Development of Chemiluminescent Lateral Flow Assay for the Detection of Nucleic Acids

Wang, Yuhong; Fill, Catherine; Nugen, Sam R.

doi:10.3390/bios2010032

Cited by 27 publications

(21 citation statements)

References 28 publications

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“…The estimated detection limit of the electrochemical detection for Salmonella in1× PBS buffer and agricultural water was 7.6×10 2 and 6.0 × 10 2 CFU/mL, respectively. The limit of detection (LOD) was estimated by the comparison of the mean value of negative control plus three standard deviations and the corresponding Salmonella concentration (Kessler, 1998;Kjems, et al, 1993;Y. Wang, Fill, & Nugen, 2012).…”

Section: Electrochemical Detection Of Salmonellamentioning

confidence: 99%

Rapid detection of Salmonella using a redox cycling-based electrochemical method

Wang

Chen

et al. 2016

Food Control

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An electrochemical method based on redox cycling combined with immunomagnetic separation and pre-concentration was developed for rapid and sensitive detection of Salmonella. Electrochemical methods for the detection of bacteria offer the advantages of instant quantification with minimal equipment. Unfortunately, the limits of detection are often poor compare to other transduction methods such as fluorescence and chemiliuminescence. We demonstrated an electrochemical method which is both rapid and has a low limit of detection. A two-step strategy, which included immunomagentic pre-concentration and redox cycling was used to amplify the signal. Magnetic beads modified with anti-Salmonella antibodies were used for separation and pre-concentration of Salmonella from phosphate buffered saline (PBS) and agricultural water. Then anti-Salmonella antibodies conjugated with alkaline phosphatase were employed for labeling the Salmonella which had been captured by magnetic beads. Alkaline phosphatase (ALP) catalyzed the substrate Lascorbic acid 2-phosphate (AAP) to electroactive species L-ascorbic acid (AA) while tris(2-carboxyethyl)phosphine (TCEP) facilitated the regeneration of AA on the gold electrode to form redox cycling resulting in an amplified signal. Under the optimal conditions, the Salmonella in PBS buffer as well as in agricultural water were detected. The limit of detection of this approach was approximately 7.6 × 10 2 CFU/mL and 6.0 × 10 2 CFU/mL in PBS buffer and agricultural water, respectively, without pre-enrichment in 3 hours. When the agricultural water has been pre-enriched for 4 hours, the limit of detection was approximately 10 CFU/mL.

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Section: Electrochemical Detection Of Salmonellamentioning

confidence: 99%

Rapid detection of Salmonella using a redox cycling-based electrochemical method

Wang

Chen

et al. 2016

Food Control

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“…Molecular diagnosis on a lateral/vertical flow system has been described for the detection of other viral and parasitic infections (41,42). However, these assays lacked an integrated nucleic acid extraction protocol and have yet to demonstrate clinical efficacy in practical settings.…”

Section: Discussionmentioning

confidence: 99%

Duplex Molecular Assay Intended for Point-of-Care Diagnosis of Influenza A/B Virus Infection

Thomas

Curran

et al. 2013

J Clin Microbiol

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Early diagnosis and management of influenza virus infection directly correlates with the effectiveness in disease control. Current molecular influenza virus tests were designed for use in diagnostic testing facilities, where sophisticated equipment and highly trained technicians are available. A longer turnaround time for the centralized testing than when testing near the sample source could delay the initiation of medical intervention, thereby reducing the efficacy of antiviral treatment. The new assay, the SAMBA (simple amplification-based assay) Flu duplex test, is a dipstick-based molecular assay developed to provide a simple, accurate, and cost-effective solution for the diagnosis of influenza A/B viruses intended for near-patient testing. The test presents an alternative format of influenza virus molecular testing that utilizes isothermal amplification and visual detection of nucleic acid on a test strip. The entire test procedure (extraction, amplification, and detection) is integrated into an enclosed semiautomated system. Analytically, the SAMBA Flu duplex test detects 95 and 85 copies of viral genomes for influenza A and B viruses, respectively, with no cross-reactivity observed against other common respiratory pathogens. The clinical performance was established by blind testing of 328 nasal/throat and nasopharyngeal swab specimens from the United Kingdom and Belgium and comparing the results with the quantitative reverse transcription-PCR method routinely used in two public health laboratories. The SAMBA Flu duplex test showed a clinical sensitivity and specificity of 100% and 97.9% for influenza virus A and 100% and 100% for influenza virus B. The test provides a new technology that could facilitate simple and timely identification of influenza virus infection, potentially resulting in more efficient control measures.

