Simultaneous Detection of Antibiotic Resistance Genes on Paper-Based Chip Using [Ru(phen)₂dppz]²⁺ Turn-on Fluorescence Probe

Abstract

Antibiotic resistance, the ability of some bacteria to resist antibiotic drugs, has been a major global health burden due to the extensive use of antibiotic agents. Antibiotic resistance is encoded via particular genes; hence the specific detection of these genes is necessary for diagnosis and treatment of antibiotic resistant cases. Conventional methods for monitoring antibiotic resistance genes require the sample to be transported to a central laboratory for tedious and sophisticated tests, which is grueling and time-consuming. We developed a paper-based chip, integrated with loop-mediated isothermal amplification (LAMP) and the "light switch" molecule [Ru(phen)dppz], to conduct turn-on fluorescent detection of antibiotic resistance genes. In this assay, the amplification reagents can be embedded into test spots of the chip in advance, thus simplifying the detection procedure. [Ru(phen)dppz] was applied to intercalate into amplicons for product analysis, enabling this assay to be operated in a wash-free format. The paper-based detection device exhibited a limit of detection (LOD) as few as 100 copies for antibiotic resistance genes. Meanwhile, it could detect antibiotic resistance genes from various bacteria. Noticeably, the approach can be applied to other genes besides antibiotic resistance genes by simply changing the LAMP primers. Therefore, this paper-based chip has the potential for point-of-care (POC) applications to detect various gene samples, especially in resource-limited conditions.

Fabrication, Flow Control, and Applications of Microfluidic Paper-Based Analytical Devices

“…A fluorescence microscope with filter was used to measure the fluorescence of each fold of paper [198]. Recent advances in various detection methods in both medical and environmental applications include metal ions [199,200], fluoride ions [201], phenolic pollutants (Figure 8C) [202], antibiotic resistance genes (Figure 8D) [203], drug analysis of adrafinil [204], immunoassay [205], and visualization of cancer cells [206].…”

“…Copyright 2018 American Chemical Society). ( D ) Antibiotic resistance gene detection via fluorescence sensing using a light source (reprinted with permission from [203]. Copyright 2018 American Chemical Society).…”

Recent Advances in Electrochemiluminescence Sensors for Pathogenic Bacteria Detection

“…Due to the advantages of simplicity, ability of capillary action and sample storage, many paper-based colorimetric analysis platforms have been built to detect nucleic acid, protein, etc. [95,96]. Benefiting from the advantages of malleability, foldability, as well as the ability to filter the sample, several paper-based platforms integrate purification of sample, nucleic acid amplification and detection [97,98,99].…”

Challenges in paper-based fluorogenic optical sensing with smartphones

“…In most paper-based point-of-care assays that utilize fluorescence, pixel intensities are extracted and converted into a concentration of a target of interest, i.e., molecule [ 38 – 40 ], protein [ 23 , 41 ], whole organism [ 42 ], or nucleic acid [ 43 , 44 ]. When trying to control and maintain constant lighting, on-board default camera settings on smartphones can be problematic, as they are constantly trying to adjust white balance, focus and exposure during multiple assays.…”