Loop-mediated isothermal amplification (LAMP) shield for Arduino DNA detection

NARCIS (Netherlands)

Velders, Aldrik H.; Schoen, Cor; Saggiomo, Vittorio

2018-01-01

Objective: Loop-mediated isothermal amplification (LAMP) of DNA is gaining relevance as a method to detect nucleic acids, as it is easier, faster, and more powerful than conventional Polymerase Chain Reaction. However, LAMP is still mostly used in laboratory settings, because of the lack of a cheap

Loop-mediated isothermal amplification (LAMP) shield for Arduino DNA detection.

Science.gov (United States)

Velders, Aldrik H; Schoen, Cor; Saggiomo, Vittorio

2018-02-01

Loop-mediated isothermal amplification (LAMP) of DNA is gaining relevance as a method to detect nucleic acids, as it is easier, faster, and more powerful than conventional Polymerase Chain Reaction. However, LAMP is still mostly used in laboratory settings, because of the lack of a cheap and easy, one-button device that can perform LAMP experiments. Here we show how to build and program an Arduino shield for a LAMP and detection of DNA. The here described Arduino Shield is cheap, easy to assemble, to program and use, it is battery operated and the detection of DNA is done by naked-eye so that it can be used in field.

Development of mRNA-based body fluid identification using reverse transcription loop-mediated isothermal amplification.

Science.gov (United States)

Satoh, Tetsuya; Kouroki, Seiya; Ogawa, Keita; Tanaka, Yorika; Matsumura, Kazutoshi; Iwase, Susumu

2018-04-25

Identifying body fluids from forensic samples can provide valuable evidence for criminal investigations. Messenger RNA (mRNA)-based body fluid identification was recently developed, and highly sensitive parallel identification using reverse transcription polymerase chain reaction (RT-PCR) has been described. In this study, we developed reverse transcription loop-mediated isothermal amplification (RT-LAMP) as a simple, rapid assay for identifying three common forensic body fluids, namely blood, semen, and saliva, and evaluated its specificity and sensitivity. Hemoglobin beta (HBB), transglutaminase 4 (TGM4), and statherin (STATH) were selected as marker genes for blood, semen, and saliva, respectively. RT-LAMP could be performed in a single step including both reverse transcription and DNA amplification under an isothermal condition within 60 min, and detection could be conveniently performed via visual fluorescence. Marker-specific amplification was performed in each assay, and no cross-reaction was observed among five representative forensically relevant body fluids. The detection limits of the assays were 0.3 nL, 30 nL, and 0.3 μL for blood, semen, and saliva, respectively, and their sensitivities were comparable with those of RT-PCR. Furthermore, RT-LAMP assays were applicable to forensic casework samples. It is considered that RT-LAMP is useful for body fluid identification.

Rapid detection of Brucella spp. using loop-mediated isothermal amplification (LAMP).

Science.gov (United States)

Chen, Shouyi; Li, Xunde; Li, Juntao; Atwill, Edward R

2013-01-01

Brucella spp. are facultative intracellular bacteria that cause zoonotic disease of brucellosis worldwide. Livestock that are most vulnerable to brucellosis include cattle, goats, and pigs. Brucella spp. cause serious health problems to humans and animals and economic losses to the livestock industry. Traditional methods for detection of Brucella spp. take 48-72 h (Kumar et al., J Commun Dis 29:131-137, 1997; Barrouin-Melo et al., Res Vet Sci 83:340-346, 2007) that do not meet the food industry's need of rapid detection. Therefore, there is an urgent need of fast, specific, sensitive, and inexpensive method for diagnosing of Brucella spp. Loop-mediated isothermal amplification (LAMP) is a method to amplify nucleic acid at constant temperatures. Amplification can be detected by visual detection, fluorescent stain, turbidity, and electrophoresis. We targeted at the Brucella-specific gene omp25 and designed LAMP primers for detection of Brucella spp. Amplification of DNA with Bst DNA polymerase can be completed at 65 °C in 60 min. Amplified products can be detected by SYBR Green I stain and 2.0% agarose gel electrophoresis. The LAMP method is feasible for detection of Brucella spp. from blood and milk samples.

Development of loop-mediated isothermal amplification method for ...

African Journals Online (AJOL)

A novel assay method to detect the highly virulent Porcine reproductive and respiratory syndrome virus (PRRSV) termed reverse transcriptase loop-mediated isothermal amplification (RT-LAMP), was reported by using hydroxynaphthol blue (HNB) as the LAMP product colorimetric judgment. By the set of special primers, ...

Loop-mediated isothermal amplification (LAMP) based detection of ...

African Journals Online (AJOL)

SAM

2014-05-07

May 7, 2014 ... 2 months for growing in a culture. Therefore, to control .... The LAMP reaction is carried out in a 25 µL reaction mixture containing ..... J. Fish Dis. 32(6):491-497. Goto M, Honda E, Ogura A, Nomoto A, Hanaki K (2009). Colorimetric detection of loop-mediated isothermal amplification reaction by using hydroxy ...

Identification of Methicillin-Resistant Staphylococcus aureus (MRSA) Using Simultaneous Detection of mecA, nuc, and femB by Loop-Mediated Isothermal Amplification (LAMP).

Science.gov (United States)

Chen, Changguo; Zhao, Qiangyuan; Guo, Jianwei; Li, Yanjun; Chen, Qiuyuan

2017-08-01

The aim of this study was to develop a rapid detection assay to identify methicillin-resistant Staphylococcus aureus by simultaneous testing for the mecA, nuc, and femB genes using the loop-mediated isothermal amplification (LAMP) method. LAMP primers were designed using online bio-software ( http://primerexplorer.jp/e/ ), and amplification reactions were performed in an isothermal temperature bath. The products were then examined using 2% agarose gel electrophoresis. MecA, nuc, and femB were confirmed by triplex TaqMan real-time PCR. For better naked-eye inspection of the reaction result, hydroxy naphthol blue (HNB) was added to the amplification system. Within 60 min, LAMP successfully amplified the genes of interest under isothermal conditions at 63 °C. The results of 2% gel electrophoresis indicated that when the Mg 2+ concentration in the reaction system was 6 μmol, the amplification of the mecA gene was relatively good, while the amplification of the nuc and femB genes was better at an Mg 2+ concentration of 8 μmol. Obvious color differences were observed by adding 1 μL (3.75 mM) of HNB into 25 μL reaction system. The LAMP assay was applied to 128 isolates cases of methicillin-resistant Staphylococcus aureus, which were separated from the daily specimens and identified by Vitek microbial identification instruments. The results were identical for both LAMP and PCR. LAMP offers an alternative detection assay for mecA, nuc, and femB and is faster than other methods.

Development of a reverse transcription loop-mediated isothermal amplification method for the rapid detection of avian influenza virus subtype H7.

Science.gov (United States)

Bao, Hongmei; Wang, Xiurong; Zhao, Yuhui; Sun, Xiaodong; Li, Yanbing; Xiong, Yongzhong; Chen, Hualan

2012-01-01

A rapid and sensitive reverse transcription loop-mediated isothermal amplification (RT-LAMP) method for the detection of the H7 avian influenza virus (H7 AIV) isotype was developed. The minimum detection limit of the RT-LAMP assay was 0.1-0.01 PFU per reaction for H7 AIV RNA, making this assay 100-fold more sensitive than the conventional RT-PCR method. This RT-LAMP assay also has the capacity to detect both high- and low-pathogenic H7 AIV strains. Using a pool of RNAs extracted from influenza viruses corresponding to all 15 HA subtypes (in addition to other avian pathogenic viruses), the RT-LAMP system was confirmed to amplify only H7 AIV RNA. Furthermore, specific pathogen free (SPF) chickens were infected artificially with H7 AIV, throat and cloacal swabs were collected, and viral shedding was examined using viral isolation, RT-PCR and RT-LAMP. Shedding was detected following viral isolation and RT-LAMP one day after infection, whereas viral detection using RT-PCR was effective only on day 3 post-infection. These results indicate that the RT-LAMP method could facilitate epidemiological surveillance and the rapid diagnosis of the avian influenza subtype H7. Copyright © 2011 Elsevier B.V. All rights reserved.

Visual Detection of Potato leafroll virus by One-step Reverse Transcription Loop-Mediated Isothermal Amplification of DNA with Hydroxynaphthol Blue Dye

NARCIS (Netherlands)

Ahmadi, S.; Almasi, A.M.; Fatehi, F.; Struik, P.C.; Moradi, A.

2013-01-01

Loop-mediated isothermal amplification (LAMP) assay is a novel technique for amplifying DNA under constant temperature, with high specificity, sensitivity, rapidity and efficiency. We applied reverse transcription loop-mediated isothermal amplification (RT-LAMP) to visually detect Potato leafroll

Development of Reverse Transcription Thermostable Helicase-Dependent DNA Amplification for the Detection of Tomato Spotted Wilt Virus.

Science.gov (United States)

Wu, Xinghai; Chen, Chanfa; Xiao, Xizhi; Deng, Ming Jun

2016-11-01

A protocol for the reverse transcription-helicase-dependent amplification (RT-HDA) of isothermal DNA was developed for the detection of tomato spotted wilt virus (TSWV). Specific primers, which were based on the highly conserved region of the N gene sequence in TSWV, were used for the amplification of virus's RNA. The LOD of RT-HDA, reverse transcriptase-loop-mediated isothermal amplification (RT-LAMP), and reverse transcriptase-polymerase chain reaction (RT-PCR) assays were conducted using 10-fold serial dilution of RNA eluates. TSWV sensitivity in RT-HDA and RT-LAMP was 4 pg RNA compared with 40 pg RNA in RT-PCR. The specificity of RT-HDA for TSWV was high, showing no cross-reactivity with other tomato and Tospovirus viruses including cucumber mosaic virus (CMV), tomato black ring virus (TBRV), tomato mosaic virus (ToMV), or impatiens necrotic spot virus (INSV). The RT-HDA method is effective for the detection of TSWV in plant samples and is a potential tool for early and rapid detection of TSWV.

Loop-mediated isothermal amplification (LAMP) assay for the diagnosis of fasciolosis in sheep and its application under field conditions

OpenAIRE

Mart?nez-Valladares, Mar?a; Rojo-V?zquez, Francisco Antonio

2016-01-01

Background Loop-mediated isothermal amplification (LAMP) is a very specific, efficient, and rapid gene amplification procedure in which the reaction can run at a constant temperature. In the current study we have developed a LAMP assay to improve the diagnosis of Fasciola spp. in the faeces of sheep. Findings After the optimisation of the LAMP assay we have shown similar results between this technique and the standard PCR using the outer primers of the LAMP reaction. In both cases the limit o...

Novel reverse transcription loop-mediated isothermal amplification for rapid detection of foot-and-mouth disease virus

DEFF Research Database (Denmark)

Dukes, J.P.; King, D.P.; Alexandersen, Søren

2006-01-01

Speed is paramount in the diagnosis of foot-and-mouth disease (FMD) and simplicity is required if a test is to be deployed in the field. The development of a one-step, reverse transcription loop-mediated amplification (RT-LAMP) assay enables FMD virus (FMDV) to be detected in under an hour...... in a single tube without thermal cycling. A fragment of the 3D RNA polymerase gene of the virus is amplified at 65 degrees C in the presence of a primer mixture and both reverse transcriptase and Bst DNA polymerase. Compared with real-time RT-PCR, RT-LAMP was consistently faster, and ten copies of FMDV...... vesicular diseases and from that of genetically related picornaviruses. Diagnostic sensitivity was validated by the amplification of reference FMDV strains and archival material from field cases of FMD. In comparison with the performance of the established diagnostic TaqMan (R) assay, RT-LAMP appears...

Rapid typing of foot-and-mouth disease serotype Asia 1 by reverse transcription loop-mediated isothermal amplification

Directory of Open Access Journals (Sweden)

Chen Hao-tai

2011-10-01

Full Text Available Abstract A reverse transcriptase loop-mediated isothermal amplification (RT-LAMP assay was rapidly used to detect serotype Asia 1 of foot-and-mouth disease virus (FMDV within 45 min at 61°C. All FMDV serotype Asia 1 reference strains were positive by RT-LAMP, while other viruses such as FMDV serotypes O, C, A and classical swine fever virus, swine vesicular disease virus, porcine reproductive and respiratory syndrome virus and Japanese encephalitis virus remained negative. Furthermore, FMDV sreotype Asia 1 positive samples were able to detect by RT-LAMP assay. This RT-LAMP assay may be suitable particularly for diagnosis of FMDV serotype Asia 1 infection in field stations.

Detection of phytoplasma by loop-mediated isothermal amplification of DNA (LAMP).

Science.gov (United States)

Obura, E; Masiga, D; Wachira, F; Gurja, B; Khan, Z R

2011-02-01

Napier stunt phytoplasma (16SrXI and 16SrIII) in eastern Africa is a serious threat to the expansion of Napier grass (Pennisetum purpureum) farming in the region, where it is widely cultivated as fodder in zero grazing livestock systems. The grass has high potential for bio-fuel production, and has been adopted by farmers as a countermeasure to cereal stem borer Lepidoptera, since it attracts and traps the insect. Diagnosis of stunt phytoplasma have been largely by nested polymerase chain reaction (nPCR) targeting the 16S rRNA gene. However, the method is laborious, costly and technically demanding. This investigation has developed a simpler but effective phytoplasma diagnostic tool, called; loop-mediated isothermal amplification of DNA (LAMP). The assay was tested on 8 symptomatic and 8 asymptomatic plants, while its detection limit was compared to nested PCR using samples serially diluted from 3 ng/μl to 0.38 pg/μl. Molecular typing of LAMP products was determined by BsrI restriction digestion and Southern blot analysis. The assay sensitivity, positive and negative predictive values were estimated, while the specificity was tested on 11 phytoplasma groups. LAMP was specific to 5 phytoplasma groups: 16SrVI, X, XI and XVI. BsrI restriction digestion produced two predicted fragments, and there was specific binding of probe DNA to the LAMP amplicons in Southern blot analysis. The assay sensitivity was 100%, while the positive and negative predictive values were 63 and 100% respectively. LAMP was 20-fold more sensitive than nested PCR. This study validates LAMP for routine diagnosis of Napier stunt and other closely related phytoplasmas. Copyright © 2010 Elsevier B.V. All rights reserved.

Development of a Rapid Detection Method for Potato virus X by Reverse Transcription Loop-Mediated Isothermal Amplification

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Joojin Jeong

2015-09-01

Full Text Available The primary step for efficient control of viral diseases is the development of simple, rapid, and sensitive virus detection. Reverse transcription loop-mediated isothermal amplification (RT-LAMP has been used to detect viral RNA molecules because of its simplicity and high sensitivity for a number of viruses. RT-LAMP for the detection of Potato virus X (PVX was developed and compared with conventional reverse transcription polymerase chain reaction (RT-PCR to demonstrate its advantages over RT-PCR. RT-LAMP reactions were conducted with or without a set of loop primers since one out of six primers showed PVX specificity. Based on real-time monitoring, RT-LAMP detected PVX around 30 min, compared to 120 min for RT-PCR. By adding a fluorescent reagent during the reaction, the extra step of visualization by gel electrophoresis was not necessary. RT-LAMP was conducted using simple inexpensive instruments and a regular incubator to evaluate whether RNA could be amplified at a constant temperature instead of using an expensive thermal cycler. This study shows the potential of RT-LAMP for the diagnosis of viral diseases and PVX epidemiology because of its simplicity and rapidness compared to RT-PCR.

Demonstration of a very inexpensive, turbidimetric, real-time, RT-LAMP detection platform using shrimp Laem-Singh virus (LSNV as a model.

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Narong Arunrut

Full Text Available Rapid and accurate detection of pathogens under field laboratory conditions is necessary for effective control of veterinary pathogens. Here we describe a prototype, portable, pathogen detection device developed for single tube, real-time, reverse transcription, loop-mediated isothermal amplification (RT-LAMP using Laem-Singh virus (LSNV as a model. LSNV is an RNA virus and a component cause of growth retardation in black tiger shrimp. We chose its RNA-dependent RNA polymerase (RdRp gene as the target for our tests. The basis for detection was measurement of turbidity arising from formation of a white, insoluble magnesium pyrophosphate precipitate byproduct upon amplification of the RdRp target sequence from 100 ng template RNA extracted from shrimp. The measurement device consisted of a heating block to maintain constant temperature in the RT-LAMP reaction for 8 Eppindorf sample tubes, a light-emitting diode (LED light source providing red light emission at 650 nm wavelength to pass through sample tubes, a light dependent resistance (LDR photo-detector and a software program to report turbidity events and could potentially be marketed for under US$3000. The device was connected to a computer to display real-time results in a variety of formats. The optimized protocol for LSNV detection consisted of incubation of the sample tubes at 65 °C for 1 h during which turbidity was continuously measured, and quantitative results could be obtained by reaction time measurement. The sensitivity of detection was comparable to that of conventional nested RT-PCR and there was no cross reaction with other common shrimp viruses. The device was used for quantitative measurement of relative copy numbers of LSNV RdRp in 8 shrimp tissues and they were found to be highest in the gills followed in order by the lymphoid organ and hemolymph (p ≤ 0.05. This platform can be easily adapted for detection of other pathogens under field laboratory settings.

LAVA: An Open-Source Approach To Designing LAMP (Loop-Mediated Isothermal Amplification DNA Signatures

Directory of Open Access Journals (Sweden)

Gardner Shea N

2011-06-01

Full Text Available Abstract Background We developed an extendable open-source Loop-mediated isothermal AMPlification (LAMP signature design program called LAVA (LAMP Assay Versatile Analysis. LAVA was created in response to limitations of existing LAMP signature programs. Results LAVA identifies combinations of six primer regions for basic LAMP signatures, or combinations of eight primer regions for LAMP signatures with loop primers, which can be used as LAMP signatures. The identified primers are conserved among target organism sequences. Primer combinations are optimized based on lengths, melting temperatures, and spacing among primer sites. We compare LAMP signature candidates for Staphylococcus aureus created both by LAVA and by PrimerExplorer. We also include signatures from a sample run targeting all strains of Mycobacterium tuberculosis. Conclusions We have designed and demonstrated new software for identifying signature candidates appropriate for LAMP assays. The software is available for download at http://lava-dna.googlecode.com/.

Real-time sequence-validated loop-mediated isothermal amplification assays for detection of Middle East respiratory syndrome coronavirus (MERS-CoV.

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Sanchita Bhadra

Full Text Available The Middle East respiratory syndrome coronavirus (MERS-CoV, an emerging human coronavirus, causes severe acute respiratory illness with a 35% mortality rate. In light of the recent surge in reported infections we have developed asymmetric five-primer reverse transcription loop-mediated isothermal amplification (RT-LAMP assays for detection of MERS-CoV. Isothermal amplification assays will facilitate the development of portable point-of-care diagnostics that are crucial for management of emerging infections. The RT-LAMP assays are designed to amplify MERS-CoV genomic loci located within the open reading frame (ORF1a and ORF1b genes and upstream of the E gene. Additionally we applied one-step strand displacement probes (OSD for real-time sequence-specific verification of LAMP amplicons. Asymmetric amplification effected by incorporating a single loop primer in each assay accelerated the time-to-result of the OSD-RT-LAMP assays. The resulting assays could detect 0.02 to 0.2 plaque forming units (PFU (5 to 50 PFU/ml of MERS-CoV in infected cell culture supernatants within 30 to 50 min and did not cross-react with common human respiratory pathogens.

Real-time DNA Amplification and Detection System Based on a CMOS Image Sensor.

Science.gov (United States)

Wang, Tiantian; Devadhasan, Jasmine Pramila; Lee, Do Young; Kim, Sanghyo

2016-01-01

In the present study, we developed a polypropylene well-integrated complementary metal oxide semiconductor (CMOS) platform to perform the loop mediated isothermal amplification (LAMP) technique for real-time DNA amplification and detection simultaneously. An amplification-coupled detection system directly measures the photon number changes based on the generation of magnesium pyrophosphate and color changes. The photon number decreases during the amplification process. The CMOS image sensor observes the photons and converts into digital units with the aid of an analog-to-digital converter (ADC). In addition, UV-spectral studies, optical color intensity detection, pH analysis, and electrophoresis detection were carried out to prove the efficiency of the CMOS sensor based the LAMP system. Moreover, Clostridium perfringens was utilized as proof-of-concept detection for the new system. We anticipate that this CMOS image sensor-based LAMP method will enable the creation of cost-effective, label-free, optical, real-time and portable molecular diagnostic devices.

Loop-mediated isothermal amplification (LAMP) based detection of Colletotrichum falcatum causing red rot in sugarcane.

Science.gov (United States)

Chandra, Amaresh; Keizerweerd, Amber T; Que, Youxiong; Grisham, Michael P

2015-08-01

Red rot, caused by Colletotrichum falcatum, is a destructive disease prevalent in most sugarcane-producing countries. Disease-free sugarcane planting materials (setts) are essential as the pathogen spreads primarily through infected setts. The present study was undertaken to develop a loop-mediated isothermal amplification (LAMP) assay for the detection of C. falcatum. C. falcatum genomic DNA was isolated from pure mycelium culture and infected tissues. Four sets of primers corresponding to a unique DNA sequence specific to C. falcatum were designed. Specificity of the LAMP test was checked with DNA of another fungal pathogen of sugarcane, Puccinia melanocephala, as well as two closely-related species, Colletotrichum fructivorum and Colletotrichum acutatum. No reaction was found with the three pathogens. When C. falcatum DNA from pure culture was used in a detection limit analysis, sensitivity of the LAMP method was observed to be ten times higher than that of conventional PCR; however, sensitivity was only 5 times higher when DNA from C. falcatum-infected tissues was used. Using the LAMP assay, C. falcatum DNA is amplified with high specificity, efficiency, and rapidity under isothermal conditions. Moreover, visual judgment of color change in <1 h without further post-amplification processing makes the LAMP method convenient, economical, and useful in diagnostic laboratories and the field.

Loop-Mediated Isothermal Amplification (LAMP): Emergence As an Alternative Technology for Herbal Medicine Identification.

Science.gov (United States)

Li, Jing-Jian; Xiong, Chao; Liu, Yue; Liang, Jun-Song; Zhou, Xing-Wen

2016-01-01

Correct identification of medicinal plant ingredients is essential for their safe use and for the regulation of herbal drug supply chain. Loop-mediated isothermal amplification (LAMP) is a recently developed approach to identify herbal medicine species. This novel molecular biology technique enables timely and accurate testing, especially in settings where infrastructures to support polymerase chain reaction facilities are lacking. Studies that used this method have altered our view on the extent and complexity of herbal medicine identification. In this review, we give an introduction into LAMP analysis, covers the basic principles and important aspects in the development of LAMP analysis method. Then we presented a critical review of the application of LAMP-based methods in detecting and identifying raw medicinal plant materials and their processed products. We also provide a practical standard operating procedure (SOP) for the utilization of the LAMP protocol in herbal authentication, and consider the prospects of LAMP technology in the future developments of herbal medicine identification and the challenges associated with its application.

Evaluation of a loop-mediated isothermal amplification (LAMP) method for rapid on-site detection of horse meat

NARCIS (Netherlands)

Aartse, Aafke; Scholtens-Toma, Ingrid; A, van der Hans J.G.; Boersma-Greve, Monique M.; Prins, Theo W.; Ginkel, van Leen A.; Kok, Esther J.; Bovee, Toine F.H.

2017-01-01

Detection of horse DNA by loop-mediated isothermal amplification (LAMP) seems one of the most promising methods to meet the criteria of fast, robust, cost efficient, specific, and sensitive on-site detection. In the present study an assessment of the specificity and sensitivity of the LAMP horse

LOOP-MEDIATED ISOTHERMAL AMPLIFICATION (LAMP) FOR THE DETECTION OF SALMONELLA SPP. ISOLATED FROM DIFFERENT FOOD TYPES

OpenAIRE

Kostas Papanotas; Petros A. Kokkinos; Panos G. Ziros; Apostolos Vantarakis

2012-01-01

The objective of this study was the application and evaluation of a loop-mediated isothermal amplification (LAMP) method for the detection of Salmonella spp. strains isolated from food samples. Salmonella specific invA gene sequences (50 strains, 15 serotypes) were amplified at 65oC in 60 min. All of the strains of Salmonella subsp. Enterica were shown to be positive using the LAMP reaction assay, whereas, all other bacteria, virus and yeasts tested in this study were negative. LAMP products ...

Urine sample used for detection of toxoplasma gondii infection by loop-mediated isothermal amplification (LAMP).

Science.gov (United States)

Hu, Xin; Pan, Chang-Wang; Li, Ya-Fei; Wang, Han; Tan, Feng

2012-02-01

In this study, a loop-mediated isothermal amplification (LAMP) assay was established to detect Toxoplasma gondii DNA in mice infected with T. gondii PRU strain. This LAMP assay was based on the sequence of highly repetitive B1 gene. The detection limit of T. gondii LAMP assay was 1 pg of T. gondii DNA, which was evaluated using 10-fold serially diluted DNA of cultured parasites. The LAMP assay was also highly specific for T. gondii and able to detect T. gondii DNA in urine of mice treated with dexamethasone at 90 day post infection (p.i.), although this assay could not detect the DNA in mice urine 2-6 days p.i. These results demonstrated that LAMP is effective for evaluation of therapy effectiveness for T. gondii infection. The established LAMP assay may represent a useful and practical tool for the routine diagnosis and therapeutic evaluation of human toxoplasmosis.

Development of a Novel Loop-Mediated Isothermal Amplification (LAMP) Assay for the Detection of Rickettsia spp.

Science.gov (United States)

Hanaoka, Nozomu; Matsutani, Minenosuke; Satoh, Masaaki; Ogawa, Motohiko; Shirai, Mutsunori; Ando, Shuji

2017-01-24

We developed a novel loop-mediated isothermal amplification (LAMP) method to detect Rickettsia spp., including Rickettsia prowazekii and R. typhi. Species-specific LAMP primers were developed for orthologous genes conserved among Rickettsia spp. The selected modified primers could detect all the Rickettsia spp. tested. The LAMP method was successfully used to detect 100 DNA copies of Rickettsia spp. within approximately 60 min at 63℃. Therefore, this method may be an excellent tool for the early diagnosis of rickettsiosis in a laboratory or in the field.

Visual detection of West Nile virus using reverse transcription loop-mediated isothermal amplification combined with a vertical flow visualization strip

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Zengguo eCao

2016-04-01

Full Text Available West Nile virus (WNV causes a severe zoonosis, which can lead to a large number of casualties and considerable economic losses. A rapid and accurate identification methodfor WNV for use in field laboratories is urgently needed. Here, a method utilizing reverse transcription loop-mediated isothermal amplification combined with a vertical flow visualization strip (RT-LAMP-VF was developed to detect the envelope (E gene of WNV. The RT-LAMP-VF assay could detect 102 copies/μl ofan WNV RNA standard using a 40 min amplification reaction followed by a 2 min incubationof the amplification product on the visualization strip, and no cross-reaction with other closely related members of theFlavivirus genus was observed. The assay was further evaluated using cells and mouse brain tissues infected with a recombinant rabies virus expressing the E protein of WNV.The assay produced sensitivities of 101.5TCID50/ml and 101.33 TCID50/ml for detection of the recombinant virus in the cells and brain tissues, respectively. Overall, the RT-LAMP-VF assay developed in this study is rapid, simple and effective, and it is therefore suitable for clinical application in the field.

Loop-mediated isothermal amplification (LAMP) method for detection of genetically modified maize T25.

Science.gov (United States)

Xu, Junyi; Zheng, Qiuyue; Yu, Ling; Liu, Ran; Zhao, Xin; Wang, Gang; Wang, Qinghua; Cao, Jijuan

2013-11-01

The loop-mediated isothermal amplification (LAMP) assay indicates a potential and valuable means for genetically modified organism (GMO) detection especially for its rapidity, simplicity, and low cost. We developed and evaluated the specificity and sensitivity of the LAMP method for rapid detection of the genetically modified (GM) maize T25. A set of six specific primers was successfully designed to recognize six distinct sequences on the target gene, including a pair of inner primers, a pair of outer primers, and a pair of loop primers. The optimum reaction temperature and time were verified to be 65°C and 45 min, respectively. The detection limit of this LAMP assay was 5 g kg(-1) GMO component. Comparative experiments showed that the LAMP assay was a simple, rapid, accurate, and specific method for detecting the GM maize T25.

Visual detection of the human metapneumovirus using reverse transcription loop-mediated isothermal amplification with hydroxynaphthol blue dye

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Wang Xiang

2012-07-01

Full Text Available Abstract Background Human metapneumovirus (hMPV is a major cause of acute respiratory infections ranging from wheezing to bronchiolitis and pneumonia in children worldwide. The objective of this study is to develop a visual reverse transcription loop-mediated isothermal amplification (RT-LAMP assay for the detection of hMPV and applied to the clinical samples. Results In this study, visual RT-LAMP assay for hMPV was performed in one step with the addition of hydroxynaphthol blue (HNB, and were used to detect respiratory samples. Six primers, including two outer primers (F3 and B3, two inner primers (FIP, BIP and two loop primers (LF and LB, were designed for hMPV N gene by the online software. Moreover, the RT-LAMP assay showed good specificity and no cross-reactivity was observed with human rhinovirus (HRV, human respiratory syncytial Virus (RSV, or influenza virus A/PR/8/34 (H1N1. The detection limit of the RT-LAMP assay was approximately ten viral RNA copies, lower than that of traditional reverse transcriptase polymerase chain reaction (RT-PCR 100 RNA copies. In the 176 nasopharyngeal samples, 23 (13.1% were conformed as hMPV positive by RT-LAMP, but 18 (10.2% positive by RT-PCR. Conclusion Compared with conventional RT-PCR, the visual hMPV RT-LAMP assay performed well in the aspect of detect time, sensitivity, specificity and visibility. It is anticipated that the RT-LAMP will be used for clinical tests in hospital or field testing during outbreaks and in emergency.

A simple and rapid identification method for newly emerged porcine Deltacoronavirus with loop-mediated isothermal amplification

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Fanfan Zhang

2017-10-01

Full Text Available Abstract Background Porcine Deltacoronavirus (PDCoV is a newly emerged enteropathogenic coronavirus that causes diarrhea and mortality in neonatal piglets. PDCoV has spread to many countries around the world, leading to significant economic losses in the pork industry. Therefore, a rapid and sensitive method for detection of PDCoV in clinical samples is urgently needed. Results In this study, we developed a single-tube one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP assay specific for nucleocapsid gene to diagnose and monitor PDCoV infections. The detection limit of RT-LAMP assay was 1 × 101 copies of PDCoV, which was approximately 100-fold more sensitive than gel-based one-step reverse transcription polymerase chain reaction (RT-PCR. This assay could specifically amplify PDCoV and had no cross amplification with porcine epidemic diarrhea virus (PEDV, transmissible gastroenteritis virus (TGEV, porcine kobuvirus (PKoV, porcine astrovirus (PAstV, porcine reproductive and respiratory syndrome virus (PRRSV, classic swine fever virus (CSFV, and porcine circovirus type 2 (PCV2. By screening a panel of clinical specimens (N = 192, this method presented a similar sensitivity with nested RT-PCR and was 1–2 log more sensitive than conventional RT-PCR in detection of PDCoV. Conclusions The RT-LAMP assay established in this study is a potentially valuable tool, especially in low-resource laboratories and filed settings, for a rapid diagnosis, surveillance, and molecular epidemiology investigation of PDCoV infections. To the best of our knowledge, this is the first work for detection of newly emerged PDCoV with LAMP technology.

