Rapid Detection of Brucella spp. and Elimination of Carryover Using Multiple Cross Displacement Amplification Coupled With Nanoparticles-Based Lateral Flow Biosensor

Li, Shijun; Liu, Ying; Wang, Yue; Wang, Ming; Liu, Chunting; Wang, Yi

doi:10.3389/fcimb.2019.00078

Cited by 33 publications

(36 citation statements)

References 23 publications

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“…The newly developed E. faecalis- MCDA-LFB method was capable of assaying 10 fg of E. faecalis genomic DNA (Figure 4), which was more sensitive than E. faecalis- LAMP-LFB detection (250 fg per reaction) reported by the previous study 25. Although the amplification products could be detected equally with the MG method used in the present study, the LFB was deemed as the preferred method as observing the amplification results is more objective and does not need any apparatus 19,27. In conclusion, the newly E. faecalis -MCDA-LFB assay in this study is a valuable method for the convenient, rapid, sensitive, specific, visual, reliable, and low-cost detection of E. faecalis in clinical and environmental samples, especially in resource-constrained settings.…”

Section: Discussionmentioning

confidence: 50%

A Novel Detection of Enterococcus faecalis Using Multiple Cross Displacement Amplification Linked with Gold Nanoparticle Lateral Flow Biosensor

Chen

et al. 2019

IDR

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BackgroundEnterococcus faecalis, an opportunistic bacterial pathogen, is one of the most frequently isolated bacterial species and cause of serious nosocomial infections in recent decades. A reliable and rapid assay for E. faecalis detection is significant for the diagnosis and follow-up treatment.MethodsA novel assay method, named multiple cross displacement amplification linked with nanoparticle-based lateral flow biosensor (MCDA-LFB), was applied for detecting E. faecalis strains. A set of special 10 primers was designed according to E. faecalis-specific gene Ef0027. The MCDA amplification conditions, including the target DNA concentration, reaction temperature and time, were optimized. The sensitivity and specificity of MCDA method were tested in the current study, and then, the MCDA-LFB technology was applied to detect the E. faecalis strain from clinical samples.ResultsThe E. faecalis specific primers were valid for the establishment of MCDA-LFB technology forthe detection of E. faecalis based on the Ef0027 gene. The MCDA amplification condition was optimized at 62°C for 35 min. The MCDA products were directly sensed and displayed with a biosensor. The full process, comprising genomic DNA template preparation (approximately 30 mins), amplification of MCDA (35 mins), and the product identification (approximately 2 mins), could be achieved in 70 mins. The MCDA technique could detect as little as 10 fg per reaction system of pure E. faecalis genomic DNA. The specificity of E. faecalis-MCDA-LFB method is 100%, with no cross-reactions to non-E. faecalis strains.ConclusionThe MCDA-LFB technique established in the present study is a reliable, simple, rapid, sensitive and specific method to assay E. faecalis and can be applied for the detection of clinical samples.

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Section: Discussionmentioning

confidence: 50%

A Novel Detection of Enterococcus faecalis Using Multiple Cross Displacement Amplification Linked with Gold Nanoparticle Lateral Flow Biosensor

Chen

et al. 2019

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“…Comparing the above three assay methods, we deemed the LFB was more visual. And it did no needs special instruments and regents [16,17]. Besides, LFB can detect both the IS6110 and mpb64 genes and make them visualized in a single test at the same time.…”

Section: Discussionmentioning

confidence: 99%

“…strains and Shigella spp [13], Vibrio parahaemolyticus [14] and Listeria monocytogenes [15]. Our team also created the MCDA method for Brucella spp [16] and Neisseria meningitidis [17] detection successfully.…”

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confidence: 99%

Two Target Genes Based Multiple Cross Displacement Amplification Combined with Lateral Flow Biosensor for the Detection of Mycobacrterium Tuberculosis Complex

