DEVELOPMENT AND EVALUATION OF A LOOP-MEDIATED ISOTHERMAL AMPLIFICATION (LAMP) METHOD FOR DETECTINGLISTERIA MONOCYTOGENESIN RAW MILK

2010 ◽  
Vol 30 (2) ◽  
pp. 251-262 ◽  
Author(s):  
DEGUO WANG ◽  
GUICHENG HUO ◽  
DAXI REN ◽  
YONGGANG LI
Keyword(s):  
Raw Milk ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Loop Mediated Isothermal Amplification

Development and Evaluation of a Loop-Mediated Isothermal Amplification (LAMP) Method for Detecting Foodborn Salmonella in Raw Milk

2013 ◽  
Vol 647 ◽  
pp. 577-582 ◽  
Author(s):  
Yong Zhen Wang ◽  
De Guo Wang
Keyword(s):  
Reference Method ◽  
Lamp Assay ◽  
Raw Milk ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Conventional Pcr ◽  
Detection Level ◽  
Loop Mediated Isothermal Amplification ◽  
Milk Samples ◽  
Food Borne

In present study, we reported the performance of a Loop-mediated isothermal amplification (LAMP) assay detecting food-borne pathogen Salmonella. Three pairs of primers were specially designed for recognizing eight distinct sequences on the target invA gene. Time and temperature conditions for amplification of Salmonella were optimized to be 40 min at 61°C. The LAMP assay gave with artificially contaminated raw milk samples detection limit level of 142 CFU/ml which corresponds to 6-9 cells per reaction tube, while the detection level of conventional PCR was 103 CFU/ml. Data on naturally contaminated raw milk samples indicated that the LAMP method was highly specific and sensitive, giving 89.58% concordance with the ISO 6579 reference method for the samples without enrichment and 100% concordance for the samples after enrichment.

Rapid Detection of Mycobacterium tuberculosis Complex by Loop-Mediated Isothermal Amplification Combined with Chemosensor

2013 ◽  
Vol 749 ◽  
pp. 449-452
Author(s):  
De Guo Wang
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Dna Amplification ◽  
Raw Milk ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Loop Mediated Isothermal Amplification ◽  
Milk Samples ◽  
Assay Time ◽  
Low Levels ◽  
Rapid Amplification

Loop-mediated isothermal amplification (LAMP) allowed rapid amplification of nucleic acids under isothermal conditions. It can be combined with a chemosensor for much more efficient, field-friendly detection of Mycobacterium tuberculosis complex. In this report, LAMP was performed at 65 °C for 10 min, followed by a rapid reaction of DNA amplification by-product, pyrophosphate ion, with chemosensor resulted in red disappearance. The detection limit of Mycobacterium tuberculosis complex by LAMP-Chemosensor was 3-5 copies, and the total assay time including 10 min for rapid DNA extraction was approximately 30 min. Data on naturally contaminated raw milk samples indicated that the LAMP method was highly specific and sensitive, giving 100% concordance with Real-time PCR. The results showed that the LAMP-Chemosensor method had the advantages of better sensitivity and speed and less dependence on equipment than the standard (PCR) for specifically detecting low levels of Mycobacterium tuberculosis complex DNA, and this can be useful in the field as a routine diagnostic tool.

Rapid and sensitive diagnosis of adenoviral keratoconjunctivitis by loop-mediated isothermal amplification (LAMP) method

2004 ◽  
Vol 28 (6) ◽  
pp. 445-450 ◽  
Author(s):  
Taketoshi Wakabayashi ◽  
Ryoko Yamashita ◽  
Tetsuhiko Kakita ◽  
Mito Kakita ◽  
Tetsuro Oshika
Keyword(s):  
Isothermal Amplification ◽  
Lamp Method ◽  
Loop Mediated Isothermal Amplification

scholarly journals Loop-Mediated Isothermal Amplification for Detection ofStaphylococcus aureusin Dairy Cow Suffering from Mastitis

2012 ◽  
Vol 2012 ◽  
pp. 1-5 ◽  
Author(s):  
Zhang Tie ◽  
Wang Chunguang ◽  
Wei Xiaoyuan ◽  
Zhao Xinghua ◽  
Zhong Xiuhui
Keyword(s):  
Staphylococcus Aureus ◽  
High Sensitivity ◽  
High Specificity ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Specific Primers ◽  
Loop Mediated Isothermal Amplification ◽  
Reaction Tube ◽  
Specificity And Sensitivity ◽  
Detectable Concentration

To develop a rapid detection method ofStaphylococcus aureususing loop-mediated isothermal amplification (LAMP), four specific primers were designed according to six distinct sequences of thenucgene. In addition, the specificity and sensitivity of LAMP were verified and compared with those of PCR. Results showed that the LAMP reaction was completed within 45 min at 62.5°C, and ladder bands were appeared in LAMP products analyzed by gel electrophoresis. After adding 1x SYBR Green l, the positive reaction tube showed green color and the negative reaction tube remained orange, indicating that the LAMP has high specificity. The minimal detectable concentration of LAMP was1×102 CFU/mL and that of PCR was1×104 CFU/mL, indicating that the LAMP was 100 times more sensitive than the PCR. The LAMP method for detection ofStaphylococcus aureushas many advantages, such as simple operation, high sensitivity, high specificity, and rapid analysis. Therefore, this method is more suitable for the rapid on-site detection ofStaphylococcus aureus.

scholarly journals Rapid identification of transgenic cotton (Gossypium hirsutum L.) plants by loop-mediated isothermal amplification