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“…The limit of detection was determined using the sum of the negative control and three standard deviations (0 + 3SD) as the lowest detectable signal. [57][58][59][60]…”

Section: Drinking Water Assaymentioning

confidence: 99%

Reporter bacteriophage T7_NLCutilizes a novel NanoLuc::CBM fusion for the ultrasensitive detection ofEscherichia coliin water

Hinkley

Garing

Singh

et al. 2018

Analyst

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Rapid detection of bacteria responsible for foodborne diseases is a growing necessity for public health. Reporter bacteriophages (phages) are robust biorecognition elements uniquely suited for the rapid and sensitive detection of bacterial species. The advantages of phages include their host specificity, ability to distinguish viable and non-viable cells, low cost, and ease of genetic engineering. Upon infection with reporter phages, target bacteria express reporter enzymes encoded within the phage genome. In this study, the T7 coliphage was genetically engineered to express the newly developed luceriferase, NanoLuc (NLuc), as an indicator of bacterial contamination. While several genetic approaches were employed to optimize reporter enzyme expression, the novel achievement of this work was the successful fusion of the NanoLuc reporter to a carbohydrate binding module (CBM) with specificity to crystalline cellulose. This novel chimeric reporter (nluc::cbm) bestows the specific and irreversible immobilization of NanoLuc onto a low-cost, widely available crystalline cellulosic substrate. We have shown the possibility of detecting the immobilized fusion protein in a filter plate which resulted from a single CFU of E. coli. We then demonstrated that microcrystalline cellulose can be used to concentrate the fusion reporter from 100 mL water samples allowing a limit of detection of <10 CFU mL-1E. coli in 3 hours. Therefore, we conclude that our phage-based detection assay displays significant aptitude as a proof-of-concept drinking water diagnostic assay for the low-cost, rapid and sensitive detection of E. coli. Additional improvements in the capture efficiency of the phage-based fusion reporter should allow a limit of detection of <10 CFU per 100 mL.

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Development of Chemiluminescent Lateral Flow Assay for the Detection of Nucleic Acids

Cited by 27 publications

References 28 publications

Rapid detection of Salmonella using a redox cycling-based electrochemical method

Rapid detection of Salmonella using a redox cycling-based electrochemical method

Duplex Molecular Assay Intended for Point-of-Care Diagnosis of Influenza A/B Virus Infection

Reporter bacteriophage T7_NLCutilizes a novel NanoLuc::CBM fusion for the ultrasensitive detection ofEscherichia coliin water

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Development of Chemiluminescent Lateral Flow Assay for the Detection of Nucleic Acids

Cited by 27 publications

References 28 publications

Rapid detection of Salmonella using a redox cycling-based electrochemical method

Rapid detection of Salmonella using a redox cycling-based electrochemical method

Duplex Molecular Assay Intended for Point-of-Care Diagnosis of Influenza A/B Virus Infection

Reporter bacteriophage T7NLCutilizes a novel NanoLuc::CBM fusion for the ultrasensitive detection ofEscherichia coliin water

Contact Info

Product

Resources

About

Reporter bacteriophage T7_NLCutilizes a novel NanoLuc::CBM fusion for the ultrasensitive detection ofEscherichia coliin water