plasmid which can be moved from one bacterial species to another. Given the rapid dissemination of resistance genes and the emergence of strains which are commonly multidrug resistant, there is a clear need to develop rapid and sensitive tests to determine antibiotic resistance, both as part of the diagnosis and management of infection and to provide a rational basis for the appropriate prescription of antibiotics
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