Loop-mediated isothermal amplification-based microfluidic chip for pathogen detection

Shang, Yuting; Sun, Jiadi; Ye, Yongli; Zhang, Jumei; Zhang, Yinzhi; Sun, Xiulan

doi:10.1080/10408398.2018.1518897

Cited by 94 publications

(48 citation statements)

References 116 publications

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“…These advantages allow CMOS image sensors to be used in some of the highest-specification industrial control devices and the finest cameras [23]. CMOS-based biosensors were developed for the detection of bacteria [24,25], water, and air toxicants [26,27], coupled with PCR [28,29], and used in food safety [30][31][32][33][34]. Only a few CMOS-based biosensor platforms were developed for use in agriculture applications [35,36].…”

Section: Introductionmentioning

confidence: 99%

The Incorporation of Amplified Metal-Enhanced Fluorescence in a CMOS-Based Biosensor Increased the Detection Sensitivity of a DNA Marker of the Pathogenic Fungus Colletotrichum gloeosporioides

2020

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Half of the global agricultural fresh produce is lost, mainly because of rots that are caused by various pathogenic fungi. In this study, a complementary metal-oxide-semiconductor (CMOS)-based biosensor was developed, which integrates specific DNA strands that allow the detection of enoyl-CoA-hydratase/isomerase, which is a quiescent marker of Colletotrichum gloeosporioides fungi. The developed biosensor mechanism is based on the metal-enhanced fluorescence (MEF) phenomenon, which is amplified by depositing silver onto a glass surface. A surface DNA strand is then immobilized on the surface, and in the presence of the target mRNA within the sample, the reporter DNA strand that is linked to horseradish peroxidase (HRP) enzyme will also bind to it. The light signal that is later produced from the HRP enzyme and its substrate is enhanced and detected by the coupled CMOS sensor. Several parameters that affect the silver-deposition procedure were examined, including silver solution temperature and volume, heating mode, and the tank material. Moreover, the effect of blocking treatment (skim milk or bovine serum albumin (BSA)) on the silver-layer stability and nonspecific DNA absorption was tested. Most importantly, the effect of the deposition reaction duration on the silver-layer formation and the MEF amplification was also investigated. In the study findings a preferred silver-deposition reaction duration was identified as 5–8 min, which increased the deposition of silver on the glass surface up to 13-times, and also resulted in the amplification of the MEF phenomenon with a maximum light signal of 50 relative light units (RLU). It was found that MEF can be amplified by a customized silver-deposition procedure that results in increased detection sensitivity. The implementation of the improved conditions increased the biosensor sensitivity to 3.3 nM (4500 RLU) with a higher detected light signal as compared to the initial protocol (400 RLU). Moreover, the light signal was amplified 18.75-, 11.11-, 5.5-, 11.25-, and 3.75-times in the improved protocol for all the tested concentrations of the target DNA strand of 1000, 100, 10, 3.3, and 2 nM, respectively. The developed biosensor system may allow the detection of the pathogenic fungus in postharvest produce and determine its pathogenicity state.

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Section: Introductionmentioning

confidence: 99%

The Incorporation of Amplified Metal-Enhanced Fluorescence in a CMOS-Based Biosensor Increased the Detection Sensitivity of a DNA Marker of the Pathogenic Fungus Colletotrichum gloeosporioides

2020

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“…The laboratory-based detection of infectious pathogens has been the basis for the diagnosis of infectious diseases. To date, numerous methods have been proposed for pathogen detection, such as conventional microbiological analysis, nucleic acid-based methods and immunoassays (Shang et al, 2020;Zhou et al, 2014). Conventional microbiological methods are the gold standard in microbe detection (Swaminathan & Feng, 1994).…”

Section: Introductionmentioning

confidence: 99%

Simultaneous detection of multiple bacterial and viral aquatic pathogens using a fluorogenic loop‐mediated isothermal amplification‐based dual‐sample microfluidic chip