Huang

Xiao

Yang

et al. 2020

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Background: Mycobacterium tuberculosis complex (MTBC) causes tuberculosis (TB), which is a global public health problem that seriously endangers public health. Hence, development of a new and rapid method to detect MTBC is of great significance for prevention and treatment of TB. Results: In this study, a multiple cross displacement amplification combined with nanoparticle-based lateral flow biosensor (MCDA-LFB) was developed to simultaneously detect two target genes (IS6110 and mpb64) of MTBC. One suit of specific multiple cross displacement amplification (MCDA) primers, which was designed for IS6110 and mpb64 gene, respectively, was validated through using the genomic DNA extracted from reference strain H37Rv. The MCDA products were analyzed using real-time turbidity curve, colorimetric indicator (Malachite Green, MG) and LFB. The conditions of amplification temperature and time were optimized and the established MCDA-LFB method was applied to detect the sputum specimens and MTBC strains from clinical samples. The results show that two sets of MCDA primers for the IS6110 and mpb64 genes have detected MTBC validly. The MCDA reaction conditions were optimized at 67 °C for 35 min. The limit of detection of MCDA assay based on IS6110 and mpb64 genes was 100 fg of genomic DNA template per reaction. The specificity of MCDA-LFB detection was 100%, and no cross-reactions for non-MTBC strains detection. The positive rate of MCDA-LFB for the detection of MTBC strains was equal to that of semi-nested automatic real-time PCR (Xpert MTB/RIF), and had a higher positive rate than acid-fast staining (AFS) when used for the detection of sputum samples. The whole procedure of MCDA-LFB, including genomic template preparation, MCDA reaction and products analysis, was completed with 70 min.Conclusion: The simplicity, rapidity, sensitivity and reliability of the MCDA-LFB based on IS6110 and mpb64 gene of MTBC developed in this study make it potentially significant for the prevention and treatment of TB.

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“…To overcome the drawbacks of such traditional detections, the multiple cross displacement amplification (MCDA) technique is a novel, low-cost, sensitive, and rapid isothermal amplification technology similar to LAMP, but with more specificity and sensitivity in microorganism assay, 13 which has been applied to detect several pathogens such as Mycobacterium tuberculosis, Neisseria meningitides, and Listeria monocytogenes. [14][15][16] Previous studies suggested that MCDA amplification products could been analyzed by various technologies, including agarose gel, turbidimetry, and colorimetric indicators (malachite green reagent).…”

Section: Introductionmentioning

confidence: 99%

Multiple Cross Displacement Amplification Linked with Nanoparticles-Based Lateral Flow Biosensor in Screening of Hepatitis B Virus in Clinical Application

Chen¹,

Zhou²,

Dong³

et al. 2021

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Background Hepatitis B virus (HBV) is a common pathogen that predominantly causes severe liver disease, and remains one of a huge challenge worldwide, especially in many resource-constrained areas. Developing a low-cost, sensitive, specific, and rapid approach for screening HBV is critical for its treatment and prevention. In the current study, a novel molecular detection approach, multiple cross displacement amplification (MCDA) coupled with polymer nanoparticle-based lateral flow biosensor (MCDA-LFB), was applied for detection of HBV in blood samples. Methods HBV standard substance and clinical donor serum samples were collected and used for the establishment and confirmation of the HBV-MCDA-LFB assay. A set of 10 MCDA primers was designed according to HBV-specific gene S . The HBV-MCDA-LFB assay conditions, including genomic template concentration, MCDA reaction temperature and time were optimized. The sensitivity and specificity of the HBV-MCDA -LFB assay were evaluated in this report. The HBV-MCDA-LFB assay was applied to detect the HBV agent from clinical samples. Results The HBV-MCDA primers based on the S gene were valid for establishment of MCDA assay. The HBV-MCDA reaction with optimized conditions could be carried out at a constant temperature 64°C for 35 min. The whole process, including sample preparation (5 min), genomic template extraction (~30 min), MCDA amplification (35 min), and LFB reading (~2 min), could be completed within 80 min. The sensitivity of this assay was 5 IU per reaction. The specificity was 100% for HBV-MCDA-LFB assay. Conclusion These results confirmed that the HBV-MCDA-LFB is a low-cost, sensitive, specific, simple, and rapid method for detecting HBV agents. This technique has great potential to develop a point-of-care testing (POCT) method in clinical practice, especially in endemic and resource-constrained regions.

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Rapid Detection of Brucella spp. and Elimination of Carryover Using Multiple Cross Displacement Amplification Coupled With Nanoparticles-Based Lateral Flow Biosensor

Cited by 33 publications

References 23 publications

<p>A Novel Detection of <em>Enterococcus</em> <em>faecalis</em> Using Multiple Cross Displacement Amplification Linked with Gold Nanoparticle Lateral Flow Biosensor</p>

<p>A Novel Detection of <em>Enterococcus</em> <em>faecalis</em> Using Multiple Cross Displacement Amplification Linked with Gold Nanoparticle Lateral Flow Biosensor</p>

Two Target Genes Based Multiple Cross Displacement Amplification Combined with Lateral Flow Biosensor for the Detection of Mycobacrterium Tuberculosis Complex

Multiple Cross Displacement Amplification Linked with Nanoparticles-Based Lateral Flow Biosensor in Screening of Hepatitis B Virus in Clinical Application

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