Loop-mediated isothermal amplification (LAMP) assay for the diagnosis of fasciolosis in sheep and its application under field conditions.

Science.gov (United States)

Martínez-Valladares, María; Rojo-Vázquez, Francisco Antonio

2016-02-05

Loop-mediated isothermal amplification (LAMP) is a very specific, efficient, and rapid gene amplification procedure in which the reaction can run at a constant temperature. In the current study we have developed a LAMP assay to improve the diagnosis of Fasciola spp. in the faeces of sheep. After the optimisation of the LAMP assay we have shown similar results between this technique and the standard PCR using the outer primers of the LAMP reaction. In both cases the limit of detection was 10 pg; also, the diagnosis of fasciolosis was confirmed during the first week post-infection in experimental infected sheep by both techniques. In eight naturally infected sheep, the infection with F. hepatica was confirmed in all animals before a treatment with triclabendazole and on day 30 post treatment in two sheep using the LAMP assay; however, when we carried out the standard PCR with the outer primers, the results before treatment were the same but on day 30 post-treatment the infection was only confirmed in one out of the two sheep. On the other hand, the standard PCR took around 3 h to obtain a result, comparing with 1 h and 10 min for the LAMP assay. The LAMP assay described here could be a good alternative to conventional diagnostic methods to detect F. hepatica in faeces since it solves the drawbacks of the standard PCR.

Detection of BCR-ABL Fusion mRNA Using Reverse Transcriptase Loop-mediated Isothermal Amplification

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Dugan, L C; Hall, S; Kohlgruber, A; Urbin, S; Torres, C; Wilson, P

2011-12-08

RT-PCR is commonly used for the detection of Bcr-Abl fusion transcripts in patients diagnosed with chronic myelogenous leukemia, CML. Two fusion transcripts predominate in CML, Br-Abl e13a2 and e14a2. They have developed reverse transcriptase isothermal loop-mediated amplification (RT-LAMP) assays to detect these two fusion transcripts along with the normal Bcr transcript.

Visual loop-mediated isothermal amplification (LAMP) for the rapid diagnosis of Enterocytozoon hepatopenaei (EHP) infection.

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T, Sathish Kumar; A, Navaneeth Krishnan; J, Joseph Sahaya Rajan; M, Makesh; K P, Jithendran; S V, Alavandi; K K, Vijayan

2018-05-01

The emerging microsporidian parasite Enterocytozoon hepatopenaei (EHP), the causative agent of hepatopancreatic microsporidiosis, has been widely reported in shrimp-farming countries. EHP infection can be detected by light microscopy observation of spores (1.7 × 1 μm) in stained hepatopancreas (HP) tissue smears, HP tissue sections, and fecal samples. EHP can also be detected by polymerase chain reaction (PCR) targeting the small subunit (SSU) ribosomal RNA (rRNA) gene or the spore wall protein gene (SWP). In this study, a rapid, sensitive, specific, and closed tube visual loop-mediated isothermal amplification (LAMP) protocol combined with FTA cards was developed for the diagnosis of EHP. LAMP primers were designed based on the SSU rRNA gene of EHP. The target sequence of EHP was amplified at constant temperature of 65 °C for 45 min and amplified LAMP products were visually detected in a closed tube system by using SYBR™ green I dye. Detection limit of this LAMP protocol was ten copies. Field and clinical applicability of this assay was evaluated using 162 field samples including 106 HP tissue samples and 56 fecal samples collected from shrimp farms. Out of 162 samples, EHP could be detected in 62 samples (47 HP samples and 15 fecal samples). When compared with SWP-PCR as the gold standard, this EHP LAMP assay had 95.31% sensitivity, 98.98% specificity, and a kappa value of 0.948. This simple, closed tube, clinically evaluated visual LAMP assay has great potential for diagnosing EHP at the farm level, particularly under low-resource circumstances.

Development of a Reverse Transcription Loop-Mediated Isothermal Amplification Method for the Rapid Detection of Subtype H7N9 Avian Influenza Virus

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Hongmei Bao

2014-01-01

Full Text Available A novel influenza A (H7N9 virus has emerged in China. To rapidly detect this virus from clinical samples, we developed a reverse transcription loop-mediated isothermal amplification (RT-LAMP method for the detection of the H7N9 virus. The minimum detection limit of the RT-LAMP assay was 0.01 PFU H7N9 virus, making this method 100-fold more sensitive to the detection of the H7N9 virus than conventional RT-PCR. The H7N9 virus RT-LAMP assays can efficiently detect different sources of H7N9 influenza virus RNA (from chickens, pigeons, the environment, and humans. No cross-reactive amplification with the RNA of other subtype influenza viruses or of other avian respiratory viruses was observed. The assays can effectively detect H7N9 influenza virus RNA in drinking water, soil, cloacal swab, and tracheal swab samples that were collected from live poultry markets, as well as human H7N9 virus, in less than 30 min. These results suggest that the H7N9 virus RT-LAMP assays were efficient, practical, and rapid diagnostic methods for the epidemiological surveillance and diagnosis of influenza A (H7N9 virus from different resource samples.

Rapid detection of Prunus necrotic ringspot virus using magnetic nanoparticle-assisted reverse transcription loop-mediated isothermal amplification.

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Zong, Xiaojuan; Wang, Wenwen; Wei, Hairong; Wang, Jiawei; Chen, Xin; Xu, Li; Zhu, Dongzi; Tan, Yue; Liu, Qingzhong

2014-11-01

Prunus necrotic ringspot virus (PNRSV) has seriously reduced the yield of Prunus species worldwide. In this study, a highly efficient and specific two-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) was developed to detect PNRSV. Total RNA was extracted from sweet cherry leaf samples using a commercial kit based on a magnetic nanoparticle technique. Transcripts were used as the templates for the assay. The results of this assay can be detected using agarose gel electrophoresis or by assessing in-tube fluorescence after adding SYBR Green I. The assay is highly specific for PNRSV, and it is more sensitive than reverse-transcription polymerase chain reaction (RT-PCR). Restriction enzyme digestion verified further the reliability of this RT-LAMP assay. To our knowledge, this is the first report of the application of RT-LAMP to PNRSV detection in Prunus species. Copyright © 2014 Elsevier B.V. All rights reserved.

A rapid and sensitive loop-mediated isothermal amplification procedure (LAMP) for Mycoplasma hyopneumoniae detection based on the p36 gene.

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Liu, M J; Du, G M; Bai, F F; Wu, Y Z; Xiong, Q Y; Feng, Z X; Li, B; Shao, G Q

2015-05-04

The aim of this study was to establish a method for sensitive and rapid diagnosis of Mycoplasma hyopneumoniae in clinical specimens. To this effect, we employed three sets of primers specifically designed for amplification of nucleic acids under isothermal conditions. After optimization of reaction conditions, M. hyopneumoniae could be successfully detected at 63°C in 45 min through use of the loop-mediated isothermal amplification (LAMP) assay. A positive reaction was identified visually as white precipitate and confirmed by gel electrophoresis. The detection limit for this assay was 10 copies/μL, as observed by electrophoretic analysis. The accuracy of the LAMP reaction was confirmed by restriction endonuclease digestion as well as by direct sequencing of the amplified product. This method can specifically detect M. hyopneumoniae; other species with high homology and other bacterial and virus strains gave negative results. To test the utility of this procedure, the LAMP assay was applied to 40 clinical samples collected from swine lung tissues experimentally challenged with M. hyopneumoniae isolates, and compared to the results from a real-time polymerase chain reaction (PCR) assay. A concordance of 100% was observed between the two assays. In conclusion, the results from our study demonstrated that the LAMP assay provided a rapid reaction and was inexpensive to perform, with no need of complex instruments or systems such as Geneamp PCR. The LAMP assay may therefore be applied in routine diagnosis in the clinical laboratory and for in-field detection of M. hyopneumoniae infection.

Reverse transcription loop-mediated isothermal amplification assays for rapid identification of eastern and western strains of bluetongue virus in India.

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Maan, S; Maan, N S; Batra, K; Kumar, A; Gupta, A; Rao, Panduranga P; Hemadri, Divakar; Reddy, Yella Narasimha; Guimera, M; Belaganahalli, M N; Mertens, P P C

2016-08-01

Bluetongue virus (BTV) infects all ruminants, including cattle, goats and camelids, causing bluetongue disease (BT) that is often severe in naïve deer and sheep. Reverse-transcription-loop-mediated-isothermal-amplification (RT-LAMP) assays were developed to detect eastern or western topotype of BTV strains circulating in India. Each assay uses four primers recognizing six distinct sequences of BTV genome-segment 1 (Seg-1). The eastern (e)RT-LAMP and western (w)RT-LAMP assay detected BTV RNA in all positive isolates that were tested (n=52, including Indian BTV-1, -2, -3, -5, -9, -10, -16, -21 -23, and -24 strains) with high specificity and efficiency. The analytical sensitivity of the RT-LAMP assays is comparable to real-time RT-PCR, but higher than conventional RT-PCR. The accelerated eRT-LAMP and wRT-LAMP assays generated detectable levels of amplified DNA, down to 0.216 fg of BTV RNA template or 108 fg of BTV RNA template within 60-90min respectively. The assays gave negative results with RNA from foot-and-mouth-disease virus (FMDV), peste des petits ruminants virus (PPRV), or DNA from Capripox viruses and Orf virus (n=10), all of which can cause clinical signs similar to BT. Both RT-LAMP assays did not show any cross-reaction among themselves. The assays are rapid, easy to perform, could be adapted as a 'penside' test making them suitable for 'front-line' diagnosis, helping to identify and contain field outbreaks of BTV. Copyright © 2016 Elsevier B.V. All rights reserved.

Development of loop-mediated isothermal amplification (LAMP assay for rapid and sensitive identification of ostrich meat.

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Amir Abdulmawjood

Full Text Available Animal species identification is one of the primary duties of official food control. Since ostrich meat is difficult to be differentiated macroscopically from beef, therefore new analytical methods are needed. To enforce labeling regulations for the authentication of ostrich meat, it might be of importance to develop and evaluate a rapid and reliable assay. In the present study, a loop-mediated isothermal amplification (LAMP assay based on the cytochrome b gene of the mitochondrial DNA of the species Struthio camelus was developed. The LAMP assay was used in combination with a real-time fluorometer. The developed system allowed the detection of 0.01% ostrich meat products. In parallel, a direct swab method without nucleic acid extraction using the HYPLEX LPTV buffer was also evaluated. This rapid processing method allowed detection of ostrich meat without major incubation steps. In summary, the LAMP assay had excellent sensitivity and specificity for detecting ostrich meat and could provide a sampling-to-result identification-time of 15 to 20 minutes.

Rapid identification and differentiation of Fasciola hepatica and Fasciola gigantica by a loop-mediated isothermal amplification (LAMP) assay.

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Ai, L; Li, C; Elsheikha, H M; Hong, S J; Chen, J X; Chen, S H; Li, X; Cai, X Q; Chen, M X; Zhu, X Q

2010-12-15

The present study developed and validated a species-specific loop-mediated isothermal amplification (LAMP) assay for the rapid detection and discrimination of Fasciola hepatica and Fasciola gigantica. The LAMP assay is inexpensive, easy to perform and shows rapid reaction, wherein the amplification can be obtained in 45 min under isothermal conditions of 61 °C or 62 °C by employing a set of four species-specific primer mixtures and results can be checked through naked-eye visualization. The optimal assay conditions with no cross-reaction with other closely related trematodes (Clonorchis sinensis, Opisthorchis viverrini, Orientobilharzia turkestanicum and Schistosoma japonicum) as well as within the two Fasciola species were established. The assay was validated by examining F. gigantica DNA in the intermediate host snails and in faecal samples. The results indicated that the LAMP assay is approximately 10(4) times more sensitive than the conventional specific PCR assays. These findings indicate that this Fasciola species-specific LAMP assay may have a potential clinical application for detection and differentiation of Fasciola species, especially in endemic countries. Copyright © 2010 Elsevier B.V. All rights reserved.

Rapid and sensitive detection of Bordetella bronchiseptica by loop-mediated isothermal amplification (LAMP

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Hui Zhang

2013-10-01

Full Text Available Bordetella bronchiseptica causes acute and chronic respiratory infections in diverse animal species and occasionally in humans. In this study, we described the establishment of a simple, sensitive and cost-efficient loop-mediated isothermal amplification (LAMP assay for the detection of B. bronchiseptica. A set of primers towards a 235 bp region within the flagellum gene of B. bronchiseptica was designed with online software.. The specificity of the LAMP assay was examined by using 6 porcine pathogens and 100 nasal swabs collected from healthy pigs and suspect infected pigs. The results indicated that positive reactions were confirmed for all B. bronchiseptica and no cross-reactivity was observed from other non-B. bronchiseptica. In sensitivity evaluations, the technique successfully detected a serial dilutions of extracted B. bronchiseptica DNA with a detection limit of 9 copies, which was 10 times more sensitive than that of PCR. Compared with conventional PCR, the higher sensitivity of LAMP method and no need for the complex instrumentation make this LAMP assay a promising alternative for the diagnosis of B. bronchiseptica in rural areas and developing countries where there lacks of complex laboratory services.

Rapid authentication of the precious herb saffron by loop-mediated isothermal amplification (LAMP) based on internal transcribed spacer 2 (ITS2) sequence.

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Zhao, Mingming; Shi, Yuhua; Wu, Lan; Guo, Licheng; Liu, Wei; Xiong, Chao; Yan, Song; Sun, Wei; Chen, Shilin

2016-05-05

Saffron is one of the most expensive species of Chinese herbs and has been subjected to various types of adulteration because of its high price and limited production. The present study introduces a loop-mediated isothermal amplification (LAMP) technique for the differentiation of saffron from its adulterants. This novel technique is sensitive, efficient and simple. Six specific LAMP primers were designed on the basis of the nucleotide sequence of the internal transcribed spacer 2 (ITS2) nuclear ribosomal DNA of Crocus sativus. All LAMP amplifications were performed successfully, and visual detection occurred within 60 min at isothermal conditions of 65 °C. The results indicated that the LAMP primers are accurate and highly specific for the discrimination of saffron from its adulterants. In particular, 10 fg of genomic DNA was determined to be the limit for template accuracy of LAMP in saffron. Thus, the proposed novel, simple, and sensitive LAMP assay is well suited for immediate on-site discrimination of herbal materials. Based on the study, a practical standard operating procedure (SOP) for utilizing the LAMP protocol for herbal authentication is provided.

Development of a highly sensitive loop-mediated isothermal amplification (LAMP) method for the detection of Loa loa.

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Fernández-Soto, Pedro; Mvoulouga, Prosper Obolo; Akue, Jean Paul; Abán, Julio López; Santiago, Belén Vicente; Sánchez, Miguel Cordero; Muro, Antonio

2014-01-01

The filarial parasite Loa loa, the causative agent of loiasis, is endemic in Central and Western Africa infecting 3-13 million people. L. loa has been associated with fatal encephalopathic reactions in high Loa-infected individuals receiving ivermectin during mass drug administration programs for the control of onchocerciasis and lymphatic filariasis. In endemic areas, the only diagnostic method routinely used is the microscopic examination of mid-day blood samples by thick blood film. Improved methods for detection of L. loa are needed in endemic regions with limited resources, where delayed diagnosis results in high mortality. We have investigated the use of a loop-mediated isothermal amplification (LAMP) assay to facilitate rapid, inexpensive, molecular diagnosis of loiasis. Primers for LAMP were designed from a species-specific repetitive DNA sequence from L. loa retrieved from GenBank. Genomic DNA of a L. loa adult worm was used to optimize the LAMP conditions using a thermocycler or a conventional heating block. Amplification of DNA in the LAMP mixture was visually inspected for turbidity as well as addition of fluorescent dye. LAMP specificity was evaluated using DNA from other parasites; sensitivity was evaluated using DNA from L. loa 10-fold serially diluted. Simulated human blood samples spiked with DNA from L. loa were also tested for sensitivity. Upon addition of fluorescent dye, all positive reactions turned green while the negative controls remained orange under ambient light. After electrophoresis on agarose gels, a ladder of multiple bands of different sizes could be observed in positive samples. The detection limit of the assay was found to be as little as 0.5 ag of L. loa genomic DNA when using a heating block. We have designed, for the first time, a highly sensitive LAMP assay for the detection of L. loa which is potentially adaptable for field diagnosis and disease surveillance in loiasis-endemic areas.

Loop-mediated isothermal amplification (LAMP): early detection of Toxoplasma gondii infection in mice.

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Kong, Qing-Ming; Lu, Shao-Hong; Tong, Qun-Bo; Lou, Di; Chen, Rui; Zheng, Bin; Kumagai, Takashi; Wen, Li-Yong; Ohta, Nobuo; Zhou, Xiao-Nong

2012-01-03

Toxoplasmosis is a widespread zoonotic parasitic disease that occurs in both animals and humans. Traditional molecular assays are often difficult to perform, especially for the early diagnosis of Toxoplasma gondii infections. Here, we established a novel loop-mediated isothermal amplification targeting the 529 bp repeat element (529 bp-LAMP) to detect T. gondii DNA in blood samples of experimental mice infected with tachyzoites of the RH strain. The assay was performed with Bst DNA polymerase at 65°C for 1 h. The detection limit of the 529 bp-LAMP assay was as low as 0.6 fg of T. gondii DNA. The sensitivity of this assay was 100 and 1000 fold higher than that of the LAMP targeting B1 gene (B1-LAMP) and nested PCR targeting 529 bp repeat element (529 bp-nested PCR), respectively. The specificity of the 529 bp-LAMP assay was determined using the DNA samples of Trypanosoma evansi, Plasmodium falciparum, Paragonimus westermani, Schistosoma japonicum, Fasciola hepatica and Angiostrongylus cantonensis. No cross-reactivity with the DNA of any parasites was found. The assay was able to detect T. gondii DNA in all mouse blood samples at one day post infection (dpi). We report the following findings: (i) The detection limit of the 529 bp-LAMP assay is 0.6 fg of T. gondii DNA; (ii) The assay does not involve any cross-reactivity with the DNA of other parasites; (iii) This is the first report on the application of the LAMP assay for early diagnosis of toxoplasmosis in blood samples from experimentally infected mice. Due to its simplicity, sensitivity and cost-effectiveness for common use, we suggest that this assay should be used as an early diagnostic tool for health control of toxoplasmosis.

Loop-mediated isothermal amplification (LAMP): Early detection of Toxoplasma gondii infection in mice

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Kong, Qing-Ming; Lu, Shao-Hong; Tong, Qun-Bo; Lou, Di; Chen, Rui; Zheng, Bin; Kumagai, Takashi; Wen, Li-Yong; Ohta, Nobuo; Zhou, Xiao-Nong

2012-01-01

Abstract Background Toxoplasmosis is a widespread zoonotic parasitic disease that occurs in both animals and humans. Traditional molecular assays are often difficult to perform, especially for the early diagnosis of Toxoplasma gondii infections. Here, we established a novel loop-mediated isothermal amplification targeting the 529 bp repeat element (529 bp-LAMP) to detect T. gondii DNA in blood samples of experimental mice infected with tachyzoites of the RH strain. Findings The assay was perf...

Development of a loop-mediated isothermal amplification (LAMP) assay for rapid and specific detection of common genetically modified organisms (GMOs).

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Feng, Jiawang; Tang, Shiming; Liu, Lideng; Kuang, Xiaoshan; Wang, Xiaoyu; Hu, Songnan; You, Shuzhu

2015-03-01

Here, we developed a loop-mediated isothermal amplification (LAMP) assay for 11 common transgenic target DNA in GMOs. Six sets of LAMP primer candidates for each target were designed and their specificity, sensitivity, and reproductivity were evaluated. With the optimized LAMP primers, this LAMP assay was simply run within 45-60 min to detect all these targets in GMOs tested. The sensitivity, specificity, and reproductivity of the LAMP assay were further analyzed in comparison with those of Real-Time PCR. In consistent with real-time PCR, detection of 0.5% GMOs in equivalent background DNA was possible using this LAMP assay for all targets. In comparison with real-time PCR, the LAMP assay showed the same results with simple instruments. Hence, the LAMP assay developed can provide a rapid and simple approach for routine screening as well as specific events detection of many GMOs.

Evaluation of an Internally Controlled Multiplex Tth Endonuclease Cleavage Loop-Mediated Isothermal Amplification (TEC-LAMP Assay for the Detection of Bacterial Meningitis Pathogens

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Owen Higgins

2018-02-01

Full Text Available Bacterial meningitis infection is a leading global health concern for which rapid and accurate diagnosis is essential to reduce associated morbidity and mortality. Loop-mediated isothermal amplification (LAMP offers an effective low-cost diagnostic approach; however, multiplex LAMP is difficult to achieve, limiting its application. We have developed novel real-time multiplex LAMP technology, TEC-LAMP, using Tth endonuclease IV and a unique LAMP primer/probe. This study evaluates the analytical specificity, limit of detection (LOD and clinical application of an internally controlled multiplex TEC-LAMP assay for detection of leading bacterial meningitis pathogens: Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae. Analytical specificities were established by testing 168 bacterial strains, and LODs were determined using Probit analysis. The TEC-LAMP assay was 100% specific, with LODs for S. pneumoniae, N. meningitidis and H. influenzae of 39.5, 17.3 and 25.9 genome copies per reaction, respectively. Clinical performance was evaluated by testing 65 archived PCR-positive samples. Compared to singleplex real-time PCR, the multiplex TEC-LAMP assay demonstrated diagnostic sensitivity and specificity of 92.3% and 100%, respectively. This is the first report of a single-tube internally controlled multiplex LAMP assay for bacterial meningitis pathogen detection, and the first report of Tth endonuclease IV incorporation into nucleic acid amplification diagnostic technology.

Evaluation of an Internally Controlled Multiplex Tth Endonuclease Cleavage Loop-Mediated Isothermal Amplification (TEC-LAMP) Assay for the Detection of Bacterial Meningitis Pathogens

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Clancy, Eoin; Cormican, Martin; Boo, Teck Wee; Cunney, Robert

2018-01-01

Bacterial meningitis infection is a leading global health concern for which rapid and accurate diagnosis is essential to reduce associated morbidity and mortality. Loop-mediated isothermal amplification (LAMP) offers an effective low-cost diagnostic approach; however, multiplex LAMP is difficult to achieve, limiting its application. We have developed novel real-time multiplex LAMP technology, TEC-LAMP, using Tth endonuclease IV and a unique LAMP primer/probe. This study evaluates the analytical specificity, limit of detection (LOD) and clinical application of an internally controlled multiplex TEC-LAMP assay for detection of leading bacterial meningitis pathogens: Streptococcus pneumoniae, Neisseria meningitidis and Haemophilus influenzae. Analytical specificities were established by testing 168 bacterial strains, and LODs were determined using Probit analysis. The TEC-LAMP assay was 100% specific, with LODs for S. pneumoniae, N. meningitidis and H. influenzae of 39.5, 17.3 and 25.9 genome copies per reaction, respectively. Clinical performance was evaluated by testing 65 archived PCR-positive samples. Compared to singleplex real-time PCR, the multiplex TEC-LAMP assay demonstrated diagnostic sensitivity and specificity of 92.3% and 100%, respectively. This is the first report of a single-tube internally controlled multiplex LAMP assay for bacterial meningitis pathogen detection, and the first report of Tth endonuclease IV incorporation into nucleic acid amplification diagnostic technology. PMID:29425124

Loop-mediated isothermal amplification (LAMP) test for specific and rapid detection of Brucella abortus in cattle.

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Karthik, K; Rathore, Rajesh; Thomas, Prasad; Arun, T R; Viswas, K N; Agarwal, R K; Manjunathachar, H V; Dhama, Kuldeep

2014-01-01

Brucella abortus, the major causative agent of abortion in cattle and a zoonotic pathogen, needs to be diagnosed at an early stage. Loop-mediated isothermal amplification (LAMP) test is easy to perform and also promising to be adapted at field level. To develop a LAMP assay for specific and rapid detection of B. abortus from clinical samples of cattle. LAMP primers were designed targeting BruAb2_0168 region using specific software tool and LAMP was optimized. The developed LAMP was tested for its specificity with 3 Brucella spp. and 11 other non-Brucella spp. Sensitivity of the developed LAMP was also carried out with known quantity of DNA. Cattle whole blood samples and aborted fetal stomach contents were collected and used for testing with developed LAMP assay and results were compared with polymerase chain reaction (PCR). The developed LAMP assay works at 61 °C for 60 min and the detection limit was observed to be 100-fold more than the conventional PCR that is commonly used for diagnosis of B. abortus. Clinical sensitivity and specificity of the developed LAMP assay was 100% when compared with Rose Bengal plate test and standard tube agglutination test. SYB® green dye I was used to visualize the result with naked eye. The novelty of the developed LAMP assay for specifically detecting B. abortus infection in cattle along with its inherent rapidness and high sensitivity can be employed for detecting this economically important pathogen of cattle at field level as well be exploited for screening of human infections.

Improved detection limit in rapid detection of human enterovirus 71 and coxsackievirus A16 by a novel reverse transcription-isothermal multiple-self-matching-initiated amplification assay.

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Ding, Xiong; Nie, Kai; Shi, Lei; Zhang, Yong; Guan, Li; Zhang, Dan; Qi, Shunxiang; Ma, Xuejun

2014-06-01

Rapid detection of human enterovirus 71 (EV71) and coxsackievirus A16 (CVA16) is important in the early phase of hand-foot-and-mouth disease (HFMD). In this study, we developed and evaluated a novel reverse transcription-isothermal multiple-self-matching-initiated amplification (RT-IMSA) assay for the rapid detection of EV71 and CVA16 by use of reverse transcriptase, together with a strand displacement DNA polymerase. Real-time RT-IMSA assays using a turbidimeter and visual RT-IMSA assays to detect EV71 and CVA16 were established and completed in 1 h, and the reported corresponding real-time reverse transcription-loop-mediated isothermal amplification (RT-LAMP) assays targeting the same regions of the VP1 gene were adopted as parallel tests. Through testing VP1 RNAs transcribed in vitro, the real-time RT-IMSA assays exhibited better linearity of quantification, with R(2) values of 0.952 (for EV71) and 0.967 (for CVA16), than the real-time RT-LAMP assays, which had R(2) values of 0.803 (for EV71) and 0.904 (for CVA16). Additionally, the detection limits of the real-time RT-IMSA assays (approximately 937 for EV71 and 67 for CVA16 copies/reaction) were higher than those of real-time RT-LAMP assays (approximately 3,266 for EV71 and 430 for CVA16 copies/reaction), and similar results were observed in the visual RT-IMSA assays. The new approaches also possess high specificities for the corresponding targets, with no cross-reactivity observed. In clinical assessment, compared to commercial reverse transcription-quantitative PCR (qRT-PCR) kits, the diagnostic sensitivities of the real-time RT-IMSA assays (96.4% for EV71 and 94.6% for CVA16) were higher than those of the real-time RT-LAMP assays (91.1% for EV71 and 90.8% for CVA16). The visual RT-IMSA assays also exhibited the same results. In conclusion, this proof-of-concept study suggests that the novel RT-IMSA assay is superior to the RT-LAMP assay in terms of detection limit and has the potential to rapidly detect EV71

The Development and Evaluation of a Loop-Mediated Isothermal Amplification Method for the Rapid Detection of Salmonella enterica serovar Typhi.

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Fenxia Fan

Full Text Available Typhoid fever remains a public health threat in many countries. A positive result in traditional culture is a gold-standard for typhoid diagnosis, but this method is time consuming and not sensitive enough for detection of samples containing a low copy number of the target organism. The availability of the loop-mediated isothermal amplification (LAMP assay, which offers high speed and simplicity in detection of specific targets, has vastly improved the diagnosis of numerous infectious diseases. However, little research efforts have been made on utilizing this approach for diagnosis of Salmonella enterica serovar Typhi by targeting a single and specific gene. In this study, a LAMP assay for rapid detection of S. Typhi based on a novel marker gene, termed STY2879-LAMP, was established and evaluated with real-time PCR (RT-PCR. The specificity tests showed that STY2879 could be amplified in all S. Typhi strains isolated in different years and regions in China, whereas no amplification was observable in non-typhoidal strains covering 34 Salmonella serotypes and other pathogens causing febrile illness. The detection limit of STY2879-LAMP for S. Typhi was 15 copies/reaction in reference plasmids, 200 CFU/g with simple heat-treatment of DNA extracted from simulated stool samples and 20 CFU/ml with DNA extracted from simulated blood samples, which was 10 fold more sensitive than the parallel RT-PCR control experiment. Furthermore, the sensitivity of STY2879-LAMP and RT-PCR combining the traditional culture enrichment method for simulated stool and blood spiked with lower S. Typhi count during the 10 h enrichment time was also determined. In comparison with LAMP, the positive reaction time for RT-PCR required additional 2-3 h enrichment time for either simulated stool or blood specimens. Therefore, STY2879-LAMP is of practical value in the clinical settings and has a good potential for application in developing regions due to its easy-to-use protocol.

Simplified Real-Time Multiplex Detection of Loop-Mediated Isothermal Amplification Using Novel Mediator Displacement Probes with Universal Reporters.

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Becherer, Lisa; Bakheit, Mohammed; Frischmann, Sieghard; Stinco, Silvina; Borst, Nadine; Zengerle, Roland; von Stetten, Felix

2018-04-03

A variety of real-time detection techniques for loop-mediated isothermal amplification (LAMP) based on the change in fluorescence intensity during DNA amplification enable simultaneous detection of multiple targets. However, these techniques depend on fluorogenic probes containing target-specific sequences. That complicates the adaption to different targets leading to time-consuming assay optimization. Here, we present the first universal real-time detection technique for multiplex LAMP. The novel approach allows simple assay design and is easy to implement for various targets. The innovation features a mediator displacement probe and a universal reporter. During amplification of target DNA the mediator is displaced from the mediator displacement probe. Then it hybridizes to the reporter generating a fluorescence signal. The novel mediator displacement (MD) detection was validated against state-of-the-art molecular beacon (MB) detection by means of a HIV-1 RT-LAMP: MD surpassed MB detection by accelerated probe design (MD: 10 min, MB: 3-4 h), shorter times to positive (MD 4.1 ± 0.1 min shorter than MB, n = 36), improved signal-to-noise fluorescence ratio (MD: 5.9 ± 0.4, MB: 2.7 ± 0.4; n = 15), and showed equally good or better analytical performance parameters. The usability of one universal mediator-reporter set in different multiplex assays was successfully demonstrated for a biplex RT-LAMP of HIV-1 and HTLV-1 and a biplex LAMP of Haemophilus ducreyi and Treponema pallidum, both showing good correlation between target concentration and time to positive. Due to its simple implementation it is suggested to extend the use of the universal mediator-reporter sets to the detection of various other diagnostic panels.