Zhou

et al. 2020

Journal of Fish Diseases

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Rapid and user-friendly diagnostic tests are necessary for early diagnosis and immediate detection of diseases, particularly for on-site screening of pathogenic microorganisms in aquaculture. In this study, we developed a dual-sample microfluidic chip integrated with a real-time fluorogenic loop-mediated isothermal amplification assay (dual-sample on-chip LAMP) to simultaneously detect 10 pathogenic microorganisms, that is Aeromonas hydrophila, Edwardsiella tarda, Vibrio harveyi, V. alginolyticus, V. anguillarum, V. parahaemolyticus, V. vulnificus, infectious hypodermal and haematopoietic necrosis virus, infectious spleen and kidney necrosis virus, and white spot syndrome virus. This on-chip LAMP provided a nearly automated protocol that can analyse two samples simultaneously, and the tests achieved limits of detection (LOD) ranging from 10 0 to 10-1 pg/μl for genomic DNA of tested bacteria and 10-4 to 10-5 pg/μl for recombinant plasmid DNA of tested viruses, with run times averaging less than 30 min. The coefficient of variation for the time-to-positive value was less than 10%, reflecting a robust reproducibility. The clinical sensitivity and specificity were 93.52% and 85.53%, respectively, compared to conventional microbiological or clinical methods. The on-chip LAMP assay provides an effective dual-sample and multiple pathogen analysis, and thus would be applicable to on-site detection and routine monitoring of multiple pathogens in aquaculture.

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“…Foodborne pathogens pose important public health concern worldwide because they can contaminate foods through any food chains from farm to fork (Shan et al, 2015;Shang et al, 2018). The Centers for Disease Control and Prevention estimated that 1 in 6 Americans (or 48 million people) each year becomes adversely affected by foodborne diseases (CDC, 2018).…”

Section: Introductionmentioning

confidence: 99%

Fluorescence immunoassay through histone-ds-poly(AT)-templated copper nanoparticles as signal transductors for the sensitive detection of Salmonella choleraesuis in milk

Xiong

Zhang

Gao

et al. 2019

Journal of Dairy Science

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The rapid and sensitive detection of foodborne pathogens is one of the most important issues in food safety control. In this work, we developed a novel fluorescence immunoassay method for the sensitive detection of Salmonella choleraesuis. The method uses the fluorescent signals of histone-ds-poly(AT)-templated copper nanoparticles (His-pAT CuNP) as signal transducers and glucose oxidase as an alternative for horseradish peroxidase for the generation of hydrogen peroxide (H 2 O 2 ) through the catalysis of glucose. The H 2 O 2 is then further converted into hydroxyl radical (•OH) by Fenton reagents. Owing to the ultrahigh sensitivity of His-pAT CuNP synthesis toward •OH, the proposed fluorescence immunoassay method exhibited excellent sensitivity for S. choleraesuis, with a limit of detection of 8.04 × 10 1 cfu/mL, which is 3 orders of magnitude lower than that of the tetramethylbenzidine-based traditional immunoassay. The reliability of the proposed method was evaluated by using spiked milk samples with S. choleraesuis concentration ranging from 8.8 × 10 1 to 8.8 × 10 4 cfu/mL. The average recoveries for the intra-and inter-assay ranged from 73.52 to 96.59% and from 66.99 to 98.24% with a coefficient of variation from 6.85 to 31.26% and 5.46 to 17.99%, respectively. These results indicated that the proposed fluorescence immunoassay possesses a great potential for ultra-sensitive detection of foodborne pathogens in food safety control.

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Loop-mediated isothermal amplification-based microfluidic chip for pathogen detection

Cited by 94 publications

References 116 publications

The Incorporation of Amplified Metal-Enhanced Fluorescence in a CMOS-Based Biosensor Increased the Detection Sensitivity of a DNA Marker of the Pathogenic Fungus Colletotrichum gloeosporioides

The Incorporation of Amplified Metal-Enhanced Fluorescence in a CMOS-Based Biosensor Increased the Detection Sensitivity of a DNA Marker of the Pathogenic Fungus Colletotrichum gloeosporioides

Simultaneous detection of multiple bacterial and viral aquatic pathogens using a fluorogenic loop‐mediated isothermal amplification‐based dual‐sample microfluidic chip

Fluorescence immunoassay through histone-ds-poly(AT)-templated copper nanoparticles as signal transductors for the sensitive detection of Salmonella choleraesuis in milk

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