2011 ◽  
Vol 47 (No. 4) ◽  
pp. 140-148 ◽  
Author(s):  
N. Rostamkhani ◽  
A. Haghnazari ◽  
M. Tohidfar ◽  
A. Moradi
Keyword(s):  
Real Time ◽  
Real Time Pcr ◽  
Lamp Assay ◽  
Transgenic Cotton ◽  
Rapid Identification ◽  
Isothermal Amplification ◽  
Detection Sensitivity ◽  
Lamp Method ◽  
Loop Mediated Isothermal Amplification ◽  
Leaf Tissues

In an attempt to speed up the process of screening of transgenic cotton (G. hirsutum L.) plants, a visual and rapid loop-mediated isothermal amplification (LAMP) assay was adopted. Genomic DNA was extracted from fresh leaf tissues of T<sub>2</sub> transgenic cotton containing chitinase (chi) and cry1A(b) genes. Detection of genes of interest was performed by polymerase chain reaction (PCR), LAMP and real-time PCR methods. In LAMP assay the amplification was performed after 30 min at 65&deg;C when loop primers were involved in the reaction. The involvement of loop primers decreased the time needed for amplification. By testing serial tenfold dilutions (10<sup>&ndash;1</sup> to 10<sup>&ndash;8</sup>) of the genes of interest, the detection sensitivity of LAMP was found to be 100-fold higher than that of PCR. The rapid DNA extraction method and LAMP assay can be performed within 30 min and the derived LAMP products can be directly observed as visually detectable based on turbidity in the reaction tube. The accuracy of LAMP method in the screening of transgenes was confirmed by PCR and real-time PCR. The developed method was efficient, rapid and sensitive in the screening of cotton transgenic plants. This method can be applied to any other crops.

scholarly journals Detection and discrimination of multiple strains of Zika virus by reverse transcription-loop-mediated isothermal amplification

2020 ◽  
Vol 48 (1) ◽  
Author(s):  
Hiroka Aonuma ◽  
Itoe Iizuka-Shiota ◽  
Tokio Hoshina ◽  
Shigeru Tajima ◽  
Fumihiro Kato ◽  
...  
Keyword(s):  
Virus Strain ◽  
Reverse Transcription ◽  
Zika Virus ◽  
Virus Disease ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Simple Method ◽  
Loop Mediated Isothermal Amplification ◽  
Specificity And Sensitivity ◽  
Virus Strains

Abstract Background Monitoring both invasion of Zika virus disease into free countries and circulation in endemic countries is essential to avoid a global pandemic. However, the difficulty lies in detecting Zika virus due to the large variety of mutations in its genomic sequence. To develop a rapid and simple method with high accuracy, reverse transcription-loop-mediated isothermal amplification (RT-LAMP) was adopted for the detection of Zika virus strains derived from several countries. Results Common primers for RT-LAMP were designed based on the genomic sequences of two standard Zika strains: African lineage, MR-766, and Asian lineage, PRVABC59. RT-LAMP reactions using a screened primer set, targeting the NS3 region, detected both Zika virus strains. The minimum detectable quantity was 3 × 10−2 ng of virus RNA. Measurable lag of reaction times among strains was observed. The RT-LAMP method amplified the target virus sequence from the urine and serum of a patient with a travel history in the Caribbean Islands and also provided a prediction about which lineage of Zika virus strain was present. Conclusions The RT-LAMP method using a well-optimized primer set demonstrated high specificity and sensitivity for the detection of Zika virus strains with a variety in genomic RNA sequences. In combination with the simplicity of LAMP reaction in isothermal conditions, the optimized primer set established in this study may facilitate rapid and accurate diagnosis of Zika fever patients with virus strain information.

scholarly journals Direct detection of Mycobacterium avium in environmental water and scale samples by loop-mediated isothermal amplification

2013 ◽  
Vol 12 (2) ◽  
pp. 211-219 ◽  
Author(s):  
Yukiko Nishiuchi ◽  
Aki Tamaru ◽  
Yasuhiko Suzuki ◽  
Seigo Kitada ◽  
Ryoji Maekura ◽  
...  
Keyword(s):  
Mycobacterium Avium ◽  
Environmental Samples ◽  
Direct Detection ◽  
Culture Method ◽  
Environmental Water ◽  
Isothermal Amplification ◽  
Lamp Method ◽  
Culture Methods ◽  
Loop Mediated Isothermal Amplification ◽  
Conventional Culture

We previously demonstrated the colonization of Mycobacterium avium complex in bathrooms by the conventional culture method. In the present study, we aimed to directly detect M. avium organisms in the environment using loop-mediated isothermal amplification (LAMP), and to demonstrate the efficacy of LAMP by comparing the results with those obtained by culture. Our data showed that LAMP analysis has detection limits of 100 fg DNA/reaction for M. avium. Using an FTA® elute card, DNA templates were extracted from environmental samples from bathrooms in the residences of 29 patients with pulmonary M. avium disease. Of the 162 environmental samples examined, 143 (88%) showed identical results by both methods; 20 (12%) and 123 (76%) samples were positive and negative, respectively, for M. avium. Of the remaining 19 samples (12%), seven (5%) and 12 (7%) samples were positive by the LAMP and culture methods, respectively. All samples that contained over 20 colony forming units/primary isolation plate, as measured by the culture method, were also positive by the LAMP method. Our data demonstrate that the combination of the FTA elute card and LAMP can facilitate prompt detection of M. avium in the environment.

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