Detection of Brucellosis in Sika Deer ( Cervus nippon ) through Loop-mediated Isothermal Amplification (LAMP).

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Liu, Qianhong; Wei, Jie; Sun, Qingsong; Wang, Ben; Wang, Yuting; Hu, Ying; Wu, Wenrong

2017-07-01

Brucellosis (Brucella bovis) in sika deer ( Cervus nippon ) can cause enormous losses to stag breeding, especially in areas in which stag breeding has become an important industry. It also poses a threat to humans because it is a zoonotic disease. Use of the loop-mediated isothermal amplification (LAMP) assay has been poorly described in the diagnosis of brucellosis in deer. We developed a LAMP assay targeting the omp25 gene sequence to detect brucellosis in sika deer. The reaction can be completed in 60 min at 63 C and, with a detection limit of 17 pg, it was more sensitive than conventional PCR, with its detection limit of 1.7 ng. No cross-reactivity was observed with four bacteria: Escherichia coli , Salmonella enterica subsp. enterica, Clostridium pasteurianum , and Pseudomonas aeruginosa . We used 263 samples of blood to evaluate the reaction. The percentage of agreement between LAMP and PCR reached 91%; relative specificity reached 87%, and relative sensitivity reached 100%. The results indicate LAMP can be a simple and rapid diagnostic tool for detecting brucellosis in sika deer, particularly in the field, where it is essential to control brucellosis in deer with a rapid and accurate diagnosis for removal of positive animals.

Comparison of reverse transcriptase PCR, reverse transcriptase loop-mediated isothermal amplification, and culture-based assays for Salmonella detection from pork processing environments.

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Techathuvanan, Chayapa; Draughon, Frances Ann; D'Souza, Doris Helen

2011-02-01

Novel rapid Salmonella detection assays without the need for sophisticated equipment or labor remain in high demand. Real-time reverse transcriptase PCR (RT-PCR) assays, though rapid and sensitive, require expensive thermocyclers, while a novel RT loop-mediated isothermal amplification (RT-LAMP) method requires only a simple water bath. Our objective was to compare the detection sensitivity of Salmonella Typhimurium from the pork processing environment by RT-LAMP, RT-PCR, and culture-based assays. Carcass and surface swabs and carcass rinses were obtained from a local processing plant. Autoclaved carcass rinses (500 ml) were spiked with Salmonella Typhimurium and filtered. Filters were placed in stomacher bags containing tetrathionate broth (TTB) and analyzed with or without 10-h enrichment at 37 °C. Natural swabs were stomached with buffered peptone water, and natural carcass rinses were filtered, preenriched, and further enriched in TTB. Serially-diluted enriched samples were enumerated by spread plating on xylose lysine Tergitol 4 agar. RNA was extracted from 5 ml of enriched TTB with TRIzol. RT-LAMP assay using previously described invA primers was conducted at 62 °C for 90 min in a water bath with visual detection and by gel electrophoresis. SYBR Green I-based-real-time RT-PCR was carried out with invA primers followed by melt temperature analysis. The results of RT-LAMP detection for spiked carcass rinses were comparable to those of RT-PCR and cultural plating, with detection limits of 1 log CFU/ml, although they were obtained significantly faster, within 24 h including preenrichment and enrichment. RT-LAMP showed 4 of 12 rinse samples positive, while RT-PCR showed 1 of 12 rinse samples positive. For swabs, 6 of 27 samples positive by RT-LAMP and 5 of 27 by RT-PCR were obtained. This 1-day RT-LAMP assay shows promise for routine Salmonella screening by the pork industry. Copyright ©, International Association for Food Protection

Development of Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Cannabis sativa.

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Kitamura, Masashi; Aragane, Masako; Nakamura, Kou; Watanabe, Kazuhito; Sasaki, Yohei

2016-07-01

In many parts of the world, the possession and cultivation of Cannabis sativa L. are restricted by law. As chemical or morphological analyses cannot identify the plant in some cases, a simple yet accurate DNA-based method for identifying C. sativa is desired. We have developed a loop-mediated isothermal amplification (LAMP) assay for the rapid identification of C. sativa. By optimizing the conditions for the LAMP reaction that targets a highly conserved region of tetrahydrocannabinolic acid (THCA) synthase gene, C. sativa was identified within 50 min at 60-66°C. The detection limit was the same as or higher than that of conventional PCR. The LAMP assay detected all 21 specimens of C. sativa, showing high specificity. Using a simple protocol, the identification of C. sativa could be accomplished within 90 min from sample treatment to detection without use of special equipment. A rapid, sensitive, highly specific, and convenient method for detecting and identifying C. sativa has been developed and is applicable to forensic investigations and industrial quality control.

Establishment of reverse transcription loop-mediated isothermal amplification for rapid detection and differentiation of canine distemper virus infected and vaccinated animals.

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Liu, Da-Fei; Liu, Chun-Guo; Tian, Jin; Jiang, Yi-Tong; Zhang, Xiao-Zhan; Chai, Hong-Liang; Yang, Tian-Kuo; Yin, Xiu-Chen; Zhang, Hong-Ying; Liu, Ming; Hua, Yu-Ping; Qu, Lian-Dong

2015-06-01

Although widespread vaccination against canine distemper virus (CDV) has been conducted for many decades, several canine distemper outbreaks in vaccinated animals have been reported frequently. In order to detect and differentiate the wild-type and vaccine strains of the CDV from the vaccinated animals, a novel reverse transcription loop-mediated isothermal amplification (RT-LAMP) method was developed. A set of four primers-two internal and two external-were designed to target the H gene for the specific detection of wild-type CDV variants. The CDV-H RT-LAMP assay rapidly amplified the target gene, within 60 min, using a water bath held at a constant temperature of 65°C. The assay was 100-fold more sensitive than conventional RT-PCR, with a detection limit of 10(-1)TCID50ml(-1). The system showed a preference for wild-type CDV, and exhibited less sensitivity to canine parvovirus, canine adenovirus type 1 and type 2, canine coronavirus, and canine parainfluenza virus. The assay was validated using 102 clinical samples obtained from vaccinated dog farms, and the results were comparable to a multiplex nested RT-PCR assay. The specific CDV-H RT-LAMP assay provides a simple, rapid, and sensitive tool for the detection of canines infected with wild-type CDV from canines vaccinated with attenuated vaccine. Copyright © 2015 Elsevier B.V. All rights reserved.

Development of Loop-Mediated Isothermal Amplification (LAMP) assay for rapid detection of Fusarium oxysporum f. sp. ciceris - wilt pathogen of chickpea.

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Ghosh, Raju; Nagavardhini, Avuthu; Sengupta, Anindita; Sharma, Mamta

2015-02-11

Fusarium oxysporum f. sp. ciceris (Foc), the causal agent of Fusarium wilt is a devastating pathogen of chickpea. In chickpea, various soil borne pathogens produce (s) similar symptoms, therefore cannot be distinguished easily at field level. There is real need for a rapid, inexpensive, and easy to operate and maintain genotyping tool to facilitate accurate disease diagnosis and surveillance for better management of Fusarium wilt outbreaks. In this study, we developed a loop-mediated isothermal amplification (LAMP) assay targeting the elongation factor 1 alpha gene sequence for visual detection of Foc. The LAMP reaction was optimal at 63°C for 60 min. When hydroxynaphthol blue (HNB) was added before amplification, samples with Foc DNA developed a characteristic sky blue colour but those without DNA or with the DNA of six other plant pathogenic fungi did not. Results obtained with LAMP and HNB were confirmed when LAMP products were subjected to gel electrophoresis. The detection limit of this LAMP assay for Foc was 10 fg of genomic DNA per reaction, while that of conventional PCR was 100 pg. In conclusion, it was found that a LAMP assay combined with HNB is simple, rapid, sensitive, and specific. The LAMP assay does not require specialized equipment, hence can be used in the field for the rapid detection of Foc. This is the first report of the use of LAMP assay for the detection of Foc. The presented LAMP method provides a specific, sensitive and rapid diagnostic tool for the distinction of Foc, with the potential to be standardized as a detection method for Foc in endemic areas and will be very useful for monitoring the disease complex in the field further suggesting the management strategies.

Detection of influenza viruses by coupling multiplex reverse-transcription loop-mediated isothermal amplification with cascade invasive reaction using nanoparticles as a sensor

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Ge Y

2017-04-01

Full Text Available Yiyue Ge,1 Qiang Zhou,2 Kangchen Zhao,1 Ying Chi,1 Bin Liu,3 Xiaoyan Min,4 Zhiyang Shi,1 Bingjie Zou,2 Lunbiao Cui1 1Institute of Pathogenic Microbiology, Key Laboratories of Enteric Pathogenic Microbiology (Ministry of Health, Jiangsu Provincial Center for Disease Control and Prevention, 2Department of Pharmacology, Jinling Hospital, Medical School of Nanjing University, 3Department of Biomedical Engineering, Nanjing Medical University, 4Department of Geriatrics, The First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu, People’s Republic of China Abstract: Influenza virus infections represent a worldwide public health and economic problem due to the significant morbidity and mortality caused by seasonal epidemics and pandemics. Sensitive and convenient methodologies for detection of influenza viruses are essential for further disease control. Loop-mediated isothermal amplification (LAMP is the most commonly used method of nucleic acid isothermal amplification. However, with regard to multiplex LAMP, differentiating the ladder-like LAMP products derived from multiple targets is still challenging today. The requirement of specialized instruments has further hindered the on-site application of multiplex LAMP. We have developed an integrated assay coupling multiplex reverse transcription LAMP with cascade invasive reaction using nanoparticles (mRT-LAMP-CIRN as a sensor for the detection of three subtypes of influenza viruses: A/H1N1pdm09, A/H3 and influenza B. The analytic sensitivities of the mRT-LAMP-CIRN assay were 101 copies of RNA for both A/H1N1pdm09 and A/H3, and 102 copies of RNA for influenza B. This assay demonstrated highly specific detection of target viruses and could differentiate them from other genetically or clinically related viruses. Clinical specimen analysis showed the mRT-LAMP-CIRN assay had an overall sensitivity and specificity of 98.3% and 100%, respectively. In summary, the mRT-LAMP-CIRN assay is

Rapid and sensitive detection of novel avian-origin influenza A (H7N9 virus by reverse transcription loop-mediated isothermal amplification combined with a lateral-flow device.

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Yiyue Ge

Full Text Available A severe disease in humans caused by a novel avian-origin influenza A (H7N9 virus emerged in China recently, which has caused at least 128 cases and 26 deaths. Rapid detection of the novel H7N9 virus is urgently needed to differentiate the disease from other infections, and to facilitate infection control as well as epidemiologic investigations. In this study, a reverse transcription loop-mediated isothermal amplification combined with a lateral flow device (RT-LAMP-LFD assay to rapidly detect H7N9 virus was developed and evaluated. The RT-LAMP primers were designed to target the haemagglutinin (HA and neuraminidase (NA genes of H7N9 virus. Results of 10-fold dilution series assays showed that analysis of RT-LAMP products by the LFD method was as sensitive as real-time turbidity detection, and that the analytic sensitivities of the HA and NA RT-LAMP assays were both 10 copies of synthetic RNA. Furthermore, both the assays showed 100% clinical specificity for identification of H7N9 virus. The performance characteristics of the RT-LAMP-LFD assay were evaluated with 80 clinical specimens collected from suspected H7N9 patients. The NA RT-LAMP-LFD assay was more sensitive than real time RT-PCR assay. Compared with a combination of virus culture and real-time RT-PCR, the sensitivity, specificity, positive predictive value, and negative predictive value of the RT-LAMP-LFD assay were all 100%. Overall, The RT-LAMP-LFD assay established in this study can be used as a reliable method for early diagnosis of the avian-origin influenza A (H7N9 virus infection.

Loop-mediated Isothermal Amplification Assay to Rapidly Detect Wheat Streak Mosaic Virus in Quarantined Plants

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Siwon Lee

2015-12-01

Full Text Available We developed a loop-mediated isothermal amplification (LAMP method to rapidly diagnose Wheat streak mosaic virus (WSMV during quarantine inspections of imported wheat, corn, oats, and millet. The LAMP method was developed as a plant quarantine inspection method for the first time, and its simplicity, quickness, specificity and sensitivity were verified compared to current reverse transcription-polymerase chain reaction (RT-PCR and nested PCR quarantine methods. We were able to quickly screen for WSMV at quarantine sites with many test samples; thus, this method is expected to contribute to plant quarantine inspections.

Preliminary validation of direct detection of foot-and-mouth disease virus within clinical samples using reverse transcription loop-mediated isothermal amplification coupled with a simple lateral flow device for detection.

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Ryan A Waters

Full Text Available Rapid, field-based diagnostic assays are desirable tools for the control of foot-and-mouth disease (FMD. Current approaches involve either; 1 Detection of FMD virus (FMDV with immuochromatographic antigen lateral flow devices (LFD, which have relatively low analytical sensitivity, or 2 portable RT-qPCR that has high analytical sensitivity but is expensive. Loop-mediated isothermal amplification (LAMP may provide a platform upon which to develop field based assays without these drawbacks. The objective of this study was to modify an FMDV-specific reverse transcription-LAMP (RT-LAMP assay to enable detection of dual-labelled LAMP products with an LFD, and to evaluate simple sample processing protocols without nucleic acid extraction. The limit of detection of this assay was demonstrated to be equivalent to that of a laboratory based real-time RT-qPCR assay and to have a 10,000 fold higher analytical sensitivity than the FMDV-specific antigen LFD currently used in the field. Importantly, this study demonstrated that FMDV RNA could be detected from epithelial suspensions without the need for prior RNA extraction, utilising a rudimentary heat source for amplification. Once optimised, this RT-LAMP-LFD protocol was able to detect multiple serotypes from field epithelial samples, in addition to detecting FMDV in the air surrounding infected cattle, pigs and sheep, including pre-clinical detection. This study describes the development and evaluation of an assay format, which may be used as a future basis for rapid and low cost detection of FMDV. In addition it provides providing "proof of concept" for the future use of LAMP assays to tackle other challenging diagnostic scenarios encompassing veterinary and human health.

Application of a Real-time Reverse Transcription Loop Mediated Amplification Method to the Detection of Rabies Virus in Arctic Foxes in Greenland

DEFF Research Database (Denmark)

Wakeley, Philip; Johnson, Nicholas; Rasmussen, Thomas Bruun

Reverse transcription loop mediated amplification (RT-LAMP) offers a rapid, isothermal method for amplification of virus RNA. In this study a panel of positive rabies virus samples originally prepared from arctic fox brain tissue was assessed for the presence of rabies viral RNA using a real time...... RT-LAMP. The method had previously been shown to work with samples from Ghana which clustered with cosmopolitan lineage rabies viruses but the assay had not been assessed using samples from animals infected with rabies from the arctic region. The assay is designed to amplify both cosmopolitan strains...... and arctic-like strains of classical rabies virus due to the primer design and is therefore expected to be universally applicable independent of region of the world where the virus is isolated. Of the samples tested all were found to be positive after incubation for 25 to 30 minutes. The method made use...

Colorimetric tests for diagnosis of filarial infection and vector surveillance using non-instrumented nucleic acid loop-mediated isothermal amplification (NINA-LAMP.

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Catherine B Poole

Full Text Available Accurate detection of filarial parasites in humans is essential for the implementation and evaluation of mass drug administration programs to control onchocerciasis and lymphatic filariasis. Determining the infection levels in vector populations is also important for assessing transmission, deciding when drug treatments may be terminated and for monitoring recrudescence. Immunological methods to detect infection in humans are available, however, cross-reactivity issues have been reported. Nucleic acid-based molecular assays offer high levels of specificity and sensitivity, and can be used to detect infection in both humans and vectors. In this study we developed loop-mediated isothermal amplification (LAMP tests to detect three different filarial DNAs in human and insect samples using pH sensitive dyes for enhanced visual detection of amplification. Furthermore, reactions were performed in a portable, non-instrumented nucleic acid amplification (NINA device that provides a stable heat source for LAMP. The efficacy of several strand displacing DNA polymerases were evaluated in combination with neutral red or phenol red dyes. Colorimetric NINA-LAMP assays targeting Brugia Hha I repeat, Onchocerca volvulus GST1a and Wuchereria bancrofti LDR each exhibit species-specificity and are also highly sensitive, detecting DNA equivalent to 1/10-1/5000th of one microfilaria. Reaction times varied depending on whether a single copy gene (70 minutes, O. volvulus or repetitive DNA (40 min, B. malayi and W. bancrofti was employed as a biomarker. The NINA heater can be used to detect multiple infections simultaneously. The accuracy, simplicity and versatility of the technology suggests that colorimetric NINA-LAMP assays are ideally suited for monitoring the success of filariasis control programs.

Preliminary application and evaluation of loop-mediated isothermal amplification (LAMP for detection of bovine theileriosis and trypanosomosis in Tanzania : research communication

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O.M.M. Thekisoe

2007-09-01

Full Text Available The sensitivity of LAMP, PCR and microscopy to detect Theileria spp. and Trypanosoma congolense in field-derived bovine blood samples from Tanzania was evaluated and compared. No parasites were detected by microscopy. Furthermore, no bovine Theileria spp. were detected by LAMP and PCR from all the 24 samples collected from Arusha. Four and one out of 24 samples were positive for Theileria congolense infection by LAMP and PCR respectively while, 18 and nine out of 40 samples from Dar es Salaam were positive by LAMP and PCR for Theileria spp. Infection, respectively. Although all samples from Dar es Salaam were negative for Trypanosoma congolense infections by PCR, 12 out of 40 samples were LAMP positive. Whilst PCR is an established gene amplification method for the detection of Theileria and trypanosome parasites, this study introduces LAMP as an alternative molecular diagnostic tool that could be used in large-scale epidemiological surveys.

Detection of foot-and-mouth disease virus rna by reverse transcription loop-mediated isothermal amplification

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Chen Hao-tai

2011-11-01

Full Text Available Abstract A reverse transcription loop-mediated isothermal amplification (RT-LAMP assay was developed for foot-and-mouth disease virus (FMDV RNA. The amplification was able to finish in 45 min under isothermal condition at 64°C by employing a set of four primers targeting FMDV 2B. The assay showed higher sensitivity than RT-PCR. No cross reactivity was observed from other RNA viruses including classical swine fever virus, swine vesicular disease, porcine reproductive and respiratory syndrome virus, Japanese encephalitis virus. Furthermore, the assay correctly detected 84 FMDV positive samples but not 65 FMDV negative specimens. The result indicated the potential usefulness of the technique as a simple and rapid procedure for the detection of FMDV infection.

Loop-mediated isothermal amplification (LAMP) assays for rapid detection and differentiation of Nosema apis and N. ceranae in honeybees.

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Ptaszyńska, Aneta A; Borsuk, Grzegorz; Woźniakowski, Grzegorz; Gnat, Sebastian; Małek, Wanda

2014-08-01

Nosemosis is a contagious disease of honeybees (Apis mellifera) manifested by increased winter mortality, poor spring build-up and even the total extinction of infected bee colonies. In this paper, loop-mediated isothermal amplifications (LAMP) were used for the first time to identify and differentiate N. apis and N. ceranae, the causative agents of nosemosis. LAMP assays were performed at a constant temperature of 60 °C using two sets of six species-specific primers, recognising eight distinct fragments of 16S rDNA gene and GspSSD polymerase with strand displacement activity. The optimal time for LAMP and its Nosema species sensitivity and specificity were assessed. LAMP only required 30 min for robust identification of the amplicons. Ten-fold serial dilutions of total DNA isolated from bees infected with microsporidia were used to determine the detection limit of N. apis and N. ceranae DNAs by LAMP and standard PCR assays. LAMP appeared to be 10(3) -fold more sensitive than a standard PCR in detecting N. apis and N. ceranae. LAMP methods developed by us are highly Nosema species specific and allow to identify and differentiate N. apis and N. ceranae. © 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.

Development of a novel loop-mediated isothermal amplification (LAMP) assay for the detection of Salmonella ser. Enteritidis from egg products

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Salmonella ser. Enteritidis is a major public health concern worldwide. Loop-mediated isothermal amplification (LAMP) is a novel simple, easy-to-operate detection technology that amplifies DNA with high speed, efficiency, and specificity under isothermal conditions. The objective of this study was t...

Identification of ochratoxin A producing Aspergillus carbonarius and A. niger clade isolated from grapes using the loop-mediated isothermal amplification (LAMP) reaction.

Science.gov (United States)

Storari, M; von Rohr, R; Pertot, I; Gessler, C; Broggini, G A L

2013-04-01

To develop two assays based on the loop-mediated isothermal amplification (LAMP) of DNA for the quick and specific identification of Aspergillus carbonarius and ochratoxigenic strains of the Aspergillus niger clade isolated from grapes. Two sets of primers were designed based on the polyketide synthase genes involved or putatively involved in ochratoxin A (OTA) biosynthesis in A. carbonarius and A. niger clade. Hydroxynaphthol blue was used as indirect method to indicate DNA amplification. The limit of detection of both assays was comparable to that of a PCR reaction. Specificities of the reactions were tested using DNA from different black aspergilli isolated from grapes. The two LAMP assays were then used to identify A. carbonarius and ochratoxigenic A. niger and A. awamori grown in pure cultures without a prior DNA extraction. The two LAMP assays permitted to quickly and specifically identify DNA from OTA-producing black aspergilli, as well as isolates grown in pure culture. Monitoring vineyards for the presence of OTA-producing strains is part of the measures to minimize the occurrence of OTA in grape products. The two LAMP assays developed here could be potentially used to speed the screening process of vineyards for the presence of OTA-producing black aspergilli. © 2013 The Society for Applied Microbiology.

Assessing the performance of a Loop Mediated Isothermal Amplification (LAMP) assay for the detection and subtyping of high-risk suptypes of Human Papilloma Virus (HPV) for Oropharyngeal Squamous Cell Carcinoma (OPSCC) without DNA purification.

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Rohatensky, Mitchell G; Livingstone, Devon M; Mintchev, Paul; Barnes, Heather K; Nakoneshny, Steven C; Demetrick, Douglas J; Dort, Joseph C; van Marle, Guido

2018-02-08

Oropharyngeal Squamous Cell Carcinoma (OPSCC) is increasing in incidence despite a decline in traditional risk factors. Human Papilloma Virus (HPV), specifically subtypes 16, 18, 31 and 35, has been implicated as the high-risk etiologic agent. HPV positive cancers have a significantly better prognosis than HPV negative cancers of comparable stage, and may benefit from different treatment regimens. Currently, HPV related carcinogenesis is established indirectly through Immunohistochemistry (IHC) staining for p16, a tumour suppressor gene, or polymerase chain reaction (PCR) that directly tests for HPV DNA in biopsied tissue. Loop mediated isothermal amplification (LAMP) is more accurate than IHC, more rapid than PCR and is significantly less costly. In previous work we showed that a subtype specific HPV LAMP assay performed similar to PCR on purified DNA. In this study we examined the performance of this LAMP assay without DNA purification. We used LAMP assays using established primers for HPV 16 and 18, and new primers for HPV 31 and 35. LAMP reaction conditions were tested on serial dilutions of plasmid HPV DNA to confirm minimum viral copy number detection thresholds. LAMP was then performed directly on different human cell line samples without DNA purification. Our LAMP assays could detect 10 5 , 10 3 , 10 4 , and 10 5 copies of plasmid DNA for HPV 16, 18, 31, and 35, respectively. All primer sets were subtype specific, with no cross-amplification. Our LAMP assays also reliably amplified subtype specific HPV DNA from samples without requiring DNA isolation and purification. The high risk OPSCC HPV subtype specific LAMP primer sets demonstrated, excellent clinically relevant, minimum copy number detection thresholds with an easy readout system. Amplification directly from samples without purification illustrated the robust nature of the assay, and the primers used. This lends further support HPV type specific LAMP assays, and these specific primer sets and assays

Application of a loop-mediated isothermal amplification (LAMP) assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples.

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Rahman, S M Mazidur; Song, Hyun Beom; Jin, Yan; Oh, Jin-Kyoung; Lim, Min Kyung; Hong, Sung-Tae; Choi, Min-Ho

2017-10-01

Clonorchiasis is prevalent in the Far East, and a major health problem in endemic areas. Infected persons may experience, if not treated, serious complications such as bile stone formation, pyogenic cholangitis, and even cholangiocarcinoma. Early diagnosis and treatment are important to prevent serious complications and, therefore, the simple and reliable diagnostic method is necessary to control clonorchiasis in endemic areas, where resources for the diagnosis are limited. The loop-mediated isothermal amplification (LAMP) assay has been applied for the detection of Clonorchis sinensis DNA. Six primers targeting eight locations on the cytochrome c oxidase subunit 1 gene of C. sinensis were designed for species-specific amplification using the LAMP assay. The LAMP assay was sensitive enough to detect as little as 100 fg of C. sinensis genomic DNA and the detection limit in 100 mg of stool was as low as one egg. The assay was highly specific because no cross-reactivity was observed with the DNA of other helminths, protozoa or Escherichia coli. Then, LAMP assay was applied to human fecal samples collected from an endemic area of clonorchiasis in Korea. Using samples showing consistent results by both Kato-Katz method and real-time PCR as reference standards, the LAMP assay showed 97.1% (95% CI, 90.1-99.2) of sensitivity and 100% (95% CI, 92.9-100) of specificity. In stool samples with more than 100 eggs per gram of feces, the sensitivity achieved 100%. To detect C. sinensis in human fecal samples, the LAMP assay was applied and achieved high sensitivity and specificity. The LAMP assay can be utilized in field laboratories as a powerful tool for diagnosis and epidemiological survey of clonorchiasis.

A rapid molecular diagnosis of cutaneous leishmaniasis by colorimetric malachite green-loop-mediated isothermal amplification (LAMP) combined with an FTA card as a direct sampling tool.

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Nzelu, Chukwunonso O; Cáceres, Abraham G; Guerrero-Quincho, Silvia; Tineo-Villafuerte, Edwin; Rodriquez-Delfin, Luis; Mimori, Tatsuyuki; Uezato, Hiroshi; Katakura, Ken; Gomez, Eduardo A; Guevara, Angel G; Hashiguchi, Yoshihisa; Kato, Hirotomo

2016-01-01

Leishmaniasis remains one of the world's most neglected diseases, and early detection of the infectious agent, especially in developing countries, will require a simple and rapid test. In this study, we established a quick, one-step, single-tube, highly sensitive loop-mediated isothermal amplification (LAMP) assay for rapid detection of Leishmania DNA from tissue materials spotted on an FTA card. An FTA-LAMP with pre-added malachite green was performed at 64°C for 60min using a heating block and/or water bath and DNA amplification was detected immediately after incubation. The LAMP assay had high detection sensitivity down to a level of 0.01 parasites per μl. The field- and clinic-applicability of the colorimetric FTA-LAMP assay was demonstrated with 122 clinical samples collected from patients suspected of having cutaneous leishmaniasis in Peru, from which 71 positives were detected. The LAMP assay in combination with an FTA card described here is rapid and sensitive, as well as simple to perform, and has great potential usefulness for diagnosis and surveillance of leishmaniasis in endemic areas. Copyright © 2015 Elsevier B.V. All rights reserved.

Clinical evaluation of a loop-mediated isothermal amplification (LAMP) assay for rapid detection of Neisseria meningitidis in cerebrospinal fluid.

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Lee, DoKyung; Kim, Eun Jin; Kilgore, Paul E; Kim, Soon Ae; Takahashi, Hideyuki; Ohnishi, Makoto; Anh, Dang Duc; Dong, Bai Qing; Kim, Jung Soo; Tomono, Jun; Miyamoto, Shigehiko; Notomi, Tsugunori; Kim, Dong Wook; Seki, Mitsuko

2015-01-01

Neisseria meningitidis (Nm) is a leading causative agent of bacterial meningitis in humans. Traditionally, meningococcal meningitis has been diagnosed by bacterial culture. However, isolation of bacteria from patients' cerebrospinal fluid (CSF) is time consuming and sometimes yields negative results. Recently, polymerase chain reaction (PCR)-based diagnostic methods of detecting Nm have been considered the gold standard because of their superior sensitivity and specificity compared with culture. In this study, we developed a loop-mediated isothermal amplification (LAMP) method and evaluated its ability to detect Nm in cerebrospinal fluid (CSF). We developed a meningococcal LAMP assay (Nm LAMP) that targets the ctrA gene. The primer specificity was validated using 16 strains of N. meningitidis (serogroup A, B, C, D, 29-E, W-135, X, Y, and Z) and 19 non-N. meningitidis species. Within 60 min, the Nm LAMP detected down to ten copies per reaction with sensitivity 1000-fold more than that of conventional PCR. The LAMP assays were evaluated using a set of 1574 randomly selected CSF specimens from children with suspected meningitis collected between 1998 and 2002 in Vietnam, China, and Korea. The LAMP method was shown to be more sensitive than PCR methods for CSF samples (31 CSF samples were positive by LAMP vs. 25 by PCR). The detection rate of the LAMP method was substantially higher than that of the PCR method. In a comparative analysis of the PCR and LAMP assays, the clinical sensitivity, specificity, positive predictive value, and negative predictive value of the LAMP assay were 100%, 99.6%, 80.6%, and 100%, respectively. Compared to PCR, LAMP detected Nm with higher analytical and clinical sensitivity. This sensitive and specific LAMP method offers significant advantages for screening patients on a population basis and for diagnosis in clinical settings.

Clinical evaluation of a loop-mediated isothermal amplification (LAMP assay for rapid detection of Neisseria meningitidis in cerebrospinal fluid.

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DoKyung Lee

Full Text Available Neisseria meningitidis (Nm is a leading causative agent of bacterial meningitis in humans. Traditionally, meningococcal meningitis has been diagnosed by bacterial culture. However, isolation of bacteria from patients' cerebrospinal fluid (CSF is time consuming and sometimes yields negative results. Recently, polymerase chain reaction (PCR-based diagnostic methods of detecting Nm have been considered the gold standard because of their superior sensitivity and specificity compared with culture. In this study, we developed a loop-mediated isothermal amplification (LAMP method and evaluated its ability to detect Nm in cerebrospinal fluid (CSF.We developed a meningococcal LAMP assay (Nm LAMP that targets the ctrA gene. The primer specificity was validated using 16 strains of N. meningitidis (serogroup A, B, C, D, 29-E, W-135, X, Y, and Z and 19 non-N. meningitidis species. Within 60 min, the Nm LAMP detected down to ten copies per reaction with sensitivity 1000-fold more than that of conventional PCR. The LAMP assays were evaluated using a set of 1574 randomly selected CSF specimens from children with suspected meningitis collected between 1998 and 2002 in Vietnam, China, and Korea. The LAMP method was shown to be more sensitive than PCR methods for CSF samples (31 CSF samples were positive by LAMP vs. 25 by PCR. The detection rate of the LAMP method was substantially higher than that of the PCR method. In a comparative analysis of the PCR and LAMP assays, the clinical sensitivity, specificity, positive predictive value, and negative predictive value of the LAMP assay were 100%, 99.6%, 80.6%, and 100%, respectively.Compared to PCR, LAMP detected Nm with higher analytical and clinical sensitivity. This sensitive and specific LAMP method offers significant advantages for screening patients on a population basis and for diagnosis in clinical settings.

Specific detection of Angiostrongylus cantonensis in the snail Achatina fulica using a loop-mediated isothermal amplification (LAMP) assay.

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Liu, Chun-Yan; Song, Hui-Qun; Zhang, Ren-Li; Chen, Mu-Xin; Xu, Min-Jun; Ai, Lin; Chen, Xiao-Guang; Zhan, Xi-Mei; Liang, Shao-Hui; Yuan, Zi-Guo; Lin, Rui-Qing; Zhu, Xing-Quan

2011-08-01

Angiostrongylus cantonensis, a rat lungworm, can cause eosinophilic meningitis and angiostrongyliasis in humans following ingestion of contaminated foods or intermediate/paratenic hosts with infective larvae. The snail Achatina fulica is one of the important intermediate hosts of A. cantonensis and is commonly eaten by humans in some countries. In the present study, we developed a loop-mediated isothermal amplification (LAMP) method for the specific detection of A. cantonensis in Ac. fulica. Primers for LAMP were designed based on the first internal transcribed spacer (ITS-1) of nuclear ribosomal DNA (rDNA) of A. cantonensis. Specificity tests showed that only the products of A. cantonensis were detected when DNA samples of A. cantonensis and the heterologous control samples Anisakis simplex s.s, Trichuris trichiura, Toxocara canis, Trichinella spiralis and Ascaris lumbricoides were amplified by LAMP. Sensitivity evaluation indicated that the LAMP assay is 10 times more sensitive than the conventional polymerase chain reaction (PCR) assay. The established LAMP assay is rapid, inexpensive and easy to be performed. It can be used in clinical applications for rapid and sensitive detection of A. cantonensis in snails, which has implications for the effective control of angiostrongyliasis. Copyright © 2011 Elsevier Ltd. All rights reserved.

Simple system for isothermal DNA amplification coupled to lateral flow detection.

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Kristina Roskos

Full Text Available Infectious disease diagnosis in point-of-care settings can be greatly improved through integrated, automated nucleic acid testing devices. We have developed an early prototype for a low-cost system which executes isothermal DNA amplification coupled to nucleic acid lateral flow (NALF detection in a mesofluidic cartridge attached to a portable instrument. Fluid handling inside the cartridge is facilitated through one-way passive valves, flexible pouches, and electrolysis-driven pumps, which promotes a compact and inexpensive instrument design. The closed-system disposable prevents workspace amplicon contamination. The cartridge design is based on standard scalable manufacturing techniques such as injection molding. Nucleic acid amplification occurs in a two-layer pouch that enables efficient heat transfer. We have demonstrated as proof of principle the amplification and detection of Mycobacterium tuberculosis (M.tb genomic DNA in the cartridge, using either Loop Mediated Amplification (LAMP or the Exponential Amplification Reaction (EXPAR, both coupled to NALF detection. We envision that a refined version of this cartridge, including upstream sample preparation coupled to amplification and detection, will enable fully-automated sample-in to answer-out infectious disease diagnosis in primary care settings of low-resource countries with high disease burden.

A loop-mediated isothermal amplification (LAMP assay for early detection of Schistosoma mansoni in stool samples: a diagnostic approach in a murine model.

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Pedro Fernández-Soto

2014-09-01

Full Text Available Human schistosomiasis, mainly due to Schistosoma mansoni species, is one of the most prevalent parasitic diseases worldwide. To overcome the drawbacks of classical parasitological and serological methods in detecting S. mansoni infections, especially in acute stage of the disease, development of cost-effective, simple and rapid molecular methods is still needed for the diagnosis of schistosomiasis. A promising approach is the loop-mediated isothermal amplification (LAMP technology. Compared to PCR-based assays, LAMP has the advantages of reaction simplicity, rapidity, specificity, cost-effectiveness and higher amplification efficiency. Additionally, as results can be inspected by the naked eye, the technique has great potential for use in low-income countries.A sequence corresponding to a mitochondrial S. mansoni minisatellite DNA region was selected as a target for designing a LAMP-based method to detect S. mansoni DNA in stool samples. We used a S. mansoni murine model to obtain well defined stool and sera samples from infected mice with S. mansoni cercariae. Samples were taken weekly from week 0 to 8 post-infection and the Kato-Katz and ELISA techniques were used for monitoring the infection. Primer set designed were tested using a commercial reaction mixture for LAMP assay and an in house mixture to compare results. Specificity of LAMP was tested using 16 DNA samples from different parasites, including several Schistosoma species, and no cross-reactions were found. The detection limit of our LAMP assay (SmMIT-LAMP was 1 fg of S. mansoni DNA. When testing stool samples from infected mice the SmMIT-LAMP detected S. mansoni DNA as soon as 1 week post-infection.We have developed, for the first time, a cost-effective, easy to perform, specific and sensitive LAMP assay for early detection of S. mansoni in stool samples. The method is potentially and readily adaptable for field diagnosis and disease surveillance in schistosomiasis-endemic areas.

Application of a loop-mediated isothermal amplification (LAMP assay targeting cox1 gene for the detection of Clonorchis sinensis in human fecal samples.

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S M Mazidur Rahman

2017-10-01

Full Text Available Clonorchiasis is prevalent in the Far East, and a major health problem in endemic areas. Infected persons may experience, if not treated, serious complications such as bile stone formation, pyogenic cholangitis, and even cholangiocarcinoma. Early diagnosis and treatment are important to prevent serious complications and, therefore, the simple and reliable diagnostic method is necessary to control clonorchiasis in endemic areas, where resources for the diagnosis are limited.The loop-mediated isothermal amplification (LAMP assay has been applied for the detection of Clonorchis sinensis DNA. Six primers targeting eight locations on the cytochrome c oxidase subunit 1 gene of C. sinensis were designed for species-specific amplification using the LAMP assay. The LAMP assay was sensitive enough to detect as little as 100 fg of C. sinensis genomic DNA and the detection limit in 100 mg of stool was as low as one egg. The assay was highly specific because no cross-reactivity was observed with the DNA of other helminths, protozoa or Escherichia coli. Then, LAMP assay was applied to human fecal samples collected from an endemic area of clonorchiasis in Korea. Using samples showing consistent results by both Kato-Katz method and real-time PCR as reference standards, the LAMP assay showed 97.1% (95% CI, 90.1-99.2 of sensitivity and 100% (95% CI, 92.9-100 of specificity. In stool samples with more than 100 eggs per gram of feces, the sensitivity achieved 100%.To detect C. sinensis in human fecal samples, the LAMP assay was applied and achieved high sensitivity and specificity. The LAMP assay can be utilized in field laboratories as a powerful tool for diagnosis and epidemiological survey of clonorchiasis.

Visual detection of human enterovirus 71 subgenotype C4 and Coxsackievirus A16 by reverse transcription loop-mediated isothermal amplification with the hydroxynaphthol blue dye.

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Nie, Kai; Zhang, Yong; Luo, Le; Yang, Meng-Jie; Hu, Xiu-Mei; Wang, Miao; Zhu, Shuang-Li; Han, Feng; Xu, Wen-Bo; Ma, Xue-Jun

2011-08-01

A sensitive reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for rapid visual detection of human enterovirus 71 subgenotype C4 (EV71-C4) and Coxsackievirus A16 (CVA16) infection, respectively. The reaction was performed in one step in a single tube at 65°C for 60 min with the addition of the hydroxynaphthol blue (HNB) dye prior to amplification. The detection limits of the RT-LAMP assay were 0.33 and 1.58 of a 50% tissue culture infective dose (TCID(50)) per reaction based on 10-fold dilutions of a titrated EV71 or CVA16 strain, respectively. No cross-reaction was observed with Coxsackievirus A (CVA) viruses (CVA2, 4, 5, 7, 9, 10, 14, and 24), Coxsackievirus B (CVB) viruses (CVB1,2,3,4, and 5) or ECHO viruses (ECHO3, 6, 11, and 19). The assay was further evaluated with 47 clinical stool specimens diagnosed previously with EV71, CVA16 or other human enterovirus infections. Virus isolates from stool samples were confirmed by virus neutralization testing and sequencing. RT-LAMP with HNB dye was demonstrated to be a sensitive and cost-effective assay for rapid visual detection of human EV71-C4 and CVA16. Copyright © 2011 Elsevier B.V. All rights reserved.

Development of multiplex loop mediated isothermal amplification (m-LAMP) label-based gold nanoparticles lateral flow dipstick biosensor for detection of pathogenic Leptospira.

Science.gov (United States)

Nurul Najian, A B; Engku Nur Syafirah, E A R; Ismail, Nabilah; Mohamed, Maizan; Yean, Chan Yean

2016-01-15

In recent years extensive numbers of molecular diagnostic methods have been developed to meet the need of point-of-care devices. Efforts have been made towards producing rapid, simple and inexpensive DNA tests, especially in the diagnostics field. We report on the development of a label-based lateral flow dipstick for the rapid and simple detection of multiplex loop-mediated isothermal amplification (m-LAMP) amplicons. A label-based m-LAMP lateral flow dipstick assay was developed for the simultaneous detection of target DNA template and a LAMP internal control. This biosensor operates through a label based system, in which probe-hybridization and the additional incubation step are eliminated. We demonstrated this m-LAMP assay by detecting pathogenic Leptospira, which causes the re-emerging disease Leptospirosis. The lateral flow dipstick was developed to detect of three targets, the LAMP target amplicon, the LAMP internal control amplicon and a chromatography control. Three lines appeared on the dipstick, indicating positive results for all representative pathogenic Leptospira species, whereas two lines appeared, indicating negative results, for other bacterial species. The specificity of this biosensor assay was 100% when it was tested with 13 representative pathogenic Leptospira species, 2 intermediate Leptospira species, 1 non-pathogenic Leptospira species and 28 other bacteria species. This study found that this DNA biosensor was able to detect DNA at concentrations as low as 3.95 × 10(-1) genomic equivalent ml(-1). An integrated m-LAMP and label-based lateral flow dipstick was successfully developed, promising simple and rapid visual detection in clinical diagnostics and serving as a point-of-care device. Copyright © 2015 Elsevier B.V. All rights reserved.

Diagnostic performance of a novel loop-mediated isothermal amplification (LAMP) assay targeting the apicoplast genome for malaria diagnosis in a field setting in sub-Saharan Africa.

Science.gov (United States)

Oriero, Eniyou C; Okebe, Joseph; Jacobs, Jan; Van Geertruyden, Jean-Pierre; Nwakanma, Davis; D'Alessandro, Umberto

2015-10-09

New diagnostic tools to detect reliably and rapidly asymptomatic and low-density malaria infections are needed as their treatment could interrupt transmission. Isothermal amplification techniques are being explored for field diagnosis of malaria. In this study, a novel molecular tool (loop-mediated isothermal amplification-LAMP) targeting the apicoplast genome of Plasmodium falciparum was evaluated for the detection of asymptomatic malaria-infected individuals in a rural setting in The Gambia. A blood was collected from 341 subjects (median age 9 years, range 1-68 years) screened for malaria. On site, a rapid diagnostic test (RDT, SD Bioline Malaria Antigen P.f) was performed, thick blood films (TBF) slides for microscopy were prepared and dry blood spots (DBS) were collected on Whatman(®) 903 Specimen collection paper. The TBF and DBS were transported to the field laboratory where microscopy and LAMP testing were performed. The latter was done on DNA extracted from the DBS using a crude (methanol/heating) extraction method. A laboratory-based PCR amplification was done on all the samples using DNA extracted with the Qiagen kit and its results were taken as reference for all the other tests. Plasmodium falciparum malaria prevalence was 37 % (127/341) as detected by LAMP, 30 % (104/341) by microscopy and 37 % (126/341) by RDT. Compared to the reference PCR method, sensitivity was 92 % for LAMP, 78 % for microscopy, and 76 % for RDT; specificity was 97 % for LAMP, 99 % for microscopy, and 88 % for RDT. Area under the receiver operating characteristic (ROC) curve in comparison with the reference standard was 0.94 for LAMP, 0.88 for microscopy and 0.81 for RDT. Turn-around time for the entire LAMP assay was approximately 3 h and 30 min for an average of 27 ± 9.5 samples collected per day, compared to a minimum of 10 samples an hour per operator by RDT and over 8 h by microscopy. The LAMP assay could produce reliable results the same day of the screening. It could

Development of a loop-mediated isothermal amplification (LAMP) assay for rapid screening of ticks and fleas for spotted fever group rickettsia.

Science.gov (United States)

Noden, Bruce H; Martin, Jaclyn; Carrillo, Yisel; Talley, Justin L; Ochoa-Corona, Francisco M

2018-01-01

The importance of tick and flea-borne rickettsia infections is increasingly recognized worldwide. While increased focus has shifted in recent years to the development of point-of-care diagnostics for various vector-borne diseases in humans and animals, little research effort has been devoted to their integration into vector surveillance and control programs, particularly in resource-challenged countries. One technology which may be helpful for large scale vector surveillance initiatives is loop-mediated isothermal amplification (LAMP). The aim of this study was to develop a LAMP assay to detect spotted fever group (SFG) rickettsia DNA from field-collected ticks and fleas and compare with published end-point PCR results. A Spotted Fever Group rickettsia-specific loop-mediated isothermal amplification (SFGR-LAMP) assay was developed using primers based on a region of the R. rickettsii 17kDa protein gene. The sensitivity, specificity, and reproducibility of the assay were evaluated. The assay was then compared with the results of end-point PCR assays for pooled tick and flea samples obtained from field-based surveillance studies. The sensitivity of the SFGR-LAMP assay was 0.00001 ng/μl (25μl volume) which was 10 times more sensitive than the 17kDa protein gene end-point PCR used as the reference method. The assay only recognized gDNA from SFG and transitional group (TRG) rickettsia species tested but did not detect gDNA from typhus group (TG) rickettsia species or closely or distantly related bacterial species. The SFGR-LAMP assay detected the same positives from a set of pooled tick and flea samples detected by end-point PCR in addition to two pooled flea samples not detected by end-point PCR. To our knowledge, this is the first study to develop a functional LAMP assay to initially screen for SFG and TRG rickettsia pathogens in field-collected ticks and fleas. With a high sensitivity and specificity, the results indicate the potential use as a field

Comparison of loop-mediated isothermal amplification (LAMP) and nested-PCR assay targeting the RE and B1 gene for detection of Toxoplasma gondii in blood samples of children with leukaemia.

Science.gov (United States)

Fallahi, Shirzad; Seyyed Tabaei, Seyyed Javad; Pournia, Yadollah; Zebardast, Nozhat; Kazemi, Bahram

2014-07-01

Toxoplasmosis diagnosis constitutes an important measure for disease prevention and control. In this paper, a newly described DNA amplification technique, loop-mediated isothermal amplification (LAMP), and nested-PCR targeting the repeated element (RE) and B1 gene, were compared to each other for the detection of Toxoplasma gondii DNA in blood samples of children with leukaemia. One hundred ten blood samples from these patients were analyzed by LAMP and nested-PCR. Out of 50 seropositive samples (IgM+, IgG+), positive results were obtained with 92% and 86% on RE, B1-LAMP and 82% and 68% on RE, B1-nested PCR analyses, respectively. Of the 50 seronegative samples, three, two and one samples were detected positive by RE-LAMP, B1-LAMP and RE-nested PCR assays, respectively, while none were detected positive by B1-nested PCR. None of the 10 IgM-, IgG+ samples was detected positive after testing LAMP and nested-PCR assays in duplicate. This is the first report of a study in which the LAMP method was applied with high sensitivity and efficacy for the diagnosis of T. gonii in blood samples of children with leukaemia. Copyright © 2014 Elsevier Inc. All rights reserved.

UHP lamp systems for projection applications

International Nuclear Information System (INIS)

Derra, Guenther; Moench, Holger; Fischer, Ernst; Giese, Hermann; Hechtfischer, Ulrich; Heusler, Gero; Koerber, Achim; Niemann, Ulrich; Noertemann, Folke-Charlotte; Pekarski, Pavel; Pollmann-Retsch, Jens; Ritz, Arnd; Weichmann, Ulrich

2005-01-01

Projection systems have found widespread use in conference rooms and other professional applications during the last decade and are now entering the home TV market at a considerable pace. Projectors as small as about one litre are able to deliver several thousand screen lumens and are, with a system efficacy of over 10 lm W -1 , the most efficient display systems realized today. Short arc lamps are a key component for projection systems of the highest efficiency for small-size projection displays. The introduction of the ultra high performance (UHP) lamp system by Philips in 1995 can be identified as one of the key enablers of the commercial success of projection systems. The UHP lamp concept features outstanding arc luminance, a well suited spectrum, long life and excellent lumen maintenance. For the first time it combines a very high pressure mercury discharge lamp with extremely short and stable arc gap with a regenerative chemical cycle keeping the discharge walls free from blackening, leading to lifetimes of over 10 000 h. Since the introduction of the UHP lamp system, many important new technology improvements have been realized: burner designs for higher lamp power, advanced ignition systems, miniaturized electronic drivers and innovative reflector concepts. These achievements enabled the impressive increase of projector light output, a remarkable reduction in projector size and even higher optical efficiency in projection systems during the last years. In this paper the concept of the UHP lamp system is described, followed by a discussion of the technological evolution the UHP lamp has undergone so far. Last, but not least, the important improvements of the UHP lamp system including the electronic driver and the reflector are discussed. (review article)

Development of loop-mediated isothermal amplification (LAMP) assays for the rapid detection of allergic peanut in processed food.

Science.gov (United States)

Sheu, Shyang-Chwen; Tsou, Po-Chuan; Lien, Yi-Yang; Lee, Meng-Shiou

2018-08-15

Peanut is a widely and common used in many cuisines around the world. However, peanut is also one of the most important food allergen for causing anaphylactic reaction. To prevent allergic reaction, the best way is to avoid the food allergen or food containing allergic ingredient such as peanut before food consuming. Thus, to efficient and precisely detect the allergic ingredient, peanut or related product, is essential and required for maintain consumer's health or their interest. In this study, a loop-mediated isothermal amplification (LAMP) assay was developed for the detection of allergic peanut using specifically designed primer sets. Two sets of the specific LAMP primers respectively targeted the internal transcribed sequence 1 (ITS1) of nuclear ribosomal DNA sequence regions and the ara h1 gene sequence of Arachia hypogeae (peanut) were used to address the application of LAMP for detecting peanut in processed food or diet. The results demonstrated that the identification of peanut using the newly designed primers for ITS 1 sequence is more sensitive rather than primers for sequence of Ara h1 gene when performing LAMP assay. Besides, the sensitivity of LAMP for detecting peanut is also higher than the traditional PCR method. These LAMP primers sets showed high specificity for the identification of the peanut and had no cross-reaction to other species of nut including walnut, hazelnut, almonds, cashew and macadamia nut. Moreover, when minimal 0.1% peanuts were mixed with other nuts ingredients at different ratios, no any cross-reactivity was evident during performing LAMP. Finally, genomic DNAs extracted from boiled and steamed peanut were used as templates; the detection of peanut by LAMP was not affected and reproducible. As to this established LAMP herein, not only can peanut ingredients be detected but commercial foods containing peanut can also be identified. This assay will be useful and potential for the rapid detection of peanut in practical food

Simultaneous detection and differentiation of dengue virus serotypes 1-4, Japanese encephalitis virus, and West Nile virus by a combined reverse-transcription loop-mediated isothermal amplification assay

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Yin Jianhua

2011-07-01

Full Text Available Abstract Background Rapid identification and differentiation of mosquito-transmitted flaviviruses in acute-phase sera of patients and field-caught vector mosquitoes are important for the prediction and prevention of large-scale epidemics. Results We developed a flexible reverse-transcription loop-mediated isothermal amplification (RT-LAMP unit for the detection and differentiation of dengue virus serotypes 1-4 (DENV1-4, Japanese encephalitis virus (JEV, and West Nile virus (WNV. The unit efficiently amplified the viral genomes specifically at wide ranges of viral template concentrations, and exhibited similar amplification curves as monitored by a real-time PCR engine. The detection limits of the RT-LAMP unit were 100-fold higher than that of RT-PCR in 5 of the six flaviviruses. The results on specificity indicated that the six viruses in the assay had no cross-reactions with each other. By examining 66 viral strains of DENV1-4 and JEV, the unit identified the viruses with 100% accuracy and did not cross-react with influenza viruses and hantaviruses. By screening a panel of specimens containing sera of 168 patients and 279 pools of field-caught blood sucked mosquitoes, results showed that this unit is high feasible in clinical settings and epidemiologic field, and it obtained results 100% correlated with real-time RT-PCR. Conclusions The RT-LAMP unit developed in this study is able to quickly detect and accurately differentiate the six kinds of flaviviruses, which makes it extremely feasible for screening these viruses in acute-phase sera of the patients and in vector mosquitoes without the need of high-precision instruments.

Loop-mediated isothermal amplification (LAMP) assays for the species-specific detection of Eimeria that infect chickens.

Science.gov (United States)

Barkway, Christopher P; Pocock, Rebecca L; Vrba, Vladimir; Blake, Damer P

2015-02-20

Eimeria species parasites, protozoa which cause the enteric disease coccidiosis, pose a serious threat to the production and welfare of chickens. In the absence of effective control clinical coccidiosis can be devastating. Resistance to the chemoprophylactics frequently used to control Eimeria is common and sub-clinical infection is widespread, influencing feed conversion ratios and susceptibility to other pathogens such as Clostridium perfringens. Despite the availability of polymerase chain reaction (PCR)-based tools, diagnosis of Eimeria infection still relies almost entirely on traditional approaches such as lesion scoring and oocyst morphology, but neither is straightforward. Limitations of the existing molecular tools include the requirement for specialist equipment and difficulties accessing DNA as template. In response a simple field DNA preparation protocol and a panel of species-specific loop-mediated isothermal amplification (LAMP) assays have been developed for the seven Eimeria recognised to infect the chicken. We now provide a detailed protocol describing the preparation of genomic DNA from intestinal tissue collected post-mortem, followed by setup and readout of the LAMP assays. Eimeria species-specific LAMP can be used to monitor parasite occurrence, assessing the efficacy of a farm's anticoccidial strategy, and to diagnose sub-clinical infection or clinical disease with particular value when expert surveillance is unavailable.

Loop-mediated isothermal amplification (LAMP) assay for the identification of Echinococcus multilocularis infections in canine definitive hosts.

Science.gov (United States)

Ni, Xingwei; McManus, Donald P; Yan, Hongbin; Yang, Jifei; Lou, Zhongzi; Li, Hongmin; Li, Li; Lei, Mengtong; Cai, Jinzhong; Fan, Yanlei; Li, Chunhua; Liu, Quanyuan; Shi, Wangui; Liu, Xu; Zheng, Yadong; Fu, Baoquan; Yang, Yurong; Jia, Wanzhong

2014-05-30

Alveolar echinococcosis, caused by the metacestode larval stage of Echinococcus multilocularis, is a zoonosis of public health significance and is highly prevalent in northwest China. To effectively monitor its transmission, we developed a new rapid and cheap diagnostic assay, based on loop-mediated isothermal amplification (LAMP), to identify canine definitive hosts infected with E. multilocularis. The primers used in the LAMP assay were based on the mitochondrial nad5 gene of E. multilocularis and were designed using Primer Explorer V4 software. The developed LAMP assay was compared with a conventional PCR assay, using DNA extracted from the feces of dogs experimentally infected with E. multilocularis, on 189 dog fecal samples collected from three E. multilocularis-endemic regions in Qinghai province, the People's Republic of China, and 30 negative control copro-samples from dogs from an area in Gansu province that had been subjected to an intensive de-worming program. Light microscopy was also used to examine the experimentally obtained and field collected dog copro-samples for the presence of E. multilocularis eggs. The E. multilocularis-positivity rates obtained for the field-collected fecal samples were 16.4% and 5.3% by the LAMP and PCR assays, respectively, and all samples obtained from the control dogs were negative. The LAMP assay was able to detect E. multilocularis DNA in the feces of experimentally infected dogs at 12 days post-infection, whereas the PCR assay was positive on the 17th day and eggs were first detectable by light microscopy at day 44 post-challenge. The earlier specific detection of an E. multilocularis infection in dog copro-samples indicates that the LAMP assay we developed is a realistic alternative method for the field surveillance of canines in echinococcosis-endemic areas.

Development of multiplex loop mediated isothermal amplification (m-LAMP) label-based gold nanoparticles lateral flow dipstick biosensor for detection of pathogenic Leptospira

International Nuclear Information System (INIS)

Nurul Najian, A.B.; Engku Nur Syafirah, E.A.R.; Ismail, Nabilah; Mohamed, Maizan; Yean, Chan Yean

2016-01-01

In recent years extensive numbers of molecular diagnostic methods have been developed to meet the need of point-of-care devices. Efforts have been made towards producing rapid, simple and inexpensive DNA tests, especially in the diagnostics field. We report on the development of a label-based lateral flow dipstick for the rapid and simple detection of multiplex loop-mediated isothermal amplification (m-LAMP) amplicons. A label-based m-LAMP lateral flow dipstick assay was developed for the simultaneous detection of target DNA template and a LAMP internal control. This biosensor operates through a label based system, in which probe-hybridization and the additional incubation step are eliminated. We demonstrated this m-LAMP assay by detecting pathogenic Leptospira, which causes the re-emerging disease Leptospirosis. The lateral flow dipstick was developed to detect of three targets, the LAMP target amplicon, the LAMP internal control amplicon and a chromatography control. Three lines appeared on the dipstick, indicating positive results for all representative pathogenic Leptospira species, whereas two lines appeared, indicating negative results, for other bacterial species. The specificity of this biosensor assay was 100% when it was tested with 13 representative pathogenic Leptospira species, 2 intermediate Leptospira species, 1 non-pathogenic Leptospira species and 28 other bacteria species. This study found that this DNA biosensor was able to detect DNA at concentrations as low as 3.95 × 10 −1 genomic equivalent ml −1 . An integrated m-LAMP and label-based lateral flow dipstick was successfully developed, promising simple and rapid visual detection in clinical diagnostics and serving as a point-of-care device. - Highlights: • We develop multiplex LAMP label-based lateral flow dipstick biosensor for detection of pathogenic Leptospira. • We design primers for multiplex LAMP targeting the conserved LipL32 gene of pathogenic Leptospira and LAMP internal

Development of multiplex loop mediated isothermal amplification (m-LAMP) label-based gold nanoparticles lateral flow dipstick biosensor for detection of pathogenic Leptospira

Energy Technology Data Exchange (ETDEWEB)

Nurul Najian, A.B.; Engku Nur Syafirah, E.A.R.; Ismail, Nabilah [Department of Medical Microbiology & Parasitology, School of Medical Sciences, Health Campus, Universiti Sains Malaysia, 16150 Kubang Kerian, Kelantan (Malaysia); Mohamed, Maizan [Faculty of Veterinary Medicine, Universiti Malaysia Kelantan, City Campus, Pengkalan Chepa, Locked Bag 36, 16100 Kota Bharu, Kelantan (Malaysia); Yean, Chan Yean, E-mail: yeancyn@yahoo.com [Department of Medical Microbiology & Parasitology, School of Medical Sciences, Health Campus, Universiti Sains Malaysia, 16150 Kubang Kerian, Kelantan (Malaysia); Institute for Research in Molecular Medicine (INFORMM), Health Campus, Universiti Sains Malaysia, 16150 Kubang Kerian, Kelantan (Malaysia)

2016-01-15

In recent years extensive numbers of molecular diagnostic methods have been developed to meet the need of point-of-care devices. Efforts have been made towards producing rapid, simple and inexpensive DNA tests, especially in the diagnostics field. We report on the development of a label-based lateral flow dipstick for the rapid and simple detection of multiplex loop-mediated isothermal amplification (m-LAMP) amplicons. A label-based m-LAMP lateral flow dipstick assay was developed for the simultaneous detection of target DNA template and a LAMP internal control. This biosensor operates through a label based system, in which probe-hybridization and the additional incubation step are eliminated. We demonstrated this m-LAMP assay by detecting pathogenic Leptospira, which causes the re-emerging disease Leptospirosis. The lateral flow dipstick was developed to detect of three targets, the LAMP target amplicon, the LAMP internal control amplicon and a chromatography control. Three lines appeared on the dipstick, indicating positive results for all representative pathogenic Leptospira species, whereas two lines appeared, indicating negative results, for other bacterial species. The specificity of this biosensor assay was 100% when it was tested with 13 representative pathogenic Leptospira species, 2 intermediate Leptospira species, 1 non-pathogenic Leptospira species and 28 other bacteria species. This study found that this DNA biosensor was able to detect DNA at concentrations as low as 3.95 × 10{sup −1} genomic equivalent ml{sup −1}. An integrated m-LAMP and label-based lateral flow dipstick was successfully developed, promising simple and rapid visual detection in clinical diagnostics and serving as a point-of-care device. - Highlights: • We develop multiplex LAMP label-based lateral flow dipstick biosensor for detection of pathogenic Leptospira. • We design primers for multiplex LAMP targeting the conserved LipL32 gene of pathogenic Leptospira and LAMP

Detection of feline Coronavirus in effusions of cats with and without feline infectious peritonitis using loop-mediated isothermal amplification.

Science.gov (United States)

Günther, Sonja; Felten, Sandra; Wess, Gerhard; Hartmann, Katrin; Weber, Karin

2018-06-01

Feline infectious peritonitis (FIP) is a fatal disease in cats worldwide. The aim of this study was to test two commercially available reaction mixtures in a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay to detect feline Coronavirus (FCoV) in body cavity effusions of cats with and without FIP, in order to minimize the time from sampling to obtaining results. RNA was extracted from body cavity effusion samples of 71 cats, including 34 samples from cats with a definitive diagnosis of FIP, and 37 samples of control cats with similar clinical signs but other confirmed diseases. Two reaction mixtures (Isothermal Mastermix, OptiGene Ltd.and PCRun™ Molecular Detection Mix, Biogal) were tested using the same primers, which were designed to bind to a conserved region of the FCoV membrane protein gene. Both assays were conducted under isothermal conditions (61 °C-62 °C). Using the Isothermal Mastermix of OptiGene Ltd., amplification times ranged from 4 and 39 min with a sensitivity of 35.3% and a specificity of 94.6% for the reported sample group. Using the PCRun™ Molecular Detection Mix of Biogal, amplification times ranged from 18 to 77 min with a sensitivity of 58.8% and a specificity of 97.3%. Although the RT-LAMP assay is less sensitive than real time reverse transcription PCR (RT-PCR), it can be performed without the need of expensive equipment and with less hands-on time. Further modifications of primers might lead to a suitable in-house test and accelerate the diagnosis of FIP. Copyright © 2018 Elsevier B.V. All rights reserved.

A rapid molecular diagnosis of cutaneous leishmaniasis bycolorimetric malachite green-loop-mediated isothermal amplification(LAMP) combined with an FTA card as a direct sampling tool

OpenAIRE

Nzelu, Chukwunonso O.; Cáceres, Abraham G.; Guerrero-Quincho, Silvia; Tineo-Villafuerte, Edwin; Rodriquez-Delfin, Luis; Mimori, Tatsuyuki; Uezato, Hiroshi; Katakura, Ken; Gomez, Eduardo A.; Guevara, Angel G.; Hashiguchi, Yoshihisa; Kato, Hirotomo

2015-01-01

Leishmaniasis remains one of the world's most neglected diseases, and early detection of the infectious agent, especially in developing countries, will require a simple and rapid test. In this study, we established a quick, one-step, single-tube, highly sensitive loop-mediated isothermal amplification (LAMP) assay for rapid detection of Leishmania DNA from tissue materials spotted on an FTA card. An FTA-LAMP with pre-added malachite green was performed at 64 degrees C for 60 mm using a heatin...

Development and Assessment of Loop-Mediated Isothermal Amplification (LAMP) Assay for the Diagnosis of Human Visceral Leishmaniasis in Iran.

Science.gov (United States)

Ghasemian, Mehrdad; Gharavi, Mohammad Javad; Akhlaghi, Lame; Mohebali, Mehdi; Meamar, Ahmad Reza; Aryan, Ehsan; Oormazdi, Hormozd

2014-03-01

Parasitological methods for the diagnosis of Visceral leishmaniasis (VL) require invasive procedures, so serological and molecular approaches have been developed but are not generally applicable in the field. We evaluated a loop mediated isothermal amplification (LAMP) assay using blood from VL patients and compared it to nested PCR. Forty-seven subjects with clinical features (fever, hepatosplenomegaly and anemia) were confirmed positive for VL by the direct agglutination test (DAT) at titers >3200. Forty DAT negative individuals from non-endemic areas with no clinical signs or symptoms of VL served as controls. A LAMP assay was performed using a set of six primers targeting Leishmania infantum kinetoplast DNA (kDNA) minicircle gene under isothermal (64 °C) conditions. For nested PCR we used primers targeting the kDNA minicircle gene. The LAMP assay provided a detection limit of 1 parasite in 1 ml of peripheral blood and detected L. infantum DNA in 44 of 47 DAT-confirmed VL cases, with diagnostic sensitivity of 93.6% (95% CI). No L. infantum DNA was amplified in controls, indicating a specificity of 100%. The nested PCR yielded sensitivity of 96% (95% CI) and a specificity of 100% (95% CI). The LAMP assay gave results similar to those of nested PCR but in a shorter time. The LAMP method is simple; requires no sophisticated equipment; has a short reaction time; and results, indicated by turbidity of the reaction mixture, are observable with the naked eye.

Development and Assessment of Loop-Mediated Isothermal Amplification (LAMP Assay for the Diagnosis of Human Visceral Leishmaniasis in Iran.

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Mehrdad Ghasemian

2014-03-01

Full Text Available Parasitological methods for the diagnosis of Visceral leishmaniasis (VL require invasive procedures, so serological and molecular approaches have been developed but are not generally applicable in the field. We evaluated a loop mediated isothermal amplification (LAMP assay using blood from VL patients and compared it to nested PCR.Forty-seven subjects with clinical features (fever, hepatosplenomegaly and anemia were confirmed positive for VL by the direct agglutination test (DAT at titers >3200. Forty DAT negative individuals from non-endemic areas with no clinical signs or symptoms of VL served as controls. A LAMP assay was performed using a set of six primers targeting Leishmania infantum kinetoplast DNA (kDNA minicircle gene under isothermal (64 °C conditions. For nested PCR we used primers targeting the kDNA minicircle gene.The LAMP assay provided a detection limit of 1 parasite in 1 ml of peripheral blood and detected L. infantum DNA in 44 of 47 DAT-confirmed VL cases, with diagnostic sensitivity of 93.6% (95% CI. No L. infantum DNA was amplified in controls, indicating a specificity of 100%. The nested PCR yielded sensitivity of 96% (95% CI and a specificity of 100% (95% CI.The LAMP assay gave results similar to those of nested PCR but in a shorter time. The LAMP method is simple; requires no sophisticated equipment; has a short reaction time; and results, indicated by turbidity of the reaction mixture, are observable with the naked eye.

Using detergent to enhance detection sensitivity of African trypanosomes in human CSF and blood by loop-mediated isothermal amplification (LAMP.

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Dennis J Grab

2011-08-01

Full Text Available The loop-mediated isothermal amplification (LAMP assay, with its advantages of simplicity, rapidity and cost effectiveness, has evolved as one of the most sensitive and specific methods for the detection of a broad range of pathogenic microorganisms including African trypanosomes. While many LAMP-based assays are sufficiently sensitive to detect DNA well below the amount present in a single parasite, the detection limit of the assay is restricted by the number of parasites present in the volume of sample assayed; i.e. 1 per µL or 10(3 per mL. We hypothesized that clinical sensitivities that mimic analytical limits based on parasite DNA could be approached or even obtained by simply adding detergent to the samples prior to LAMP assay.For proof of principle we used two different LAMP assays capable of detecting 0.1 fg genomic DNA (0.001 parasite. The assay was tested on dilution series of intact bloodstream form Trypanosoma brucei rhodesiense in human cerebrospinal fluid (CSF or blood with or without the addition of the detergent Triton X-100 and 60 min incubation at ambient temperature. With human CSF and in the absence of detergent, the LAMP detection limit for live intact parasites using 1 µL of CSF as the source of template was at best 10(3 parasites/mL. Remarkably, detergent enhanced LAMP assay reaches sensitivity about 100 to 1000-fold lower; i.e. 10 to 1 parasite/mL. Similar detergent-mediated increases in LAMP assay analytical sensitivity were also found using DNA extracted from filter paper cards containing blood pretreated with detergent before card spotting or blood samples spotted on detergent pretreated cards.This simple procedure for the enhanced detection of live African trypanosomes in biological fluids by LAMP paves the way for the adaptation of LAMP for the economical and sensitive diagnosis of other protozoan parasites and microorganisms that cause diseases that plague the developing world.

Using detergent to enhance detection sensitivity of African trypanosomes in human CSF and blood by loop-mediated isothermal amplification (LAMP).

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Grab, Dennis J; Nikolskaia, Olga V; Inoue, Noboru; Thekisoe, Oriel M M; Morrison, Liam J; Gibson, Wendy; Dumler, J Stephen

2011-08-01

The loop-mediated isothermal amplification (LAMP) assay, with its advantages of simplicity, rapidity and cost effectiveness, has evolved as one of the most sensitive and specific methods for the detection of a broad range of pathogenic microorganisms including African trypanosomes. While many LAMP-based assays are sufficiently sensitive to detect DNA well below the amount present in a single parasite, the detection limit of the assay is restricted by the number of parasites present in the volume of sample assayed; i.e. 1 per µL or 10(3) per mL. We hypothesized that clinical sensitivities that mimic analytical limits based on parasite DNA could be approached or even obtained by simply adding detergent to the samples prior to LAMP assay. For proof of principle we used two different LAMP assays capable of detecting 0.1 fg genomic DNA (0.001 parasite). The assay was tested on dilution series of intact bloodstream form Trypanosoma brucei rhodesiense in human cerebrospinal fluid (CSF) or blood with or without the addition of the detergent Triton X-100 and 60 min incubation at ambient temperature. With human CSF and in the absence of detergent, the LAMP detection limit for live intact parasites using 1 µL of CSF as the source of template was at best 10(3) parasites/mL. Remarkably, detergent enhanced LAMP assay reaches sensitivity about 100 to 1000-fold lower; i.e. 10 to 1 parasite/mL. Similar detergent-mediated increases in LAMP assay analytical sensitivity were also found using DNA extracted from filter paper cards containing blood pretreated with detergent before card spotting or blood samples spotted on detergent pretreated cards. This simple procedure for the enhanced detection of live African trypanosomes in biological fluids by LAMP paves the way for the adaptation of LAMP for the economical and sensitive diagnosis of other protozoan parasites and microorganisms that cause diseases that plague the developing world.

Diagnostic accuracy of loop-mediated isothermal amplification (LAMP for screening patients with imported malaria in a non-endemic setting

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Ponce Camille

2017-01-01

Full Text Available Background: Sensitive and easy-to-perform methods for the diagnosis of malaria are not yet available. Improving the limit of detection and following the requirements for certification are issues to be addressed in both endemic and non-endemic settings. The aim of this study was to test whether loop-mediated isothermal amplification of DNA (LAMP may be an alternative to microscopy or real-time PCR for the screening of imported malaria cases in non-endemic area. Results: 310 blood samples associated with 829 suspected cases of imported malaria were tested during a one year period. Microscopy (thin and thick stained blood slides, reference standard was used for the diagnosis. Real-time PCR was used as a standard of truth, and LAMP (Meridian Malaria Plus was used as an index test in a prospective study conducted following the Standards for Reporting Diagnosis Accuracy Studies. In the 83 positive samples, species identification was P. falciparum (n = 66, P. ovale (n = 9, P. vivax (n = 3 P. malariae (n = 3 and 2 co-infections with P. falciparum + P.malariae. Using LAMP methods, 93 samples gave positive results, including 4 false-positives. Sensitivity, specificity, positive predictive value and negative predictive value for LAMP tests were 100%, 98.13%, 95.51%, and 100% compared to PCR. Conclusion: High negative predictive value, and limit of detection suggest that LAMP can be used for screening of imported malaria cases in non-endemic countries when expert microscopists are not immediately available. However, the rare occurrence of non-valid results and the need for species identification and quantification of positive samples preclude the use of LAMP as a single reference method.

Detection of Zika virus using reverse-transcription LAMP coupled with reverse dot blot analysis in saliva.

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Maite Sabalza

Full Text Available In recent years, there have been increasing numbers of infectious disease outbreaks that spread rapidly to population centers resulting from global travel, population vulnerabilities, environmental factors, and ecological disasters such as floods and earthquakes. Some examples of the recent outbreaks are the Ebola epidemic in West Africa, Middle East respiratory syndrome coronavirus (MERS-Co in the Middle East, and the Zika outbreak through the Americas. We have created a generic protocol for detection of pathogen RNA and/or DNA using loop-mediated isothermal amplification (LAMP and reverse dot-blot for detection (RDB and processed automatically in a microfluidic device. In particular, we describe how a microfluidic assay to detect HIV viral RNA was converted to detect Zika virus (ZIKV RNA. We first optimized the RT-LAMP assay to detect ZIKV RNA using a benchtop isothermal amplification device. Then we implemented the assay in a microfluidic device that will allow analyzing 24 samples simultaneously and automatically from sample introduction to detection by RDB technique. Preliminary data using saliva samples spiked with ZIKV showed that our diagnostic system detects ZIKV RNA in saliva. These results will be validated in further experiments with well-characterized ZIKV human specimens of saliva. The described strategy and methodology to convert the HIV diagnostic assay and platform to a ZIKV RNA detection assay provides a model that can be readily utilized for detection of the next emerging or re-emerging infectious disease.

Miniaturized isothermal nucleic acid amplification, a review.

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Asiello, Peter J; Baeumner, Antje J

2011-04-21

Micro-Total Analysis Systems (µTAS) for use in on-site rapid detection of DNA or RNA are increasingly being developed. Here, amplification of the target sequence is key to increasing sensitivity, enabling single-cell and few-copy nucleic acid detection. The several advantages to miniaturizing amplification reactions and coupling them with sample preparation and detection on the same chip are well known and include fewer manual steps, preventing contamination, and significantly reducing the volume of expensive reagents. To-date, the majority of miniaturized systems for nucleic acid analysis have used the polymerase chain reaction (PCR) for amplification and those systems are covered in previous reviews. This review provides a thorough overview of miniaturized analysis systems using alternatives to PCR, specifically isothermal amplification reactions. With no need for thermal cycling, isothermal microsystems can be designed to be simple and low-energy consuming and therefore may outperform PCR in portable, battery-operated detection systems in the future. The main isothermal methods as miniaturized systems reviewed here include nucleic acid sequence-based amplification (NASBA), loop-mediated isothermal amplification (LAMP), helicase-dependent amplification (HDA), rolling circle amplification (RCA), and strand displacement amplification (SDA). Also, important design criteria for the miniaturized devices are discussed. Finally, the potential of miniaturization of some new isothermal methods such as the exponential amplification reaction (EXPAR), isothermal and chimeric primer-initiated amplification of nucleic acids (ICANs), signal-mediated amplification of RNA technology (SMART) and others is presented.

Loop-mediated isothermal amplification for rapid and convenient detection of Mycoplasma hyopneumoniae.

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Li, Jiahe; Minion, F Chris; Petersen, Andrew C; Jiang, Fei; Yang, Sheng; Guo, Panpan; Li, Jinxiang; Wu, Wenxue

2013-04-01

Loop-mediated isothermal amplification (LAMP), a novel method of gene amplification, was employed in this study for detecting Mycoplasma hyopneumoniae in the respiratory tract or lungs of swine. The pathogen can be detected in LAMP reactions containing as few as 10 fg purified target DNA (10 copies of M. hyopneumoniae genome) within 30 min, which was comparable to real-time PCR. After 30-min reaction at 63 °C, the addition of a certain amount of dye (SYBR Green I and hydroxyl naphthol blue at a proper ratio) into the LAMP reaction system makes the results easily determined as positive or negative by visual inspection. In addition, the LAMP was able to distinguish between M. hyopneumoniae and other closely-related mycoplasma strains, indicating a high degree of specificity. The LAMP assay was more simple and cheap, since the reaction could be completed under isothermal conditions and less laboratorial infrastructure are required. And, it was proven reliable for M. hyopneumoniae diagnosis of nasal swab and lung samples from the field.

Stability of Loop-Mediated Isothermal Amplification (LAMP) Reagents and its Amplification Efficiency on Crude Trypanosome DNA Templates

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Thekisoe, Oriel M. M; Bazie, Raoul S. B; Coronel-Servian, Andrea M; Sugimoto, Chihiro; Kawazu, Shin-ichiro; Inoue, Noboru

2009-01-01

This study evaluated the stability of LAMP reagents when stored at 25C and 37C, and also assessed its detection efficiency on different DNA template preparations. Accordingly, LAMP using reagents stored at 25C and 37C amplified DNA of in vitro cultured T. b. brucei (GUTat 3.1) from day 1 to day 15 of reagent storage. There were no significant differences (P>0.05) in detection sensitivity of LAMP among the reagents stored at 25C, 37C and –20C (recommended storage temperature). LAMP usin...

A novel CMOS image sensor system for quantitative loop-mediated isothermal amplification assays to detect food-borne pathogens.

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Wang, Tiantian; Kim, Sanghyo; An, Jeong Ho

2017-02-01

Loop-mediated isothermal amplification (LAMP) is considered as one of the alternatives to the conventional PCR and it is an inexpensive portable diagnostic system with minimal power consumption. The present work describes the application of LAMP in real-time photon detection and quantitative analysis of nucleic acids integrated with a disposable complementary-metal-oxide semiconductor (CMOS) image sensor. This novel system works as an amplification-coupled detection platform, relying on a CMOS image sensor, with the aid of a computerized circuitry controller for the temperature and light sources. The CMOS image sensor captures the light which is passing through the sensor surface and converts into digital units using an analog-to-digital converter (ADC). This new system monitors the real-time photon variation, caused by the color changes during amplification. Escherichia coli O157 was used as a proof-of-concept target for quantitative analysis, and compared with the results for Staphylococcus aureus and Salmonella enterica to confirm the efficiency of the system. The system detected various DNA concentrations of E. coli O157 in a short time (45min), with a detection limit of 10fg/μL. The low-cost, simple, and compact design, with low power consumption, represents a significant advance in the development of a portable, sensitive, user-friendly, real-time, and quantitative analytic tools for point-of-care diagnosis. Copyright © 2016 Elsevier B.V. All rights reserved.

Development of a new LAMP assay for the detection of CSFV strains from Cuba: a proof-of-concept study.

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Postel, Alexander; Pérez, Lester J; Perera, Carmen L; Schmeiser, Stefanie; Meyer, Denise; Meindl-Boehmer, Alexandra; Rios, Liliam; Austermann-Busch, Sophia; Frias-Lepoureau, Maria T; Becher, Paul

2015-06-01

Classical swine fever (CSF) is a devastating animal disease of great economic impact worldwide. In many countries, CSF has been endemic for decades, and vaccination of domestic pigs is one of the measures to control the disease. Consequently, differentiating infected from vaccinated animals by antibody ELISA screening is not applicable. In some countries, such as Cuba, lack of molecular techniques for sensitive, rapid and reliable detection of virus genomes is a critical point. To overcome this problem, an easy-to-use one-tube assay based on the loop-mediated isothermal amplification (LAMP) principle has been developed for detection of the genome of CSF virus (CSFV) of endemic Cuban genotype 1.4 isolates. The assay reliably detected recent isolates from three different regions of Cuba with an analytical sensitivity 10-100 times lower than that of quantitative reverse transcription RT-qPCR. Diagnostic test sensitivity was examined using reference sera from two groups of pigs experimentally infected with Cuban virulent strain CSF0705 "Margarita" and the recent field isolate CSF1058 "Pinar del Rio". Differences in pathogenicity of the two viruses were reflected in the clinical course of disease as well as in virus loads of blood samples. Low viral RNA loads in samples from pigs infected with the field isolate caused serious detection problems in RT-LAMP as well as in RT-qPCR. Thus, it will be necessary in future research to focus on targeted sampling of diseased animals and to restrict diagnosis to the herd level in order to establish LAMP as an efficient tool for diagnosing CSF under field conditions.

Stability of Loop-Mediated Isothermal Amplification (LAMP) reagents and its amplification efficiency on crude trypanosome DNA templates.

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Thekisoe, Oriel M M; Bazie, Raoul S B; Coronel-Servian, Andrea M; Sugimoto, Chihiro; Kawazu, Shin-Ichiro; Inoue, Noboru

2009-04-01

This study evaluated the stability of LAMP reagents when stored at 25 degrees C and 37 degrees C, and also assessed its detection efficiency on different DNA template preparations. Accordingly, LAMP using reagents stored at 25 degrees C and 37 degrees C amplified DNA of in vitro cultured T. b. brucei (GUTat 3.1) from day 1 to day 15 of reagent storage. There were no significant differences (P>0.05) in detection sensitivity of LAMP among the reagents stored at 25 degrees C, 37 degrees C and -20 degrees C (recommended storage temperature). LAMP using the reagents stored at above-mentioned temperatures amplified serially diluted DNAs (genomic DNA extracted by phenol-chloroform method, FTA card and hemolysed blood) of T. b. gambiense (IL2343) with high sensitivity. Reactions were conducted on the reagents stored from 1 day to 30 days. LAMP detection sensitivity was poor when fresh blood as DNA template was added directly into reactive solution. Results of this study demonstrated that LAMP has the potential to be used in field conditions for diagnosis of trypanosome infections without being affected by ambient temperatures of tropical and sub-tropical countries where trypanosomosis is endemic.

Diode-laser-illuminated automotive lamp systems

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Marinelli, Michael A.; Remillard, Jeffrey T.

1998-05-01

We have utilized the high brightness of state-of-the-art diode laser sources, and a variety of emerging optical technologies to develop a new class of thin, uniquely styled automotive brake and signal lamps. Using optics based on thin (5 mm) plastic sheets, these lamps provide appearance and functional advantages not attainable with traditional automotive lighting systems. The light is coupled into the sheets using a 1 mm diameter glass fiber, and manipulated using refraction and reflection from edges, surfaces, and shaped cut-outs. Light can be extracted with an efficiency of approximately 50% and formed into a luminance distribution that meets the Society of Automotive Engineers (SAE) photometric requirements. Prototype lamps using these optics have been constructed and are less than one inch in thickness. Thin lamps reduce sheet metal costs, complexity, material usage, weight, and allow for increased trunk volume. In addition, these optics enhance lamp design flexibility. When the lamps are not energized, they can appear body colored, and when lighted, the brightness distribution across the lamp can be uniform or structured. A diode laser based brake lamp consumes seven times less electrical power than one using an incandescent source and has instant on capability. Also, diode lasers have the potential to be 10-year/150,000 mile light sources.

Comparison of DNA Microarray, Loop-Mediated Isothermal Amplification (LAMP) and Real-Time PCR with DNA Sequencing for Identification of Fusarium spp. Obtained from Patients with Hematologic Malignancies.

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de Souza, Marcela; Matsuzawa, Tetsuhiro; Sakai, Kanae; Muraosa, Yasunori; Lyra, Luzia; Busso-Lopes, Ariane Fidelis; Levin, Anna Sara Shafferman; Schreiber, Angélica Zaninelli; Mikami, Yuzuru; Gonoi, Tohoru; Kamei, Katsuhiko; Moretti, Maria Luiza; Trabasso, Plínio

2017-08-01

The performance of three molecular biology techniques, i.e., DNA microarray, loop-mediated isothermal amplification (LAMP), and real-time PCR were compared with DNA sequencing for properly identification of 20 isolates of Fusarium spp. obtained from blood stream as etiologic agent of invasive infections in patients with hematologic malignancies. DNA microarray, LAMP and real-time PCR identified 16 (80%) out of 20 samples as Fusarium solani species complex (FSSC) and four (20%) as Fusarium spp. The agreement among the techniques was 100%. LAMP exhibited 100% specificity, while DNA microarray, LAMP and real-time PCR showed 100% sensitivity. The three techniques had 100% agreement with DNA sequencing. Sixteen isolates were identified as FSSC by sequencing, being five Fusarium keratoplasticum, nine Fusarium petroliphilum and two Fusarium solani. On the other hand, sequencing identified four isolates as Fusarium non-solani species complex (FNSSC), being three isolates as Fusarium napiforme and one isolate as Fusarium oxysporum. Finally, LAMP proved to be faster and more accessible than DNA microarray and real-time PCR, since it does not require a thermocycler. Therefore, LAMP signalizes as emerging and promising methodology to be used in routine identification of Fusarium spp. among cases of invasive fungal infections.

[Remote Slit Lamp Microscope Consultation System Based on Web].

Science.gov (United States)

Chen, Junfa; Zhuo, Yong; Liu, Zuguo; Chen, Yanping

2015-11-01

To realize the remote operation of the slit lamp microscope for department of ophthalmology consultation, and visual display the real-time status of remote slit lamp microscope, a remote slit lamp microscope consultation system based on B/S structure is designed and implemented. Through framing the slit lamp microscope on the website system, the realtime acquisition and transmission of remote control and image data is realized. The three dimensional model of the slit lamp microscope is established and rendered on the web by using WebGL technology. The practical application results can well show the real-time interactive of the remote consultation system.

Rapid and sensitive detection of Bartonella bacilliformis in experimentally infected sand flies by loop-mediated isothermal amplification (LAMP of the Pap31 gene.

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Nasikarn Angkasekwinai

2014-12-01

Full Text Available Carrion' disease, caused by Bartonella bacilliformis, remains truly neglected due to its focal geographical nature. A wide spectrum of clinical manifestations, including asymptomatic bacteremia, and lack of a sensitive diagnostic test can potentially lead to a spread of the disease into non-endemic regions where competent sand fly vectors may be present. A reliable test capable of detecting B. bacilliformis is urgently needed. Our objective is to develop a loop-mediated isothermal amplification (LAMP assay targeting the pap31 gene to detect B. bacilliformis.The sensitivity of the LAMP was evaluated in comparison to qPCR using plasmid DNA containing the target gene and genomic DNA in the absence and presence of human or sand fly DNA. The detection limit of LAMP was 1 to 10 copies/µL, depending on the sample metrics. No cross-reaction was observed when testing against a panel of various closely related bacteria. The utility of the LAMP was further compared to qPCR by the examination of 74 Lutzomyia longipalpis sand flies artificially fed on blood spiked with B. bacilliformis and harvested at days (D 1, 3, 5, 7 and 9 post feeding. Only 86% of sand flies at D1 and 63% of flies at D3 were positive by qPCR. LAMP was able to detect B. bacilliformis in all those flies confirmed positive by qPCR. However, none of the flies after D3 were positive by either LAMP or qPCR. In addition to demonstrating the sensitivity of the LAMP assay, these results suggest that B. bacilliformis cannot propagate in artificially fed L. longipalpis.The LAMP assay is as sensitive as qPCR for the detection of B. bacilliformis and could be useful to support diagnosis of patients in low-resource settings and also to identify B. bacilliformis in the sand fly vector.

LOOP mediated isothermal AMPlification (LAMP) in diagnosis of ...

African Journals Online (AJOL)

2012-02-28

Feb 28, 2012 ... Due to disadvantages of known ... advantages, as compared to traditional diagnostic methods like ... Products of LAMP reaction are DNA fragments with stemloop ..... "Differentiation of Cryptococcus neoformans varieties and.

Giardia and Cryptosporidium spp. dissemination during wastewater treatment and comparative detection via immunofluorescence assay (IFA), nested polymerase chain reaction (nested PCR) and loop mediated isothermal amplification (LAMP).

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Gallas-Lindemann, Carmen; Sotiriadou, Isaia; Plutzer, Judit; Noack, Michael J; Mahmoudi, Mohammad Reza; Karanis, Panagiotis

2016-06-01

Environmental water samples from the Lower Rhine area in Germany were investigated via immunofluorescence assays (IFAs), nested polymerase chain reaction (nested PCR) and loop-mediated isothermal amplification (LAMP) to detect the presence of Giardia spp. (n=185) and Cryptosporidium spp. (n=227). The samples were concentrated through filtration or flocculation, and oocysts were purified via centrifugation through a sucrose density gradient. For all samples, IFA was performed first, followed by DNA extraction for the nested PCR and LAMP assays. Giardia cysts were detected in 105 samples (56.8%) by IFA, 62 samples (33.5%) by nested PCR and 79 samples (42.7%) by LAMP. Cryptosporidium spp. were detected in 69 samples (30.4%) by IFA, 95 samples (41.9%) by nested PCR and 99 samples (43.6%) by LAMP. According to these results, the three detection methods are complementary for monitoring Giardia and Cryptosporidium in environmental waters. Copyright © 2016 Elsevier B.V. All rights reserved.

Rapid genome detection of Schmallenberg virus and bovine viral diarrhea virus by use of isothermal amplification methods and high-speed real-time reverse transcriptase PCR.

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Aebischer, Andrea; Wernike, Kerstin; Hoffmann, Bernd; Beer, Martin

2014-06-01

Over the past few years, there has been an increasing demand for rapid and simple diagnostic tools that can be applied outside centralized laboratories by using transportable devices. In veterinary medicine, such mobile test systems would circumvent barriers associated with the transportation of samples and significantly reduce the time to diagnose important infectious animal diseases. Among a wide range of available technologies, high-speed real-time reverse transcriptase quantitative PCR (RT-qPCR) and the two isothermal amplification techniques loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA) represent three promising candidates for integration into mobile pen-side tests. The aim of this study was to investigate the performance of these amplification strategies and to evaluate their suitability for field application. In order to enable a valid comparison, novel pathogen-specific assays have been developed for the detection of Schmallenberg virus and bovine viral diarrhea virus. The newly developed assays were evaluated in comparison with established standard RT-qPCR using samples from experimentally or field-infected animals. Even though all assays allowed detection of the target virus in less than 30 min, major differences were revealed concerning sensitivity, specificity, robustness, testing time, and complexity of assay design. These findings indicated that the success of an assay will depend on the integrated amplification technology. Therefore, the application-specific pros and cons of each method that were identified during this study provide very valuable insights for future development and optimization of pen-side tests. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Detection of early and single infections of Schistosoma japonicum in the intermediate host snail, Oncomelania hupensis, by PCR and loop-mediated isothermal amplification (LAMP) assay.

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Kumagai, Takashi; Furushima-Shimogawara, Rieko; Ohmae, Hiroshi; Wang, Tian-Ping; Lu, Shaohong; Chen, Rui; Wen, Liyong; Ohta, Nobuo

2010-09-01

Polymerase chain reaction (PCR) with the specific primer set amplifying 28S ribosomal DNA (rDNA) of Schistosoma japonicum was able to detect genomic DNA of S. japonicum, but not S. mansoni, at 100 fg. This procedure enabled us to detect the DNA from a single miracidium and a snail infected with one miracidium at just 1 day after infection. We compared these results with those from loop-mediated isothermal amplification (LAMP) targeting 28S rDNA and found similar results. The LAMP could amplify the specific DNA from a group of 100 normal snails mixed with one infected snail A PCR screening of infected snails from endemic regions in Anhui Province revealed schistosomal DNA even in snails found negative by microscopy. PCR and LAMP show promise for monitoring the early infection rate in snails, and they may be useful for predicting the risk of infection in the endemic places.

The development and evaluation of a loop-mediated isothermal amplification (LAMP) method for detection of Babesia spp. infective to sheep and goats in China.

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Guan, Guiquan; Chauvin, Alain; Luo, Jianxun; Inoue, Noboru; Moreau, Emmanuelle; Liu, Zhijie; Gao, Jinliang; Thekisoe, Oriel M M; Ma, Miling; Liu, Aihong; Dang, Zhisheng; Liu, Junlong; Ren, Qiaoyun; Jin, Yurong; Sugimoto, Chihiro; Yin, Hong

2008-09-01

The loop-mediated isothermal amplification (LAMP) reaction is a method that amplifies with high sensitivity, efficiency, and rapidity, deoxyribonucleic acid (DNA) under isothermal condition in simple incubators. Two primer sets for the LAMP method were designed using the nucleotide sequences of 18S rRNA gene of Babesia sp. BQ1 (Lintan) and Babesia sp. Xinjiang-2005 isolated in China. The primers were used to detect parasite DNA extracted from infected blood and purified parasites by LAMP. The specific ladder bands were amplified from the autologous genomic DNA of two Babesia species, respectively, and did not cross-react with the genomic DNA of Theileria sp. China 1, Theileria sp. China 2, B. bovis, Theileria sp. (Japan) and sheep. The LAMP was sensitive enough to detect 0.02 pg and 0.2 pg genomic DNA of Babesia sp. BQ1 (Lintan) and Babesia sp. Xinjiang-2005, respectively, from 10-fold serially diluted samples corresponding to the amount of DNA present in 50 microl of 0.000002% and 0.00002% parasitemic erythrocytes. Furthermore, DNA extracted from blood of intact (non-splenectomized) sheep experimentally infected with Babesia sp. BQ1 (Lintan) and Babesia sp. Xinjiang-2005 was amplified by the LAMP from week 1 to 9 and week 2 and 3 post-infection, respectively, demonstrating the high sensitivity of these primers. Of 365 samples collected from Gansu province, 14.3% (52/365) were positively detected by the LAMP. Of 145 samples collected on filter papers (Whatman) from the grazing sheep in Xinjiang province, 3.5% (5/145) were positive. These results show that the LAMP could be an alternative diagnostic tool for the detection of babesial infection in sheep and goats.

Rapid screening method for male DNA by using the loop-mediated isothermal amplification assay.

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Kitamura, Masashi; Kubo, Seiji; Tanaka, Jin; Adachi, Tatsushi

2017-08-12

Screening for male-derived biological material from collected samples plays an important role in criminal investigations, especially those involving sexual assaults. We have developed a loop-mediated isothermal amplification (LAMP) assay targeting multi-repeat sequences of the Y chromosome for detecting male DNA. Successful amplification occurred with 0.5 ng of male DNA under isothermal conditions of 61 to 67 °C, but no amplification occurred with up to 10 ng of female DNA. Under the optimized conditions, the LAMP reaction initiated amplification within 10 min and amplified for 20 min. The LAMP reaction was sensitive at levels as low as 1-pg male DNA, and a quantitative LAMP assay could be developed because of the strong correlation between the reaction time and the amount of template DNA in the range of 10 pg to 10 ng. Furthermore, to apply the LAMP assay to on-site screening for male-derived samples, we evaluated a protocol using a simple DNA extraction method and a colorimetric intercalating dye that allows detection of the LAMP reaction by evaluating the change in color of the solution. Using this protocol, samples of male-derived blood and saliva stains were processed in approximately 30 min from DNA extraction to detection. Because our protocol does not require much hands-on time or special equipment, this LAMP assay promises to become a rapid and simple screening method for male-derived samples in forensic investigations.

Allele specific LAMP- gold nanoparticle for characterization of single nucleotide polymorphisms

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Fábio Ferreira Carlos

2017-12-01

Full Text Available Due to their relevance as disease biomarkers and for diagnostics, screening of single nucleotide polymorphism (SNPs requires simple and straightforward strategies capable to provide results in medium throughput settings. Suitable approaches relying on isothermal amplification techniques have been evolving to substitute the cumbersome and highly specialized PCR amplification detection schemes. Nonetheless, identification of an individual’s genotype still requires sophisticated equipment and laborious methods.Here, we present a low-cost and reliable approach based on the allele specific loop-mediated isothermal amplification (AS-LAMP coupled to ssDNA functionalized gold nanoparticle (Au-nanoprobe colorimetric sequence discrimination. The Au-nanoprobe integration allows for the colorimetric detection of AS-LAMP amplification product that can be easily interpreted in less than 15 min. We targeted a clinical relevant SNP responsible for lactose intolerance (-13910C/T dbSNP rs#: 4988235 to demonstrate its proof of concept and full potential of this novel approach. Keywords: SNP, Isothermal amplification, Gold nanoparticles, Gold nanoprobes, Lactose intolerance

A comparative study of three different gene expression analysis methods.

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Choe, Jae Young; Han, Hyung Soo; Lee, Seon Duk; Lee, Hanna; Lee, Dong Eun; Ahn, Jae Yun; Ryoo, Hyun Wook; Seo, Kang Suk; Kim, Jong Kun

2017-12-04

TNF-α regulates immune cells and acts as an endogenous pyrogen. Reverse transcription polymerase chain reaction (RT-PCR) is one of the most commonly used methods for gene expression analysis. Among the alternatives to PCR, loop-mediated isothermal amplification (LAMP) shows good potential in terms of specificity and sensitivity. However, few studies have compared RT-PCR and LAMP for human gene expression analysis. Therefore, in the present study, we compared one-step RT-PCR, two-step RT-LAMP and one-step RT-LAMP for human gene expression analysis. We compared three gene expression analysis methods using the human TNF-α gene as a biomarker from peripheral blood cells. Total RNA from the three selected febrile patients were subjected to the three different methods of gene expression analysis. In the comparison of three gene expression analysis methods, the detection limit of both one-step RT-PCR and one-step RT-LAMP were the same, while that of two-step RT-LAMP was inferior. One-step RT-LAMP takes less time, and the experimental result is easy to determine. One-step RT-LAMP is a potentially useful and complementary tool that is fast and reasonably sensitive. In addition, one-step RT-LAMP could be useful in environments lacking specialized equipment or expertise.

Shining a light on LAMP assays--a comparison of LAMP visualization methods including the novel use of berberine.

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Fischbach, Jens; Xander, Nina Carolin; Frohme, Marcus; Glökler, Jörn Felix

2015-04-01

The need for simple and effective assays for detecting nucleic acids by isothermal amplification reactions has led to a great variety of end point and real-time monitoring methods. Here we tested direct and indirect methods to visualize the amplification of potato spindle tuber viroid (PSTVd) by loop-mediated isothermal amplification (LAMP) and compared features important for one-pot in-field applications. We compared the performance of magnesium pyrophosphate, hydroxynaphthol blue (HNB), calcein, SYBR Green I, EvaGreen, and berberine. All assays could be used to distinguish between positive and negative samples in visible or UV light. Precipitation of magnesium-pyrophosphate resulted in a turbid reaction solution. The use of HNB resulted in a color change from violet to blue, whereas calcein induced a change from orange to yellow-green. We also investigated berberine as a nucleic acid-specific dye that emits a fluorescence signal under UV light after a positive LAMP reaction. It has a comparable sensitivity to SYBR Green I and EvaGreen. Based on our results, an optimal detection method can be chosen easily for isothermal real-time or end point screening applications.

Tuberculosis Biomarker Extraction and Isothermal Amplification in an Integrated Diagnostic Device.

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Amy Creecy

Full Text Available In this study, we integrated magnetic bead-based sample preparation and isothermal loop mediated amplification (LAMP of TB in a single tube. Surrogate sputum samples produced by the Program for Appropriate Technology in Health containing inactivated TB bacteria were used to test the diagnostic. In order to test the sample preparation method, samples were lysed, and DNA was manually extracted and eluted into water in the tube. In a thermal cycler, LAMP amplified TB DNA from 103 TB cells/mL of sputum at 53.5 ± 3.3 minutes, 104 cells/mL at 46.3 ± 2.2 minutes, and 105 cells/mL at 41.6 ± 1.9 minutes. Negative control samples did not amplify. Next, sample preparation was combined with in-tubing isothermal LAMP amplification by replacing the water elution chamber with a LAMP reaction chamber. In this intermediate configuration, LAMP amplified 103 cells/mL at 74 ± 10 minutes, 104 cells/mL at 60 ± 9 minutes, and 105 TB cells/mL of sputum at 54 ± 9 minutes. Two of three negative controls did not amplify; one amplified at 100 minutes. In the semi-automated system, DNA was eluted directly into an isothermal reaction solution containing the faster OptiGene DNA polymerase. The low surrogate sputum concentration, 103 TB cells/mL, amplified at 52.8 ± 3.3 minutes, 104 cells/mL at 45.4 ± 11.3 minutes, and 105 cells/mL at 31.8 ± 2.9 minutes. TB negative samples amplified at 66.4 ± 7.4 minutes. This study demonstrated the feasibility of a single tube design for integrating sample preparation and isothermal amplification, which with further development could be useful for point-of-care applications, particularly in a low-resource setting.

LOOP mediated isothermal AMPlification (LAMP) in diagnosis of ...

African Journals Online (AJOL)

We collected a quantity of 35 serum samples of HIVpositive patients and a number of 107 cerebrospinal fluid (CSF) samples of patients who had shown symptoms of meningitis. We designed target specific primers for PCR and LAMP techniques to trace C. neoformans and C. gattii. From the total 142 clinical specimens, five ...

Development of an allele-specific, loop-mediated, isothermal amplification method (AS-LAMP to detect the L1014F kdr-w mutation in Anopheles gambiae s. l.

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Badolo Athanase

2012-07-01

Full Text Available Abstract Background Malaria control relies heavily on treated bed nets and indoor residual spraying with pyrethroid insecticides. Unfortunately, the resistance to pyrethroid insecticides, mainly due to the kdr mutation, is spreading in the main malaria vector Anopheles gambiae s.l., decreasing the insecticides’ efficacy. To manage the insecticide resistance rapidly and flexibly, simple and effective tools for the early detection of resistant mosquitoes are needed. This study aimed to develop an allele-specific, loop-mediated, isothermal amplification (AS-LAMP method to detect the West African-type kdr mutation (kdr-w; L1014F in field-collected mosquitoes. Methods DNA fragments of the wild-type and the mutated kdr gene were used to select the primers and develop the method. The primers were designed with the mutation at the 5’ end of the backward inner primer (BIP. The AS-LAMP method was compared to the AS-PCR method using the genomic DNA of 120 field-collected mosquitoes. Results The AS-LAMP method could discriminate between the wild-type homozygote, the heterozygote, and the kdr-w homozygote within 75 min. The AS-LAMP method has the advantage of being faster and at least as sensitive and specific as the AS-PCR method. Conclusions The AS-LAMP method can be used to detect the kdr mutation for quick decision-making, even in less well-equipped laboratories.

Rapid, sensitive and effective diagnostic tools for foot-and-mouth disease virus in Africa

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Christopher J. Kasanga

2014-04-01

Full Text Available Speed is paramount in the diagnosis of highly infectious diseases, such as foot-and-mouth disease (FMD, as well as for emerging diseases; however, simplicity is required if a test is to be deployed in the field. Recent developments in molecular biology have enabled the specific detection of FMD virus (FMDV by reverse-transcription loop-mediated isothermal amplification (RT-LAMP, real-time  reverse-transcription polymerase chain reaction (RT-qPCR and sequencing. RT-LAMP enables amplification of the FMDV RNA-dependent RNA polymerase 3D(pol gene at 63 °C (in the presence of a primer mixture and both reverse transcriptase and Bst DNA polymerase for 1 h, whilst RT-qPCR amplifies the same gene in approximately 2 h 30 min. In this study, we compared the sensitivity and effectiveness of RT-LAMP against RT-qPCR for the detection of the FMDV 3D(pol gene in 179 oesophageal-pharyngeal scraping samples (collected by probang obtained from clinically healthy cattle and buffalo in Malawi, Mozambique and Tanzania in 2010. The FMDV detection rate was higher with RT-LAMP (30.2%; n = 54 than with RT-qPCR (17.3%; n = 31. All samples positive by RT-qPCR (Cq ≤ 32.0 were also positive for the RT-LAMP assay; and both assays proved to be highly specific for the FMDV target sequence. In addition, the VP1 sequences of 10 viruses isolated from positive samples corresponded to the respective FMDV serotypes and genotypes. Our findings indicate that the performance of RT-LAMP is superior to RT-qPCR. Accordingly, we consider this test to have great potential with regard to the specific detection and surveillance of infectious diseases of humans and animals in resource-compromised developing countries.

Development and evaluation of a Loop Mediated Isothermal Amplification (LAMP) technique for the detection of hookworm (Necator americanus) infection in fecal samples.

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Mugambi, Robert Muriuki; Agola, Eric L; Mwangi, Ibrahim N; Kinyua, Johnson; Shiraho, Esther Andia; Mkoji, Gerald M

2015-11-06

Hookworm infection is a major concern in sub-Saharan Africa, particularly in children and pregnant women. Necator americanus and Ancylostoma duodenale are responsible for this condition. Hookworm disease is one of the Neglected tropical diseases (NTDs) that are targeted for elimination through global mass chemotherapy. To support this there is a need for reliable diagnostic tools. The conventional diagnostic test, Kato-Katz that is based on microscopic detection of parasite ova in faecal samples, is not effective due to its low sensitivity that is brought about mainly by non-random distribution of eggs in stool and day to day variation in egg output. It is tedious, cumbersome to perform and requires experience for correct diagnosis. LAMP-based tests are simple, relatively cheap, offer greater sensitivity, specificity than existing tests, have high throughput capability, and are ideal for use at the point of care. We have developed a LAMP diagnostic test for detection of hookworm infection in faecal samples. LAMP relies on auto cycling strand displacement DNA synthesis performed at isothermal temperature by Bst polymerase and a set of 4 specific primers. The primers used in the LAMP assay were based on the second Internal Transcribed Spacer (ITS-2) region and designed using Primer Explorer version 4 Software. The ITS-2 region of the ribosomal gene (rDNA) was identified as a suitable target due to its low mutation rates and substantial differences between species. DNA was extracted directly from human faecal samples, followed by LAMP amplification at isothermal temperature of 63 °C for 1 h. Amplicons were visualized using gel electrophoresis and SYBR green dye. Both specificity and sensitivity of the assay were determined. The LAMP based technique developed was able to detect N. americanus DNA in faecal samples. The assay showed 100 % specificity and no cross-reaction was observed with other helminth parasites (S. mansoni, A. lumbricoides or T. trichiura). The

Modelling the Dynamic Interaction Power System Lamp - Application to High Pressure Mercury Gas Discharge Lamps

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ZIANE, M.

2007-11-01

Full Text Available The aim of this paper is to study the dynamic behaviour of a plant constituted by an electrical power system and a gas discharge lamp, this latter, increasingly used in street lighting, remains a nonlinear load element. Various approaches are used to represent it, one is the approximation of the discharge represented by a hot "channel", which verifies the assumption of local thermodynamic equilibrium [LTE] or the polynomial form of the conductance variation. A calculation procedure, based on "channel" approximation of the high pressure mercury (HPM gas-discharge lamp, is developed to determine the physical and electric magnitudes, which characterize the dynamic behavior of the couple "lamp-electrical power system". The evolution of the lamp properties when principal parameters of the discharge (pressure of mercury, voltage supply, frequency are varying were studied and analyzed. We show the concordance between simulation, calculations and measurements for electric, energetic or irradiative characteristics. The model reproduces well the evolution of properties of the supply when principal parameters of the discharge vary.

Development of a loop-mediated isothermal amplification assay for rapid detection of BK virus.

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Bista, Bipin Raj; Ishwad, Chandra; Wadowsky, Robert M; Manna, Pradip; Randhawa, Parmjeet Singh; Gupta, Gaurav; Adhikari, Meena; Tyagi, Rakhi; Gasper, Gina; Vats, Abhay

2007-05-01

Loop-mediated isothermal amplification (LAMP) is a novel method for rapid amplification of DNA. Its advantages include rapidity and minimal equipment requirement. The LAMP assay was developed for BK virus (BKV), which is a leading cause of morbidity in renal transplant recipients. The characteristics of the assay, including its specificity and sensitivity, were evaluated. BKV LAMP was performed using various incubation times with a variety of specimens, including unprocessed urine and plasma samples. A ladder pattern on gel electrophoresis, typical of successful LAMP reactions, was observed specifically only for BKV and not for other viruses. The sensitivity of the assay with 1 h of incubation was 100 copies/tube of a cloned BKV fragment. Additionally, a positive reaction was visually ascertained by a simple color reaction using SYBR green dye. BKV LAMP was also successful for urine and plasma specimens without the need for DNA extraction. Due to its simplicity and specificity, the LAMP assay can potentially be developed for "point of care" screening of BKV.

Loop-mediated isothermal amplification (LAMP) assay-A rapid detection tool for identifying red fox (Vulpes vulpes) DNA in the carcasses of harbour porpoises (Phocoena phocoena).

Science.gov (United States)

Heers, Teresa; van Neer, Abbo; Becker, André; Grilo, Miguel Luca; Siebert, Ursula; Abdulmawjood, Amir

2017-01-01

Carcasses of wild animals are often visited by different scavengers. However, determining which scavenger caused certain types of bite marks is particularly difficult and knowledge thereof is lacking. Therefore, a loop-mediated isothermal amplification (LAMP) assay (target sequence cytochrome b) was developed to detect red fox DNA in carcasses of harbour porpoises. The MSwab™ method for direct testing without prior DNA isolation was validated. As a detection device, the portable real-time fluorometer Genie® II was used, which yields rapid results and can be used in field studies without huge laboratory equipment. In addition to in vitro evaluation and validation, a stranded and scavenged harbour porpoise carcass was successfully examined for red fox DNA residues. The developed LAMP method is a valuable diagnostic tool for confirming presumable red fox bite wounds in harbour porpoises without further DNA isolation steps.

A Portable Automatic Endpoint Detection System for Amplicons of Loop Mediated Isothermal Amplification on Microfluidic Compact Disk Platform

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Shah Mukim Uddin

2015-03-01

Full Text Available In recent years, many improvements have been made in foodborne pathogen detection methods to reduce the impact of food contamination. Several rapid methods have been developed with biosensor devices to improve the way of performing pathogen detection. This paper presents an automated endpoint detection system for amplicons generated by loop mediated isothermal amplification (LAMP on a microfluidic compact disk platform. The developed detection system utilizes a monochromatic ultraviolet (UV emitter for excitation of fluorescent labeled LAMP amplicons and a color sensor to detect the emitted florescence from target. Then it processes the sensor output and displays the detection results on liquid crystal display (LCD. The sensitivity test has been performed with detection limit up to 2.5 × 10−3 ng/µL with different DNA concentrations of Salmonella bacteria. This system allows a rapid and automatic endpoint detection which could lead to the development of a point-of-care diagnosis device for foodborne pathogens detection in a resource-limited environment.

LED lamp power management system and method

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Gaines, James; Clauberg, Bernd; Van Erp, Josephus A. M.

2013-03-19

An LED lamp power management system and method including an LED lamp having an LED controller 58; a plurality of LED channels 60 operably connected to the LED controller 58, each of the plurality of LED channels 60 having a channel switch 62 in series with at least one shunted LED circuit 83, the shunted LED circuit 83 having a shunt switch 68 in parallel with an LED source 80. The LED controller 58 reduces power loss in one of the channel switch 62 and the shunt switch 68 when LED lamp electronics power loss (P.sub.loss) exceeds an LED lamp electronics power loss limit (P.sub.lim); and each of the channel switches 62 receives a channel switch control signal 63 from the LED controller 58 and each of the shunt switches 68 receives a shunt switch control signal 69 from the LED controller 58.

Application of loop-mediated isothermal amplification (LAMP) of the ...

African Journals Online (AJOL)

Administrator

2011-05-23

May 23, 2011 ... evaluate the RIME - LAMP for the detection of T. evansi in naturally infected camels in Sudan. MATERIALS AND METHODS. Study design. Cross sectional ... from each camel using 5 ml disposable syringe. Three drops of the fresh collected blood were immediately placed in FTA card. (Whatman classic ...

Detection of genetically modified organisms (GMOs using isothermal amplification of target DNA sequences

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La Mura Maurizio

2009-02-01

Full Text Available Abstract Background The most common method of GMO detection is based upon the amplification of GMO-specific DNA amplicons using the polymerase chain reaction (PCR. Here we have applied the loop-mediated isothermal amplification (LAMP method to amplify GMO-related DNA sequences, 'internal' commonly-used motifs for controlling transgene expression and event-specific (plant-transgene junctions. Results We have tested the specificity and sensitivity of the technique for use in GMO studies. Results show that detection of 0.01% GMO in equivalent background DNA was possible and dilutions of template suggest that detection from single copies of the template may be possible using LAMP. Conclusion This work shows that GMO detection can be carried out using LAMP for routine screening as well as for specific events detection. Moreover, the sensitivity and ability to amplify targets, even with a high background of DNA, here demonstrated, highlights the advantages of this isothermal amplification when applied for GMO detection.

Detection of genetically modified organisms (GMOs) using isothermal amplification of target DNA sequences.

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Lee, David; La Mura, Maurizio; Allnutt, Theo R; Powell, Wayne

2009-02-02

The most common method of GMO detection is based upon the amplification of GMO-specific DNA amplicons using the polymerase chain reaction (PCR). Here we have applied the loop-mediated isothermal amplification (LAMP) method to amplify GMO-related DNA sequences, 'internal' commonly-used motifs for controlling transgene expression and event-specific (plant-transgene) junctions. We have tested the specificity and sensitivity of the technique for use in GMO studies. Results show that detection of 0.01% GMO in equivalent background DNA was possible and dilutions of template suggest that detection from single copies of the template may be possible using LAMP. This work shows that GMO detection can be carried out using LAMP for routine screening as well as for specific events detection. Moreover, the sensitivity and ability to amplify targets, even with a high background of DNA, here demonstrated, highlights the advantages of this isothermal amplification when applied for GMO detection.

Development of a Loop-Mediated Isothermal Amplification Assay for Rapid Detection of BK Virus▿

Science.gov (United States)

Bista, Bipin Raj; Ishwad, Chandra; Wadowsky, Robert M.; Manna, Pradip; Randhawa, Parmjeet Singh; Gupta, Gaurav; Adhikari, Meena; Tyagi, Rakhi; Gasper, Gina; Vats, Abhay

2007-01-01

Loop-mediated isothermal amplification (LAMP) is a novel method for rapid amplification of DNA. Its advantages include rapidity and minimal equipment requirement. The LAMP assay was developed for BK virus (BKV), which is a leading cause of morbidity in renal transplant recipients. The characteristics of the assay, including its specificity and sensitivity, were evaluated. BKV LAMP was performed using various incubation times with a variety of specimens, including unprocessed urine and plasma samples. A ladder pattern on gel electrophoresis, typical of successful LAMP reactions, was observed specifically only for BKV and not for other viruses. The sensitivity of the assay with 1 h of incubation was 100 copies/tube of a cloned BKV fragment. Additionally, a positive reaction was visually ascertained by a simple color reaction using SYBR green dye. BKV LAMP was also successful for urine and plasma specimens without the need for DNA extraction. Due to its simplicity and specificity, the LAMP assay can potentially be developed for “point of care” screening of BKV. PMID:17314224

An optimized protocol for DNA extraction from wheat seeds and Loop-Mediated Isothermal Amplification (LAMP) to detect Fusarium graminearum contamination of wheat grain.

Science.gov (United States)

Abd-Elsalam, Kamel; Bahkali, Ali; Moslem, Mohamed; Amin, Osama E; Niessen, Ludwig

2011-01-01

A simple, rapid, and efficient method for isolating genomic DNA from germinated seeds of wheat that is free from polysaccharides and polyphenols is reported. DNA was extracted, treated with RNase, measured and tested for completeness using agarose gel electrophoresis. DNA purification from wheat grains yielded abundant, amplifiable DNA with yields typically between 100 and 200 ng DNA/mg. The effectiveness and reliability of the method was tested by assessing quantity and quality of the isolated DNA using three PCR-based markers. Inter-simple sequence repeats (ISSRs) were used to assess the genetic diversity between different wheat varieties. Specific PCR primer pair Tox5-1/Tox5-2 and a loop-mediated isothermal amplification (LAMP) procedure were used to detect genomic DNA of Fusarium graminearum in contaminated wheat seeds. In this method there is no need to use liquid nitrogen for crushing germinated seedlings. The protocol takes approximately one hour to prepare high quality DNA. In combination with the LAMP assay it is a fast and cost-effective alternative to traditional diagnostic methods for the early detection of toxigenic fusaria in cereals.

Loop-mediated isothermal amplification (LAMP) as an alternative to PCR: A rapid on-site detection of gene doping.

Science.gov (United States)

Salamin, Olivier; Kuuranne, Tiia; Saugy, Martial; Leuenberger, Nicolas

2017-11-01

Innovation in medical research has been diverted at multiple occasions to enhance human performance. The predicted great progress in gene therapy has raised some concerns regarding its misuse in the world of sports (gene doping) for several years now. Even though there is no evidence that gene doping has ever been used in sports, the continuous improvement of gene therapy techniques increases the likelihood of abuse. Therefore, since 2004, efforts have been invested by the anti-doping community and WADA for the development of detection methods. Several nested PCR and qPCR-based strategies exploiting the absence of introns in the transgenic DNA have been proposed for the long-term detection of transgene in blood. Despite their great sensitivity, those protocols are hampered by limitations of the techniques that can be cumbersome and costly. The purpose of this perspective is to describe a new approach based on loop-mediated isothermal amplification (LAMP) for the detection of gene doping. This protocol enables a rapid and simple method to amplify nucleic acids with a high sensitivity and specificity and with a simple visual detection of the results. LAMP is already being used in clinical application for the detection of viruses or mutations. Therefore, this technique has the potential to be further developed for the detection of foreign genetic material in elite athletes. Copyright © 2017 John Wiley & Sons, Ltd. Copyright © 2017 John Wiley & Sons, Ltd.

GMO detection in food and feed through screening by visual loop-mediated isothermal amplification assays.

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Wang, Cong; Li, Rong; Quan, Sheng; Shen, Ping; Zhang, Dabing; Shi, Jianxin; Yang, Litao

2015-06-01

Isothermal DNA/RNA amplification techniques are the primary methodology for developing on-spot rapid nucleic acid amplification assays, and the loop-mediated isothermal amplification (LAMP) technique has been developed and applied in the detection of foodborne pathogens, plant/animal viruses, and genetically modified (GM) food/feed contents. In this study, one set of LAMP assays targeting on eight frequently used universal elements, marker genes, and exogenous target genes, such as CaMV35S promoter, FMV35S promoter, NOS, bar, cry1Ac, CP4 epsps, pat, and NptII, were developed for visual screening of GM contents in plant-derived food samples with high efficiency and accuracy. For these eight LAMP assays, their specificity was evaluated by testing commercial GM plant events and their limits of detection were also determined, which are 10 haploid genome equivalents (HGE) for FMV35S promoter, cry1Ac, and pat assays, as well as five HGE for CaMV35S promoter, bar, NOS terminator, CP4 epsps, and NptII assays. The screening applicability of these LAMP assays was further validated successfully using practical canola, soybean, and maize samples. The results suggested that the established visual LAMP assays are applicable and cost-effective for GM screening in plant-derived food samples.

Rapid and specific identification of Brucella abortus using the loop-mediated isothermal amplification (LAMP) assay.

Science.gov (United States)

Kang, Sung-Il; Her, Moon; Kim, Ji-Yeon; Lee, Jin Ju; Lee, Kichan; Sung, So-Ra; Jung, Suk Chan

2015-06-01

A rapid and accurate diagnosis of brucellosis is required to reduce and prevent the spread of disease among animals and the risk of transfer to humans. In this study, a Brucella abortus-specific (Ba) LAMP assay was developed, that had six primers designed from the BruAb2_0168 region of chromosome I. The specificity of this LAMP assay was confirmed with Brucella reference strains, B. abortus vaccine strains, B. abortus isolates and phylogenetically or serologically related strains. The detection limit of target DNA was up to 20 fg/μl within 60 min. The sensitivity of the new LAMP assay was equal to or slightly higher than other PCR based assays. Moreover, this Ba-LAMP assay could specifically amplify all B. abortus biovars compared to previous PCR assays. To our knowledge, this is the first report of specific detection of B. abortus using a LAMP assay. The Ba-LAMP assay can offer a rapid, sensitive and accurate diagnosis of bovine brucellosis in the field. Copyright © 2015 Elsevier Ltd. All rights reserved.

[Colorimetric detection of HPV6 and HPV16 by loop mediated isothermal amplification].

Science.gov (United States)

Lu, Chun-bin; Luo, Le; Yang, Meng-jie; Nie, Kai; Wang, Miao; Ma, Xue-Jun

2011-01-01

A simple, rapid and sensitive colorimetric loop mediated isothermal amplification (LAMP) method was established to detect HPV6 and HPV 16 respectively. The method employed a set of four specially designed primers that recognized six distinct sequences of HPV6-E6 or HPV16-E7 for amplification of nucleic acid under isothermal conditions at 63 degrees C for one hour. The amplification process of LAMP was monitored by the addition of HNB (hydroxy naphthol blue) dye prior to amplification. A positive reaction was indicated by a color change from violet to sky blue and confirmed by real-time turbidimeter and agarose electrophoresis. Thirteen cervical swab samples having single infection with 13 different HPV genotypes were examined to evaluate the specificity. A serial dilution of a cloned plasmid containing HPV-E6 or HPV-E7 gene was examined to evaluate the sensitivity. The results showed that no cross-reaction with other HPV genotypes was observed. The colorimetric LAMP assay could achieve a sensitivity of 1000 copies, 10-20 times lower than that of real-time PCR. The assay was further evaluated with 62 clinical specimens and consistent results were obtained compared with the detection using Kai Pu HPV Genotyping Kit. We concluded that this colorimetric LAMP assay had potential usefulness for the rapid screening of the HPV6 or HPV16 infection in the laboratories and hospitals of provincial and municipal region in China.

Identification and differentiation of the twenty six bluetongue virus serotypes by RT-PCR amplification of the serotype-specific genome segment 2.

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Narender S Maan

Full Text Available Bluetongue (BT is an arthropod-borne viral disease, which primarily affects ruminants in tropical and temperate regions of the world. Twenty six bluetongue virus (BTV serotypes have been recognised worldwide, including nine from Europe and fifteen in the United States. Identification of BTV serotype is important for vaccination programmes and for BTV epidemiology studies. Traditional typing methods (virus isolation and serum or virus neutralisation tests (SNT or VNT are slow (taking weeks, depend on availability of reference virus-strains or antisera and can be inconclusive. Nucleotide sequence analyses and phylogenetic comparisons of genome segment 2 (Seg-2 encoding BTV outer-capsid protein VP2 (the primary determinant of virus serotype were completed for reference strains of BTV-1 to 26, as well as multiple additional isolates from different geographic and temporal origins. The resulting Seg-2 database has been used to develop rapid (within 24 h and reliable RT-PCR-based typing assays for each BTV type. Multiple primer-pairs (at least three designed for each serotype were widely tested, providing an initial identification of serotype by amplification of a cDNA product of the expected size. Serotype was confirmed by sequencing of the cDNA amplicons and phylogenetic comparisons to previously characterised reference strains. The results from RT-PCR and sequencing were in perfect agreement with VNT for reference strains of all 26 BTV serotypes, as well as the field isolates tested. The serotype-specific primers showed no cross-amplification with reference strains of the remaining 25 serotypes, or multiple other isolates of the more closely related heterologous BTV types. The primers and RT-PCR assays developed in this study provide a rapid, sensitive and reliable method for the identification and differentiation of the twenty-six BTV serotypes, and will be updated periodically to maintain their relevance to current BTV distribution and

Detection of the 35S promoter in transgenic maize via various isothermal amplification techniques: a practical approach.

Science.gov (United States)

Zahradnik, Celine; Kolm, Claudia; Martzy, Roland; Mach, Robert L; Krska, Rudolf; Farnleitner, Andreas H; Brunner, Kurt

2014-11-01

In 2003 the European Commission introduced a 0.9% threshold for food and feed products containing genetically modified organism (GMO)-derived components. For commodities containing GMO contents higher than this threshold, labelling is mandatory. To provide a DNA-based rapid and simple detection method suitable for high-throughput screening of GMOs, several isothermal amplification approaches for the 35S promoter were tested: strand displacement amplification, nicking-enzyme amplification reaction, rolling circle amplification, loop-mediated isothermal amplification (LAMP) and helicase-dependent amplification (HDA). The assays developed were tested for specificity in order to distinguish between samples containing genetically modified (GM) maize and non-GM maize. For those assays capable of this discrimination, tests were performed to determine the lower limit of detection. A false-negative rate was determined to rule out whether GMO-positive samples were incorrectly classified as GMO-negative. A robustness test was performed to show reliable detection independent from the instrument used for amplification. The analysis of three GM maize lines showed that only LAMP and HDA were able to differentiate between the GMOs MON810, NK603, and Bt11 and non-GM maize. Furthermore, with the HDA assay it was possible to realize a detection limit as low as 0.5%. A false-negative rate of only 5% for 1% GM maize for all three maize lines shows that HDA has the potential to be used as an alternative strategy for the detection of transgenic maize. All results obtained with the LAMP and HDA assays were compared with the results obtained with a previously reported real-time PCR assay for the 35S promoter in transgenic maize. This study presents two new screening assays for detection of the 35S promoter in transgenic maize by applying the isothermal amplification approaches HDA and LAMP.

A lab-on-a-chip system with integrated sample preparation and loop-mediated isothermal amplification for rapid and quantitative detection of Salmonella spp. in food samples

DEFF Research Database (Denmark)

Sun, Yi; Than Linh, Quyen; Hung, Tran Quang

2015-01-01

was capable to detect Salmonella at concentration of 50 cells per test within 40 min. The simple design, together with high level of integration, isothermal amplification, and quantitative analysis of multiple samples in short time will greatly enhance the practical applicability of the LOC system for rapid...... amplification (LAMP) for rapid and quantitative detection of Salmonella spp. in food samples. The whole diagnostic procedures including DNA isolation, isothermal amplification, and real-time detection were accomplished in a single chamber. Up to eight samples could be handled simultaneously and the system...... and usually take a few hours to days to complete. In response to the demand for rapid on line or at site detection of pathogens, in this study, we describe for the first time an eight-chamber lab-on-a-chip (LOC) system with integrated magnetic beads-based sample preparation and loop-mediated isothermal...

Evaluation of four novel isothermal amplification assays towards simple and rapid genotyping of chloroquine resistant Plasmodium falciparum.

Science.gov (United States)

Chahar, Madhvi; Anvikar, Anup; Dixit, Rajnikant; Valecha, Neena

2018-07-01

Loop mediated isothermal amplification (LAMP) assay is sensitive, prompt, high throughput and field deployable technique for nucleic acid amplification under isothermal conditions. In this study, we have developed and optimized four different visualization methods of loop-mediated isothermal amplification (LAMP) assay to detect Pfcrt K76T mutants of P. falciparum and compared their important features for one-pot in-field applications. Even though all the four tested LAMP methods could successfully detect K76T mutants of P. falciparum, however considering the time, safety, sensitivity, cost and simplicity, the malachite green and HNB based methods were found more efficient. Among four different visual dyes uses to detect LAMP products accurately, hydroxynaphthol blue and malachite green could produce long stable color change and brightness in a close tube-based approach to prevent cross-contamination risk. Our results indicated that the LAMP offers an interesting novel and convenient best method for the rapid, sensitive, cost-effective, and fairly user friendly tool for detection of K76T mutants of P. falciparum and therefore presents an alternative to PCR-based assays. Based on our comparative analysis, better field based LAMP visualization method can be chosen easily for the monitoring of other important drug targets (Kelch13 propeller region). Copyright © 2018 Elsevier Inc. All rights reserved.

Development and application of loop-mediated isothermal amplification methods targeting the seM gene for detection of Streptococcus equi subsp. equi.

Science.gov (United States)

Hobo, Seiji; Niwa, Hidekazu; Oku, Kazuomi

2012-03-01

Loop-mediated isothermal amplification (LAMP) constitutes a potentially valuable diagnostic tool for rapid diagnosis of contagious diseases. In this study, we developed a novel LAMP method (seM-LAMP) to detect the seM gene of Streptococcus equi subsp. equi (S. equi), the causative agent of strangles in equids. The seM-LAMP successfully amplified the target sequence of the seM gene at 63°C within 60 min. The sensitivity of the seM-LAMP was slightly lower than the 2nd reaction of the seM semi-nested PCR. To evaluate the species specificity of the seM-LAMP, we tested 100 S. equi and 189 non-S. equi strains. Significant amplification of the DNA originating from S. equi was observed within 60 min incubation, but no amplification of non-S. equi DNA occurred. The results were identical to those of seM semi-nested PCR. To investigate the clinical usefulness of the methods, the seM-LAMP and the seM semi-nested PCR were used to screen 590 nasal swabs obtained during an outbreak of strangles. Both methods showed that 79 and 511 swabs were S. equi positive and negative, respectively, and the results were identical to those of the culture examination. These results indicate that the seM-LAMP is potentially useful for the reliable routine diagnosis of Streptococcus equi subsp. equi infections.

Detection of Bar Transgenic Sugarcane with a Rapid and Visual Loop-Mediated Isothermal Amplification Assay.

Science.gov (United States)

Zhou, Dinggang; Wang, Chunfeng; Li, Zhu; Chen, Yun; Gao, Shiwu; Guo, Jinlong; Lu, Wenying; Su, Yachun; Xu, Liping; Que, Youxiong

2016-01-01

Genetic engineering offers an attractive alternative in sugarcane breeding for increasing cane and sugar yields as well as disease and insect resistance. Bar transgenic sugarcane employing the herbicide tolerance is a useful agronomical trait in weed control. In this study, a loop-mediated isothermal amplification (LAMP) assay for rapid detection of the bar gene in transgenic sugarcane has been developed and evaluated. A set of six primers was designed for LAMP-based amplification of the bar gene. The LAMP reaction conditions were optimized as follows: 5.25 mM of Mg(2+), 6:1 ratio of inner vs. outer primer, and 6.0 U of Bst DNA polymerase in a reaction volume of 25.0 μL. The detection limit of the recombinant plasmid 1Ac0229 was as low as 10 copies in the developed LAMP, which was 10-fold higher sensitive than that of conventional PCR. In 100 putative transgenic lines, the bar gene was detected in 100/100 cases (100%) by LAMP and 97/100 cases (97%) by conventional PCR, respectively. In conclusion, the developed LAMP assay is visual, rapid, sensitive, reliable, and cost-effective for detection of the bar specific transgenic sugarcane.

Detection of bar transgenic sugarcane with a rapid and visual loop-mediated isothermal amplification assay

Directory of Open Access Journals (Sweden)

Dinggang eZhou

2016-03-01

Full Text Available Genetic engineering offers an attractive alternative in sugarcane breeding for increasing cane and sugar yields as well as disease and insect resistance. Bar transgenic sugarcane employing the herbicide tolerance is a useful agronomical trait in weed control. In this study, a loop-mediated isothermal amplification (LAMP assay for rapid detection of the bar gene in transgenic sugarcane has been developed and evaluated. A set of six primers was designed for LAMP-based amplification of the bar gene. The LAMP reaction conditions were optimized as follows: 5.25 mM of Mg2+, 6:1 ratio of inner vs outer primer, and 6.0 U of Bst DNA polymerase in a reaction volume of 25.0 μL. The detection limit of the recombinant plasmid 1Ac0229 was as low as 10 copies in the developed LAMP, which was ten-fold higher sensitive than that of conventional PCR. In 100 putative transgenic lines, the bar gene was detected in 100/100 cases (100% by LAMP and 97/100 cases (97% by conventional PCR, respectively. In conclusion, the developed LAMP assay is visual, rapid, sensitive, reliable and cost-effective for detection of the bar specific transgenic sugarcane.

Loop-mediated isothermal amplification assay for rapid and sensitive detection of sheep pox and goat pox viruses in clinical samples.

Science.gov (United States)

Venkatesan, G; Balamurugan, V; Bhanuprakash, V; Singh, R K; Pandey, A B

2016-06-01

A Loop-mediated isothermal amplification (LAMP) assay targeting the highly conserved DNA polymerase gene of capripox virus genome was developed and evaluated for rapid detection of sheep pox and goat pox viruses. The optimized LAMP assay is found specific and sensitive for amplification of target DNA with a diagnostic sensitivity and specificity of 96.6% and 100% respectively compared to quantitative PCR. The detection rate of LAMP, PCR and Q-PCR assays is found to be 81.5%, 67% and 83% respectively. This LAMP assay has the potential for rapid clinical diagnosis and surveillance of sheep pox and goat pox in field diagnostic laboratories. Copyright © 2016 Elsevier Ltd. All rights reserved.

Rapid detection of Streptococcus uberis in raw milk by loop-mediated isothermal amplification

NARCIS (Netherlands)

Cornelissen, J.B.W.J.; Greeff, De A.; Heuvelink, A.E.; Swarts, M.; Smith, H.E.; Wal, Van der F.J.

2016-01-01

A loop-mediated isothermal amplification (LAMP) method to detect Streptococcus uberis in raw milk was developed and evaluated. Three genes (sodA, pauA, cpn60) were assessed for their suitability as targets in LAMP. The analytical sensitivity was 120, 120, and 12 fg per assay for the sodA, pauA,

Loop-Mediated Isothermal Amplification untuk Mendeteksi Gen blaTEM sebagai Penyandi Extended-Spectrum Beta-Lactamase pada Isolat Enterobacteriaceae

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Bayu A. P. Wilopo

2015-12-01

Full Text Available Extended-spectrum beta-lactamase (ESBL is a beta-lactamase enzyme that is capable of hydrolyzing penicillin, cephalosporin, and monobactam, and can be inhibited by clavulanic acid. This enzyme is encoded by multiple genes, one of them is blaTEM. Polymerase chain reaction (PCR is one of the DNA amplification methods that are frequently used; however, there are other methods that can be used including, among others, loop-mediated isothermal amplification (LAMP. LAMP requires simple equipment with quicker and easy-to-read results compared to PCR. This study was a diagnostic test to explore the sensitivity and specificity of LAMP method compared to PCR in detecting blaTEM gene. Furthermore, the concordance between LAMP and PCR methods was assessed. A total of 92 Enterobacteriaceae isolates were examined by PCR and LAMP methods and compared. The result showed that the LAMP method had a sensitivity of 91.4% and a specificity of 91.2% with a concordance value (kappa of 85.4%. In conclusion, LAMP method has a good validity and a very good conformity compared to the PCR method. Therefore, LAMP method can be used as an alternative diagnostic test, especially in limited settings.

Diagnosis of brugian filariasis by loop-mediated isothermal amplification.

Directory of Open Access Journals (Sweden)

Catherine B Poole

Full Text Available In this study we developed and evaluated a Brugia Hha I repeat loop-mediated isothermal amplification (LAMP assay for the rapid detection of Brugia genomic DNA. Amplification was detected using turbidity or fluorescence as readouts. Reactions generated a turbidity threshold value or a clear visual positive within 30 minutes using purified genomic DNA equivalent to one microfilaria. Similar results were obtained using DNA isolated from blood samples containing B. malayi microfilariae. Amplification was specific to B. malayi and B. timori, as no turbidity was observed using DNA from the related filarial parasites Wuchereria bancrofti, Onchocerca volvulus or Dirofilaria immitis, or from human or mosquito. Furthermore, the assay was most robust using a new strand-displacing DNA polymerase termed Bst 2.0 compared to wild-type Bst DNA polymerase, large fragment. The results indicate that the Brugia Hha I repeat LAMP assay is rapid, sensitive and Brugia-specific with the potential to be developed further as a field tool for diagnosis and mapping of brugian filariasis.

Gold nanoparticle-based RT-PCR and real-time quantitative RT-PCR assays for detection of Japanese encephalitis virus

International Nuclear Information System (INIS)

Huang, S-H; Tsai, M-H; Lin, C-W; Yang, T-C; Chuang, P-H; Tsai, I-S; Lu, H-C; Wan Lei; Lin, Y-J; Lai, C-H

2008-01-01

Virus isolation and antibody detection are routinely used for diagnosis of Japanese encephalitis virus (JEV) infection, but the low level of transient viremia in some JE patients makes JEV isolation from clinical and surveillance samples very difficult. We describe the use of gold nanoparticle-based RT-PCR and real-time quantitative RT-PCR assays for detection of JEV from its RNA genome. We tested the effect of gold nanoparticles on four different PCR systems, including conventional PCR, reverse-transcription PCR (RT-PCR), and SYBR green real-time PCR and RT-PCR assays for diagnosis in the acute phase of JEV infection. Gold nanoparticles increased the amplification yield of the PCR product and shortened the PCR time compared to the conventional reaction. In addition, nanogold-based real-time RT-PCR showed a linear relationship between Ct and template amount using ten-fold dilutions of JEV. The nanogold-based RT-PCR and real-time quantitative RT-PCR assays were able to detect low levels (1-10 000 copies) of the JEV RNA genomes extracted from culture medium or whole blood, providing early diagnostic tools for the detection of low-level viremia in the acute-phase infection. The assays described here were simple, sensitive, and rapid approaches for detection and quantitation of JEV in tissue cultured samples as well as clinical samples

Detection of HbsAg and hATIII genetically modified goats (Caprahircus) by loop-mediated isothermal amplification.

Science.gov (United States)

Tao, Chenyu; Zhang, Qingde; Zhai, Shanli; Liu, Bang

2013-11-01

In this study, sensitive and rapid detection systems were designed using a loop-mediated isothermal amplification (LAMP) method to detect the genetically modified goats. A set of 4 primers were designed for each exogenous nucleic acids HBsAg and hATIII. The DNA samples were first amplified with the outer and inner primers and released a single-stranded DNA,of which both ends were stem-loop structure. Then one inner primer hybridized with the loop, and initiated displacement synthesis in less than 1 h. The result could be visualized by both agarose gel electrophoresis and unaided eyes directly after adding SYBR GREEN 1. The detection limit of LAMP was ten copies of target molecules, indicating that LAMP was tenfold more sensitive than the classical PCR. Furthermore, all the samples of genetically modified goats were tested positively by LAMP, and the results demonstrated that the LAMP was a rapid and sensitive method for detecting the genetically modified organism.

A Closed-tube Loop-Mediated Isothermal Amplification Assay for the Visual Endpoint Detection of Brucella spp. and Mycobacterium avium subsp. paratuberculosis.

Science.gov (United States)

Trangoni, Marcos D; Gioffré, Andrea K; Cravero, Silvio L

2017-01-01

LAMP (loop-mediated isothermal amplification) is an isothermal nucleic acid amplification technique that is characterized by its efficiency, rapidity, high yield of final product, robustness, sensitivity, and specificity, with the blueprint that it can be implemented in laboratories of low technological complexity. Despite the conceptual complexity underlying the mechanistic basis for the nucleic acid amplification, the technique is simple to use and the amplification and detection can be carried out in just one step. In this chapter, we present a protocol based on LAMP for the rapid identification of isolates of Brucella spp. and Mycobacterium avium subsp. paratuberculosis, two major bacterial pathogens in veterinary medicine.

New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination.

Science.gov (United States)

Karthik, K; Rathore, Rajesh; Thomas, Prasad; Arun, T R; Viswas, K N; Dhama, Kuldeep; Agarwal, R K

2014-01-01

Loop mediated isothermal amplification (LAMP) assay, a promising diagnostic test, has been developed for detection of different pathogens of human as well as animals. Various positive points support its use as a field level test but the major problem is product cross contamination leading to false positive results. Different methods were adopted by various researchers to control this false positive amplification due to cross contamination but all have their own advantages and disadvantages. A new closed tube LAMP assay based on agar dye capsule was developed in the present study and this technique has some advantages over the other closed tube technique.•Agar at the concentration of 1.5% was used to sandwich SYBR green dye I with the aid of intradermal syringe. This agar dye capsule was placed over the LAMP reaction mixture before it was amplified.•To eliminate the hazardous nature of Ultra Violet (UV) light during result visualization of LAMP products, the present study demonstrates the use of Light Emitting Diode (LED) lights for result visualization.•LAMP was carried out for Brucella species detection using this modified techniques yielding good results without any cross contamination and LED showed similar fluorescence compared to UV.

Lamp system with conditioned water coolant and diffuse reflector of polytetrafluorethylene(PTFE)

Science.gov (United States)

Zapata, Luis E.; Hackel, Lloyd

1999-01-01

A lamp system with a very soft high-intensity output is provided over a large area by water cooling a long-arc lamp inside a diffuse reflector of polytetrafluorethylene (PTFE) and titanium dioxide (TiO.sub.2) white pigment. The water is kept clean and pure by a one micron particulate filter and an activated charcoal/ultraviolet irradiation system that circulates and de-ionizes and biologically sterilizes the coolant water at all times, even when the long-arc lamp is off.

Non-crosslinking gold nanoprobe-LAMP for simple, colorimetric, and specific detection of Salmonella typhi

Energy Technology Data Exchange (ETDEWEB)

Bozorgmehr, Ali; Yazdanparast, Razieh, E-mail: ryazdan@ut.ac.ir [University of Tehran, Institute of Biochemistry and Biophysics (Iran, Islamic Republic of); Mollasalehi, Hamidreza [Shahid Beheshti University, Protein Research Center (Iran, Islamic Republic of)

2016-12-15

In this study, we developed a non-crosslinking gold nanoprobe loop-mediated isothermal amplification (LAMP) method for nanodiagnosis of bacterial typhoid fever source, Salmonella typhi. Therefore, a unique region in the S. typhi genomic DNA was targeted for LAMP amplification using a specific set of four precisely designed primers. Also, for specific colorimetric visualization of the amplicons, a thiolated oligonucleotide probe, complementary to the single-stranded loop region of the amplicons between F2 and F1C segments, was designed. The probe was bound to the surface of gold nanoparticles via covalent bonds. Increasing the salt concentration in the detection reaction medium led to aggregation of nanoprobes in the blank and the negative vessels in a time-dependent form. That was followed by a change in the surface plasmon resonance (SPR) leading to blue/black color that was observable by the naked eyes after about 5 min. Meanwhile, the original pink/red color was retained in the positive sample due to the large interparticle spaces and the stability against the ionic strength elevation which persisted for about 30 min. The whole process of DNA extraction, amplification, and detection took less than 2 h with a sensitivity of 20 CFU/ml. The developed gold nanoprobe-LAMP could serve as a simple, rapid, and cost-effective method for nanodiagnosis of S. typhi in point-of-need applications.

Non-crosslinking gold nanoprobe-LAMP for simple, colorimetric, and specific detection of Salmonella typhi

International Nuclear Information System (INIS)

Bozorgmehr, Ali; Yazdanparast, Razieh; Mollasalehi, Hamidreza

2016-01-01

In this study, we developed a non-crosslinking gold nanoprobe loop-mediated isothermal amplification (LAMP) method for nanodiagnosis of bacterial typhoid fever source, Salmonella typhi. Therefore, a unique region in the S. typhi genomic DNA was targeted for LAMP amplification using a specific set of four precisely designed primers. Also, for specific colorimetric visualization of the amplicons, a thiolated oligonucleotide probe, complementary to the single-stranded loop region of the amplicons between F2 and F1C segments, was designed. The probe was bound to the surface of gold nanoparticles via covalent bonds. Increasing the salt concentration in the detection reaction medium led to aggregation of nanoprobes in the blank and the negative vessels in a time-dependent form. That was followed by a change in the surface plasmon resonance (SPR) leading to blue/black color that was observable by the naked eyes after about 5 min. Meanwhile, the original pink/red color was retained in the positive sample due to the large interparticle spaces and the stability against the ionic strength elevation which persisted for about 30 min. The whole process of DNA extraction, amplification, and detection took less than 2 h with a sensitivity of 20 CFU/ml. The developed gold nanoprobe-LAMP could serve as a simple, rapid, and cost-effective method for nanodiagnosis of S. typhi in point-of-need applications.

Detection of hepatitis A virus by the nucleic acid sequence-based amplification technique and comparison with reverse transcription-PCR.

Science.gov (United States)

Jean, J; Blais, B; Darveau, A; Fliss, I

2001-12-01

A nucleic acid sequence-based amplification (NASBA) technique for the detection of hepatitis A virus (HAV) in foods was developed and compared to the traditional reverse transcription (RT)-PCR technique. Oligonucleotide primers targeting the VP1 and VP2 genes encoding the major HAV capsid proteins were used for the amplification of viral RNA in an isothermal process resulting in the accumulation of RNA amplicons. Amplicons were detected by hybridization with a digoxigenin-labeled oligonucleotide probe in a dot blot assay format. Using the NASBA, as little as 0.4 ng of target RNA/ml was detected per comparison to 4 ng/ml for RT-PCR. When crude HAV viral lysate was used, a detection limit of 2 PFU (4 x 10(2) PFU/ml) was obtained with NASBA, compared to 50 PFU (1 x 10(4) PFU/ml) obtained with RT-PCR. No interference was encountered in the amplification of HAV RNA in the presence of excess nontarget RNA or DNA. The NASBA system successfully detected HAV recovered from experimentally inoculated samples of waste water, lettuce, and blueberries. Compared to RT-PCR and other amplification techniques, the NASBA system offers several advantages in terms of sensitivity, rapidity, and simplicity. This technique should be readily adaptable for detection of other RNA viruses in both foods and clinical samples.

Rapid, highly sensitive and highly specific gene detection by combining enzymatic amplification and DNA chip detection simultaneously

Directory of Open Access Journals (Sweden)

Koji Hashimoto

2016-05-01

Full Text Available We have developed a novel gene detection method based on the loop-mediated isothermal amplification (LAMP reaction and the DNA dissociation reaction on the same DNA chip surface to achieve a lower detection limit, broader dynamic range and faster detection time than are attainable with a conventional DNA chip. Both FAM- and thiol-labeled DNA probe bound to the complementary sequence accompanying Dabcyl was immobilized on the gold surface via Au/thiol bond. The LAMP reaction was carried out on the DNA probe fixed gold surface. At first, Dabcyl molecules quenched the FAM fluorescence. According to the LAMP reaction, the complementary sequence with Dabcyl was competitively reacted with the amplified targeted sequence. As a result, the FAM fluorescence increased owing to dissociation of the complementary sequence from the DNA probe. The simultaneous reaction of LAMP and DNA chip detection was achieved, and 103 copies of the targeted gene were detected within an hour by measuring fluorescence intensity of the DNA probe. Keywords: Biosensor, DNA chip, Loop-mediated isothermal amplification (LAMP, Fluorescence detection, Gold substrate, Au/thiol bond

Simple Identification of Human Taenia Species by Multiplex Loop-Mediated Isothermal Amplification in Combination with Dot Enzyme-Linked Immunosorbent Assay

Science.gov (United States)

Nkouawa, Agathe; Sako, Yasuhito; Okamoto, Munehiro; Ito, Akira

2016-01-01

For differential detection of Taenia solium, Taenia saginata, and Taenia asiatica, loop-mediated isothermal amplification (LAMP) assay targeting the cytochrome c oxidase subunit 1 gene has been recently developed and shown to be sensitive, specific, and effective. However, to achieve differential identification, one specimen requires three reaction mixtures containing a primer set of each Taenia species separately, which is complex and time consuming and increases the risk of cross-contamination. In this study, we developed a simple differential identification of human Taenia species using multiplex LAMP (mLAMP) in combination with dot enzyme-linked immunosorbent assay (dot-ELISA). Forward inner primers of T. solium, T. saginata, and T. asiatica labeled with fluorescein isothiocyanate (FITC), digoxigenin (DIG), and tetramethylrhodamine (TAMRA), respectively, and biotin-labeled backward inner primers were used in mLAMP. The mLAMP assay succeeded in specific amplification of each respective target gene in a single tube. Furthermore, the mLAMP product from each species was easily distinguished by dot-ELISA with an antibody specific for FITC, DIG, or TAMRA. The mLAMP assay in combination with dot-ELISA will make identification of human Taenia species simpler, easier, and more practical. PMID:27044566

Time Course of Detection of Human Male DNA from Stained Blood Sample on Various Surfaces by Loop Mediated Isothermal Amplification and Polymerase Chain Reaction

Directory of Open Access Journals (Sweden)

Panan Kanchanaphum

2018-01-01

Full Text Available This study explores determining the sex of humans from blood stains taken from different surfaces and compares the time course of detection with the conventional PCR, Conventional Loop Mediated Isothermal Amplification (LAMP, and LAMP-Lateral Flow Dipstick (LFD. For the DNA templates, 7 male and 7 female blood stained samples were extracted and added to LAMP and PCR reaction solution to amplify the SRY gene. The DNA samples were extracted from the following blood stained materials: cloth, wood, clay, and tile. Then, the samples were stored at room temperature for 1, 7, 30, and 60 day(s. After the DNA amplification, the gel electrophoresis process was applied to detect LAMP product. The LFD was combined with the LAMP to detect LAMP product on the male cloth samples. For the male samples, the time course of detection on the first and seventh days indicated positive for both LAMP and PCR products on all the surfaces while no DNA amplification was found on any of the female samples. On day 30, positive LAMP product was still found on all the male samples. However, it had faded on the tiles. Moreover, all the male samples, which had tested positive for PCR product, were blurred and unclear. On day 60, LAMP product was still found on all the male samples. Conversely, the PCR method resulted in no bands showing for any of the male samples. However, the LAMP-LFD method detected product on all the male samples of cloth. The results show that the LAMP is an effective, practical, and reliable molecular-biological method. Moreover, the LFD can increase the efficiency and sensitivity of the LAMP, making it more suitable for field studies because gel electrophoresis apparatus is not required.

Development of Loop-Mediated Isothermal Amplification Assay for Detection of Entamoeba histolytica▿

Science.gov (United States)

Liang, Shih-Yu; Chan, Yun-Hsien; Hsia, Kan-Tai; Lee, Jing-Lun; Kuo, Ming-Chu; Hwa, Kuo-Yuan; Chan, Chi-Wen; Chiang, Ting-Yi; Chen, Jung-Sheng; Wu, Fang-Tzy; Ji, Dar-Der

2009-01-01

A novel one-step, closed-tube, loop-mediated isothermal amplification (LAMP) assay for detecting Entamoeba histolytica, one of the leading causes of morbidity in developing countries, was developed. The sensitivity of the LAMP assay is 1 parasite per reaction. A total of 130 clinical samples were analyzed, and the results compared with those of conventional nested PCR to validate the practicability of this assay. No DNA was amplified from other diarrheal pathogens, such as other Entamoeba species, bacteria, and viruses. These results indicate that LAMP is a rapid, simple, and valuable diagnostic tool for epidemiological studies of amebiasis. PMID:19321720

LED lamp color control system and method

Science.gov (United States)

Gaines, James; Clauberg, Bernd; Van Erp, Josephus A.M.

2013-02-05

An LED lamp color control system and method including an LED lamp having an LED controller 58; and a plurality of LED channels 60 operably connected to the LED controller 58, each of the plurality of LED channels 60 having a channel switch 62 in series with at least one shunted LED circuit 83, the shunted LED circuit 83 having a shunt switch 68 in parallel with an LED source 80. The LED controller 58 determines whether the LED source 80 is in a feedback controllable range, stores measured optical flux for the LED source 80 when the LED source 80 is in the feedback controllable range, and bypasses storing the measured optical flux when the LED source 80 is not in the feedback controllable range.

Survey and visual detection of Zaire ebolavirus in clinical samples targeting the nucleoprotein gene in Sierra Leone

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Jing Yuan

2015-12-01

Full Text Available Ebola virus (EBOV can lead to severe hemorrhagic fever with a high risk of death in humans and other primates. To guide treatment and prevent spread of the viral infection, a rapid and sensitive detection method is required for clinical samples. Here, we described and evaluated a reverse transcription loop-mediated isothermal amplification (RT-LAMP method to detect Zaire ebolavirus using the nucleoprotein gene (NP as a target sequence. Two different techniques were used, a calcein/Mn2+ complex chromogenic method and real-time turbidity monitoring. The RT-LAMP assay detected the NP target sequence with a limit of 4.56 copies/μL within 45 min under 61°C, a similar even or increase in sensitivity than that of real-time reverse transcription-polymerase chain reaction (RT-PCR. Additionally, all pseudoviral particles or non- Zaire EBOV genomes were negative for LAMP detection, indicating that the assay was highly specific for EBOV. To appraise the availability of the RT-LAMP method for use in clinical diagnosis of EBOV, of 417 blood or swab samples collected from patients with clinically suspected infections in Sierra Leone, 307 were identified for RT-LAMP-based surveillance of EBOV. Therefore, the highly specific and sensitive RT-LAMP method allows the rapid detection of EBOV, and is a suitable tool for clinical screening, diagnosis, and primary quarantine purposes.

Quantitative real-time RT-PCR and chromogenic in situ hybridization

DEFF Research Database (Denmark)

Rosa, Fabíola E; Silveira, Sara M; Silveira, Cássia G T

2009-01-01

. METHODS: To elucidate the molecular profile of HER-2 status, mRNA and protein expression in 75 invasive breast carcinomas were analyzed by real time quantitative RT-PCR (qRT-PCR) and IHC, respectively. Amplifications were evaluated in 43 of these cases by CISH and in 11 by FISH. RESULTS: The concordance...

Microfluidic method for rapid turbidimetric detection of the DNA of Mycobacterium tuberculosis using loop-mediated isothermal amplification in capillary tubes

International Nuclear Information System (INIS)

Rafati, Adele; Gill, Pooria

2015-01-01

We describe a microfluidic method for rapid isothermal turbidimetric detection of the DNA of Mycobacterium tuberculosis. Loop-mediated isothermal amplification is accomplished in capillary tubes for amplifying DNA in less than 15 min, and sensitivity and specificity were compared to conventional loop-mediated isothermal amplification (LAMP). The method can detect as little as 1 pg mL −1 DNA in a sample. Results obtained with clinical specimens indicated 90 % sensitivity and 95 % specificity for microfluidic LAMP in comparison to culture methods. No interference occurred due to the presence of nonspecific DNAs. The findings demonstrate the power of the new microfluidic LAMP test for rapid molecular detection of microorganisms even when using bare eyes. (author)

Generation of recombinant pestiviruses using a full genome amplification strategy

DEFF Research Database (Denmark)

Rasmussen, Thomas Bruun; Reimann, Ilona; Uttenthal, Åse

Aim Complete genome amplification of viral RNA provides a new tool for generation of modified pestiviruses. We have recently reported a full genome amplification strategy for direct recovery of infectious pestivirus (Rasmussen et al., 2008). This comprised rescue of BDV strain “Gifhorn” from a full......-length RT-PCR amplicon demonstrating that long RT-PCR can be used for direct generation of an infectious pestivirus. The strategy is not limited to amplification of BDV “Gifhorn”, but can be further utilized for amplification of a diverse selection of pestivirus strains and for the generation of modified...... was reverse transcribed to cDNA at 50C for 90 minutes using SuperScript III reverse transcriptase (Invitrogen). Full-length PCR amplification was performed using primers specific for the extreme 5’- and 3’-ends of the viral genomes. A T7 promoter was incorporated in the 5’-primers for direct in vitro...

Detection of Nosema bombycis by FTA cards and loop-mediated isothermal amplification (LAMP).

Science.gov (United States)

Yan, Wei; Shen, Zhongyuan; Tang, Xudong; Xu, Li; Li, Qianlong; Yue, Yajie; Xiao, Shengyan; Fu, Xuliang

2014-10-01

We successfully established a detection method which exhibited a markedly higher sensitivity than previously developed detection methods for Nosema bombycis by combining glass beads, FTA card, and LAMP. Spores of N. bombycis were first broken by acid-washed glass beads; the DNA was subsequently extracted and purified with the FTA card, and LAMP was performed using primers (LSU296) designed based on the sequence of the LSU rRNA of N. bombycis. The minimum detection concentration was 10 spores/mL. When this method was used to detect pebrine disease in silkworm egg, the detection rate for 500 silkworm eggs, in which only one egg was infected with N. bombycis, was 100 % under our optimized conditions. If the number of eggs in the sample increased to 800 or 1,000, the sample was divided into two equal portions, and the eggs were smashed with glass beads after the addition of 1 mL of TE buffer. The liquid in two tubes was later mixed and applied to the FTA card, and the detection rates were 100 %. Furthermore, the LAMP method established in our study could detect N. bombycis infection in silkworm 24 h earlier than microscopy.

Evaluation of Nucleic Acid Isothermal Amplification Methods for Human Clinical Microbial Infection Detection

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Brett E. Etchebarne

2017-12-01

Full Text Available Battling infection is a major healthcare objective. Untreated infections can rapidly evolve toward the condition of sepsis in which the body begins to fail and resuscitation becomes critical and tenuous. Identification of infection followed by rapid antimicrobial treatment are primary goals of medical care, but precise identification of offending organisms by current methods is slow and broad spectrum empirical therapy is employed to cover most potential pathogens. Current methods for identification of bacterial pathogens in a clinical setting typically require days of time, or a 4- to 8-h growth phase followed by DNA extraction, purification and PCR-based amplification. We demonstrate rapid (70–120 min genetic diagnostics methods utilizing loop-mediated isothermal amplification (LAMP to test for 15 common infection pathogen targets, called the Infection Diagnosis Panel (In-Dx. The method utilizes filtration to rapidly concentrate bacteria in sample matrices with lower bacterial loads and direct LAMP amplification without DNA purification from clinical blood, urine, wound, sputum and stool samples. The In-Dx panel was tested using two methods of detection: (1 real-time thermocycler fluorescent detection of LAMP amplification and (2 visual discrimination of color change in the presence of Eriochrome Black T (EBT dye following amplification. In total, 239 duplicate samples were collected (31 blood, 122 urine, 73 mucocutaneous wound/swab, 11 sputum and two stool from 229 prospectively enrolled hospital patients with suspected clinical infection and analyzed both at the hospital and by In-Dx. Sensitivity (Se of the In-Dx panel targets pathogens from urine samples by In-Dx was 91.1% and specificity (Sp was 97.3%, with a positive predictive value (PPV of 53.7% and a negative predictive value (NPV of 99.7% as compared to clinical microbial detection methods. Sensitivity of detection of the In-Dx panel from mucocutaneous swab samples was 65.5% with a

Materials for incandescent and fluorescent lamps

DEFF Research Database (Denmark)

Thorsen, Knud Aage

1996-01-01

The article gives an overview of the materials systems used for incandescent lamps as well as a brief introduction to the systems used for fluorescent lamps. The materials used for incandescent lamps are doped tungsten used for the filaments, metals and alloys used for terminal and support posts......, lead wires and internal reflectors and screens as well as glasses for the envelope. The physics of bulbs and changes in bulbs during use are elucidated. The cost and energy savings and environmental benefits by replacement of incandescent lamps by fluorescent lamps are presented....

Immunocapture loop-mediated isothermal amplification assays for the detection of canine parvovirus.

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Sun, Yu-Ling; Yen, Chon-Ho; Tu, Ching-Fu

2017-11-01

A loop-mediated isothermal amplification (LAMP) assay was used for rapid canine parvovirus (CPV) diagnosis. To reduce the time required and increase the sensitivity of the assay, an immunocapture (IC) technique was developed in this study to exclude the DNA extraction step in molecular diagnostic procedures for CPV. A polyclonal rabbit anti-CPV serum was produced against VP2-EpC that was cloned via DNA recombination. The polyclonal anti-VP2-EpC serum was used for virus capture to prepare microtubes. IC-LAMP was performed to amplify a specific CPV target gene sequence from the CPV viral particles that were captured on the microtubes, and the amplicons were analyzed using agarose electrophoresis or enzyme-linked immunosorbent assay (IC-LAMP-ELISA) and lateral-flow dipstick (IC-LAMP-LFD). The detection sensitivities of IC-LAMP, IC-LAMP-ELISA, and IC-LAMP-LFD were 10 -1 , 10 -1 , and 10 -1 TCID 50 /mL, respectively. Using the IC-LAMP-ELISA and IC-LAMP-LFD assays, the complete CPV diagnostic process can be achieved within 1.5h. Both of the developed IC-LAMP-based assays are simple, direct visual and efficient techniques that are applicable to the detection of CPV. Copyright © 2017 Elsevier B.V. All rights reserved.

Simple Identification of Human Taenia Species by Multiplex Loop-Mediated Isothermal Amplification in Combination with Dot Enzyme-Linked Immunosorbent Assay.

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Nkouawa, Agathe; Sako, Yasuhito; Okamoto, Munehiro; Ito, Akira

2016-06-01

For differential detection of Taenia solium, Taenia saginata, and Taenia asiatica, loop-mediated isothermal amplification (LAMP) assay targeting the cytochrome c oxidase subunit 1 gene has been recently developed and shown to be sensitive, specific, and effective. However, to achieve differential identification, one specimen requires three reaction mixtures containing a primer set of each Taenia species separately, which is complex and time consuming and increases the risk of cross-contamination. In this study, we developed a simple differential identification of human Taenia species using multiplex LAMP (mLAMP) in combination with dot enzyme-linked immunosorbent assay (dot-ELISA). Forward inner primers of T. solium, T. saginata, and T. asiatica labeled with fluorescein isothiocyanate (FITC), digoxigenin (DIG), and tetramethylrhodamine (TAMRA), respectively, and biotin-labeled backward inner primers were used in mLAMP. The mLAMP assay succeeded in specific amplification of each respective target gene in a single tube. Furthermore, the mLAMP product from each species was easily distinguished by dot-ELISA with an antibody specific for FITC, DIG, or TAMRA. The mLAMP assay in combination with dot-ELISA will make identification of human Taenia species simpler, easier, and more practical. © The American Society of Tropical Medicine and Hygiene.

Loop-Mediated Isothermal Amplification for Detection of Endogenous Sad1 Gene in Cotton: An Internal Control for Rapid Onsite GMO Testing.

Science.gov (United States)

Singh, Monika; Bhoge, Rajesh K; Randhawa, Gurinderjit

2018-04-20

Background : Confirming the integrity of seed samples in powdered form is important priorto conducting a genetically modified organism (GMO) test. Rapid onsite methods may provide a technological solution to check for genetically modified (GM) events at ports of entry. In India, Bt cotton is the commercialized GM crop with four approved GM events; however, 59 GM events have been approved globally. GMO screening is required to test for authorized GM events. The identity and amplifiability of test samples could be ensured first by employing endogenous genes as an internal control. Objective : A rapid onsite detection method was developed for an endogenous reference gene, stearoyl acyl carrier protein desaturase ( Sad1 ) of cotton, employing visual and real-time loop-mediated isothermal amplification (LAMP). Methods : The assays were performed at a constant temperature of 63°C for 30 min for visual LAMP and 62ºC for 40 min for real-time LAMP. Positive amplification was visualized as a change in color from orange to green on addition of SYBR ® Green or detected as real-time amplification curves. Results : Specificity of LAMP assays was confirmed using a set of 10 samples. LOD for visual LAMP was up to 0.1%, detecting 40 target copies, and for real-time LAMP up to 0.05%, detecting 20 target copies. Conclusions : The developed methods could be utilized to confirm the integrity of seed powder prior to conducting a GMO test for specific GM events of cotton. Highlights : LAMP assays for the endogenous Sad1 gene of cotton have been developed to be used as an internal control for onsite GMO testing in cotton.

Development of a loop-mediated isothermal amplification assay for detection of Trichomonas vaginalis.

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Reyes, John Carlo B; Solon, Juan Antonio A; Rivera, Windell L

2014-07-01

A loop-mediated isothermal amplification (LAMP) assay targeting the 2-kbp repeated DNA species-specific sequence was developed for detection of Trichomonas vaginalis, the causative agent of trichomoniasis. The analytical sensitivity and specificity of the LAMP assay were evaluated using pooled genital swab and urine specimens, respectively, spiked with T. vaginalis trophozoites. Genital secretion and urine did not inhibit the detection of the parasite. The sensitivity of the LAMP was 10-1000 times higher than the PCR performed. The detection limit of LAMP was 1 trichomonad for both spiked genital swab and urine specimens. Also, LAMP did not exhibit cross-reactivity with closely-related trichomonads, Trichomonas tenax and Pentatrichomonas hominis, and other enteric and urogenital microorganisms, Entamoeba histolytica, Candida albicans, Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus. This is the first report of a LAMP assay for the detection of T. vaginalis and has prospective application for rapid diagnosis and control of trichomoniasis. Copyright © 2014 Elsevier Inc. All rights reserved.

Development of Temperature Control Solutions for Non-Instrumented Nucleic Acid Amplification Tests (NINAAT

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Tamás Pardy

2017-06-01

Full Text Available Non-instrumented nucleic acid amplification tests (NINAAT are a novel paradigm in portable molecular diagnostics. They offer the high detection accuracy characteristic of nucleic acid amplification tests (NAAT in a self-contained device, without the need for any external instrumentation. These Point-of-Care tests typically employ a Lab-on-a-Chip for liquid handling functionality, and perform isothermal nucleic acid amplification protocols that require low power but high accuracy temperature control in a single well-defined temperature range. We propose temperature control solutions based on commercially available heating elements capable of meeting these challenges, as well as demonstrate the process by which such elements can be fitted to a NINAAT system. Self-regulated and thermostat-controlled resistive heating elements were evaluated through experimental characterization as well as thermal analysis using the finite element method (FEM. We demonstrate that the proposed solutions can support various NAAT protocols, as well as demonstrate an optimal solution for the loop-mediated isothermal amplification (LAMP protocol. Furthermore, we present an Arduino-compatible open-source thermostat developed for NINAAT applications.

Detection of natural infection of infectious spleen and kidney necrosis virus in farmed tilapia by hydroxynapthol blue-loop-mediated isothermal amplification assay.

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Suebsing, R; Pradeep, P J; Jitrakorn, S; Sirithammajak, S; Kampeera, J; Turner, W A; Saksmerprome, V; Withyachumnarnkul, B; Kiatpathomchai, W

2016-07-01

Infectious spleen and kidney necrosis virus (ISKNV) has recently been recognized as a causative agent of serious systemic disease in tilapia. Our objective was to establish a new colorimetric loop-mediated isothermal amplification (LAMP) assay with pre-addition of hydroxynapthol blue (blue-LAMP) to investigate ISKNV transmission in tilapia. The blue-LAMP, targeting a major capsid protein gene of ISKNV, was conducted at 65°C for 45 min, allowing unaided visual detection of the pathogen based on colour change without cross-amplification of other known fish pathogens tested. Comparison of blue-LAMP and PCR assays revealed a higher detection level for blue-LAMP assay (41·33%) in a population of farmed tilapia infected with ISKNV. The investigation of ISKNV transmission pattern in farmed red tilapia using the blue-LAMP revealed a possible matroclinical form. The presence of ISKNV in the gonad samples was confirmed by in situ LAMP assay. Positive signals only appeared in ovarian follicles, and not in oocytes. Moreover, tissue tropism assay revealed that the brain was the main target organ in both farmed red tilapia (40%) and Nile tilapia (20%). The developed blue-LAMP assay has the potential to be used as a viable tool for screening covert and natural infections of ISKNV in tilapia. The evidence of vertical transmission of ISKNV infection in tilapia indicates the seriousness of this disease and will require a close attention and collaboration between tilapia hatcheries and disease experts in order to find a solution. The new blue-LAMP assay is a time-saving and economically viable detection tool, which allows unaided visual detection for ISKNV in tilapia, and it could be applicable for field applications. Evidence on the vertical transmission of ISKNV in farmed tilapia suggests a need for developing farm management practices to control the spread of virus in aquaculture industries. © 2016 The Society for Applied Microbiology.

Exponential isothermal amplification of nucleic acids and amplified assays for proteins, cells, and enzyme activities.

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Reid, Michael S; Le, X Chris; Zhang, Hongquan

2018-04-27

Isothermal exponential amplification techniques, such as strand-displacement amplification (SDA), rolling circle amplification (RCA), loop-mediated isothermal amplification (LAMP), nucleic acid sequence-based amplification (NASBA), helicase-dependent amplification (HDA), and recombinase polymerase amplification (RPA), have great potential for on-site, point-of-care, and in-situ assay applications. These amplification techniques eliminate the need for temperature cycling required for polymerase chain reaction (PCR) while achieving comparable amplification yield. We highlight here recent advances in exponential amplification reaction (EXPAR) for the detection of nucleic acids, proteins, enzyme activities, cells, and metal ions. We discuss design strategies, enzyme reactions, detection techniques, and key features. Incorporation of fluorescence, colorimetric, chemiluminescence, Raman, and electrochemical approaches enables highly sensitive detection of a variety of targets. Remaining issues, such as undesirable background amplification resulting from non-specific template interactions, must be addressed to further improve isothermal and exponential amplification techniques. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Loop-mediated isothermal amplification for detection of Staphylococcus aureus in dairy cow suffering from mastitis.

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Tie, Zhang; Chunguang, Wang; Xiaoyuan, Wei; Xinghua, Zhao; Xiuhui, Zhong

2012-01-01

To develop a rapid detection method of Staphylococcus aureus using loop-mediated isothermal amplification (LAMP), four specific primers were designed according to six distinct sequences of the nuc gene. 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 was 1 × 10² CFU/mL and that of PCR was 1 × 10⁴ CFU/mL, indicating that the LAMP was 100 times more sensitive than the PCR. The LAMP method for detection of Staphylococcus aureus has 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 of Staphylococcus aureus.

Development of a loop-mediated isothermal amplification method for rapid mass-screening of sand flies for Leishmania infection.

Science.gov (United States)

Nzelu, Chukwunonso O; Gomez, Eduardo A; Cáceres, Abraham G; Sakurai, Tatsuya; Martini-Robles, Luiggi; Uezato, Hiroshi; Mimori, Tatsuyuki; Katakura, Ken; Hashiguchi, Yoshihisa; Kato, Hirotomo

2014-04-01

Entomological monitoring of Leishmania infection in leishmaniasis endemic areas offers epidemiologic advantages for predicting the risk and expansion of the disease, as well as evaluation of the effectiveness of control programs. In this study, we developed a highly sensitive loop-mediated isothermal amplification (LAMP) method for the mass screening of sand flies for Leishmania infection based on the 18S rRNA gene. The LAMP technique could detect 0.01 parasites, which was more sensitive than classical PCR. The method was robust and could amplify the target DNA within 1h from a crude sand fly template without DNA purification. Amplicon detection could be accomplished by the newly developed colorimetric malachite green (MG)--mediated naked eye visualization. Pre-addition of MG to the LAMP reaction solution did not inhibit amplification efficiency. The field applicability of the colorimetric MG-based LAMP assay was demonstrated with 397 field-caught samples from the endemic areas of Ecuador and eight positive sand flies were detected. The robustness, superior sensitivity, and ability to produce better visual discriminatory reaction products than existing LAMP fluorescence and turbidity assays indicated the field potential usefulness of this new method for surveillance and epidemiological studies of leishmaniasis in developing countries. Copyright © 2013 Elsevier B.V. All rights reserved.

Development and evaluation of a novel and rapid detection assay for Botrytis cinerea based on loop-mediated isothermal amplification.

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Ya-Bing Duan

Full Text Available Botrytis cinerea is a devastating plant pathogen that causes grey mould disease. In this study, we developed a visual detection method of B. cinerea based on the Bcos5 sequence using loop-mediated isothermal amplification (LAMP with hydroxynaphthol blue dye (HNB. The LAMP reaction was optimal at 63 °C for 45 min. When HNB was added prior to amplification, samples with B. cinerea DNA developed a characteristic sky blue color after the reaction but those without DNA or with DNA of other plant pathogenic fungi did not. Results of HNB staining method were reconfirmed when LAMP products were subjected to gel electrophoresis. The detection limit of this LAMP assay for B. cinerea was 10(-3 ng µL(-1 of genomic DNA per reaction, which was 10-fold more sensitive than conventional PCR (10(-2 ng µL(-1. Detection of the LAMP assay for inoculum of B. cinerea was possible in the inoculated tomato and strawberry petals. In the 191 diseased samples, 180 (94.2% were confirmed as positive by LAMP, 172 (90.1% positive by the tissue separation, while 147 (77.0% positive by PCR. Because the LAMP assay performed well in aspects of sensitivity, specificity, repeatability, reliability, and visibility, it is suitable for rapid detection of B. cinerea in infected plant materials prior to storage and during transportation, such as cut flowers, fruits and vegetables.

One simple DNA extraction device and its combination with modified visual loop-mediated isothermal amplification for rapid on-field detection of genetically modified organisms.

Science.gov (United States)

Zhang, Miao; Liu, Yinan; Chen, Lili; Quan, Sheng; Jiang, Shimeng; Zhang, Dabing; Yang, Litao

2013-01-02

Quickness, simplicity, and effectiveness are the three major criteria for establishing a good molecular diagnosis method in many fields. Herein we report a novel detection system for genetically modified organisms (GMOs), which can be utilized to perform both on-field quick screening and routine laboratory diagnosis. In this system, a newly designed inexpensive DNA extraction device was used in combination with a modified visual loop-mediated isothermal amplification (vLAMP) assay. The main parts of the DNA extraction device included a silica gel membrane filtration column and a modified syringe. The DNA extraction device could be easily operated without using other laboratory instruments, making it applicable to an on-field GMO test. High-quality genomic DNA (gDNA) suitable for polymerase chain reaction (PCR) and isothermal amplification could be quickly isolated from plant tissues using this device within 15 min. In the modified vLAMP assay, a microcrystalline wax encapsulated detection bead containing SYBR green fluorescent dye was introduced to avoid dye inhibition and cross-contaminations from post-LAMP operation. The system was successfully applied and validated in screening and identification of GM rice, soybean, and maize samples collected from both field testing and the Grain Inspection, Packers, and Stockyards Administration (GIPSA) proficiency test program, which demonstrated that it was well-adapted to both on-field testing and/or routine laboratory analysis of GMOs.

Loop-Mediated Isothermal Amplification as a Fast Noninvasive Method of Helicobacter pylori Diagnosis.

Science.gov (United States)

Yari, Farideh; Abiri, Ramin; Aryan, Ehsan; Ahmadi Jouybari, Touraj; Navabi, Jafar; Alvandi, Amirhooshang

2016-09-01

Helicobacter pylori infection is etiologically associated with some important health problems such as gastric cancer. Because of the high clinical importance of H. pylori infection, development of a noninvasive test for the detection of H. pylori is desirable. In this study, a loop-mediated isothermal amplification (LAMP) targeted ureC of H. pylori was evaluated on 100 stool specimens and compared with a stool antigen test. Culture and rapid urease test were considered as gold standards. The overall detection rate of the fecal antigen test and LAMP was 58% and 82%, respectively. The analytical sensitivity of the fecal antigen test and LAMP was 500 and 10 H. pylori cells/g and 10 fg DNA/reaction, which is equal to six H. pylori genome. LAMP technique has been characterized by high sensitivity and low detection limit for the detection of H. pylori in stool specimen. Clinical diagnostic performance of LAMP was better than the stool antigen test. © 2015 Wiley Periodicals, Inc.

Intelligent Automatic Right-Left Sign Lamp Based on Brain Signal Recognition System

Science.gov (United States)

Winda, A.; Sofyan; Sthevany; Vincent, R. S.

2017-12-01

Comfort as a part of the human factor, plays important roles in nowadays advanced automotive technology. Many of the current technologies go in the direction of automotive driver assistance features. However, many of the driver assistance features still require physical movement by human to enable the features. In this work, the proposed method is used in order to make certain feature to be functioning without any physical movement, instead human just need to think about it in their mind. In this work, brain signal is recorded and processed in order to be used as input to the recognition system. Right-Left sign lamp based on the brain signal recognition system can potentially replace the button or switch of the specific device in order to make the lamp work. The system then will decide whether the signal is ‘Right’ or ‘Left’. The decision of the Right-Left side of brain signal recognition will be sent to a processing board in order to activate the automotive relay, which will be used to activate the sign lamp. Furthermore, the intelligent system approach is used to develop authorized model based on the brain signal. Particularly Support Vector Machines (SVMs)-based classification system is used in the proposed system to recognize the Left-Right of the brain signal. Experimental results confirm the effectiveness of the proposed intelligent Automatic brain signal-based Right-Left sign lamp access control system. The signal is processed by Linear Prediction Coefficient (LPC) and Support Vector Machines (SVMs), and the resulting experiment shows the training and testing accuracy of 100% and 80%, respectively.

Development of Loop-Mediated Isothermal Amplification for Detection of Leifsonia xyli subsp. xyli in Sugarcane

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Jing Liu

2013-01-01

Full Text Available Ratoon stunt, caused by the xylem-limited coryneform bacterium Leifsonia xyli subsp. xyli (Lxx, is a deep bacteriosis and prevalent in most of sugarcane-producing countries. Based on loop-mediated isothermal amplification (LAMP, we developed a method for detecting Lxx. The major advantages of the LAMP method are visual judgment by color and time saving with only 60 min for identification of Lxx and without the need for costly PCR apparatus and gel scanner. In the present study, positive and negative samples detected by the LAMP method were clearly distinguishable. When total DNA extracted from internode juice was used as the template, the sensitivity of LAMP was 10 times higher than that of the conventional PCR detection. The LAMP assay is a highly specific, rapid, and sensitive method for the diagnosis of ratoon stunt caused by Lxx in sugarcane. This is the first report of LAMP-based assay for the detection of Lxx in sugarcane.

Harmonics Monitoring Survey on LED Lamps

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Abdelrahman Ahmed Akila

2017-03-01

Full Text Available Light Emitting Diode (LED lamps are being increasingly used in many applications. These LED lamps operate using a driver, which is a switching device. Hence, LED lamps will be a source of harmonics in the power system. These harmonics if not well treated, may cause severe performance and operational problems. In this paper, harmonics (amplitude and phase angles generated by both LED lamps and conventional fluorescent lamps will be studied practically. Then they will be analyzed and evaluated. Compared to each other harmonics generated by both LED and conventional florescent lamps, self mitigation may occur based on the phase angle of these harmonics. All data will be measured using power analyzer and will be done on a sample of actual lamps.

New closed tube loop mediated isothermal amplification assay for prevention of product cross-contamination

OpenAIRE

Karthik, K.; Rathore, Rajesh; Thomas, Prasad; Arun, T.R.; Viswas, K.N.; Dhama, Kuldeep; Agarwal, R.K.

2014-01-01

Loop mediated isothermal amplification (LAMP) assay, a promising diagnostic test, has been developed for detection of different pathogens of human as well as animals. Various positive points support its use as a field level test but the major problem is product cross contamination leading to false positive results. Different methods were adopted by various researchers to control this false positive amplification due to cross contamination but all have their own advantages and disadvantages. A...

Rapid and sensitive detection of Didymella bryoniae by visual loop-mediated isothermal amplification assay

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Xiefeng Yao

2016-08-01

Full Text Available Didymella bryoniae is a pathogenic fungus that causes gummy stem blight (GSB in Cucurbitaceae crops (e.g. cantaloupe, muskmelon, cucumber, and watermelon. GSB produces lesions on the stems and leaves, and can also be spread by seeds. Here, we developed a rapid, visual, and sensitive loop-mediated amplification (LAMP assay for D. bryoniae detection based on sequence-characterized amplified regions (GenBank accession nos GQ872461 and GQ872462 common to the two random amplification of polymorphic DNA group genotypes (RGI and RGII of D. bryoniae; ideal conditions for detection were optimized for completion in 45 min at 63°C. The sensitivity and specificity of the LAMP assay were further analyzed in comparison with those of a conventional polymerase chain reaction (PCR. The sensitivity of the LAMP assay was 1000-fold higher than that of conventional PCR with a detection limit of 0.1 fg μL−1 of targeted DNA. The LAMP assay could be accomplished in about 45 min, with the results visible to the naked eye. The assay showed high specificity in discriminating all D. bryoniae isolates from seven other fungal pathogens that occur in Cucurbitaceae crops. The LAMP assay also detected D. bryoniae infection in young muskmelon leaves with suspected early symptoms of GSB disease. Hence, the technique has great potential for developing rapid and sensitive visual detection methods for the D. bryoniae pathogen in crops and seeds. This method has potential application in early prediction of disease and reducing the risk of epidemics.

Lamp for sunshine simulation

DEFF Research Database (Denmark)

2016-01-01

A lamp system is provided, comprising a lamp with a lamp housing accommodating a plurality of light sources for emission of visible light, including blue light, a time keeping unit, a light sensor for sensing intensity of light incident upon it, and a light controller configured for controlling...... the plurality of light sources in response to the intensity of light sensed by the light sensor and the time provided by the time keeping unit, characterized in that the lamp emits blue light for a selected time period, wherein the blue light has a luminous flux ranging from 50 lux to 200 lux and, preferably......, an irradiance that is larger than 5 mW/nm/m2 in a selected wavelength range, such as in the wavelength range from 440 nm to 500 nm, as measured at a distance of 3 metres from the lamp....

Field Evaluation of a High Throughput Loop Mediated Isothermal Amplification Test for the Detection of Asymptomatic Plasmodium Infections in Zanzibar.

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Berit Aydin-Schmidt

Full Text Available New field applicable diagnostic tools are needed for highly sensitive detection of residual malaria infections in pre-elimination settings. Field performance of a high throughput DNA extraction system for loop mediated isothermal amplification (HTP-LAMP was therefore evaluated for detecting malaria parasites among asymptomatic individuals in Zanzibar.HTP-LAMP performance was evaluated against real-time PCR on 3008 paired blood samples collected on filter papers in a community-based survey in 2015.The PCR and HTP-LAMP determined malaria prevalences were 1.6% (95%CI 1.3-2.4 and 0.7% (95%CI 0.4-1.1, respectively. The sensitivity of HTP-LAMP compared to PCR was 40.8% (CI95% 27.0-55.8 and the specificity was 99.9% (CI95% 99.8-100. For the PCR positive samples, there was no statistically significant difference between the geometric mean parasite densities among the HTP-LAMP positive (2.5 p/μL, range 0.2-770 and HTP-LAMP negative (1.4 p/μL, range 0.1-7 samples (p = 0.088. Two lab technicians analysed up to 282 samples per day and the HTP-LAMP method was experienced as user friendly.Although field applicable, this high throughput format of LAMP as used here was not sensitive enough to be recommended for detection of asymptomatic low-density infections in areas like Zanzibar, approaching malaria elimination.

Detection of Puccinia kuehnii Causing Sugarcane Orange Rust with a Loop-Mediated Isothermal Amplification-Based Assay.

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Chandra, Amaresh; Keizerweerd, Amber T; Grisham, Michael P

2016-03-01

Puccinia kuehnii is a fungal pathogen that causes orange rust in sugarcane, which is now prevalent in many countries. At the early stage of disease, it is almost indistinguishable from brown rust, which is caused by Puccinia melanocephala. Although several PCR assays are available to detect these diseases, the loop-mediated isothermal amplification (LAMP)-based assay has been reported to be more economical and easier to perform. Under isothermal conditions, DNA is amplified with high specificity and rapidity. Moreover, visual judgment of color change without further post-amplification processing makes the method convenient. The present study was undertaken to detect P. kuehnii genomic DNA using four primers corresponding to a unique DNA sequence of P. kuehnii. The LAMP assay was found to be optimal when 8 mM MgSO4 was used and the reaction was incubated at 63 °C for 90 min. Positive samples showed a color change from orange to green upon SYBR Green I dye addition. Specificity of the LAMP test was checked with DNA of P. melanocephala, which showed no reaction. Sensitivity of the LAMP method was observed to be the same as real-time PCR at 0.1 ng, thus providing a rapid and more affordable option for early disease detection.

Application of Wireless Intelligent Control System for HPS Lamps and LEDs Combined Illumination in Road Tunnel

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Lai, Jinxing; Qiu, Junling; Chen, Jianxun; Wang, Yaqiong; Fan, Haobo

2014-01-01

Because of the particularity of the environment in the tunnel, the rational tunnel illumination system should be developed, so as to optimize the tunnel environment. Considering the high cost of traditional tunnel illumination system with high-pressure sodium (HPS) lamps as well as the effect of a single light source on tunnel entrance, the energy-saving illumination system with HPS lamps and LEDs combined illumination in road tunnel, which could make full use of these two kinds of lamps, was proposed. The wireless intelligent control system based on HPS lamps and LEDs combined illumination and microcontrol unit (MCU) Si1000 wireless communication technology was designed. And the remote monitoring, wireless communication, and PWM dimming module of this system were designed emphatically. Intensity detector and vehicle flow detector can be configured in wireless intelligent control system, which gather the information to the master control unit, and then the information is sent to the monitoring center through the Ethernet. The control strategies are got by the monitoring center according to the calculated results, and the control unit wirelessly sends parameters to lamps, which adjust the luminance of each segment of the tunnel and realize the wireless intelligent control of combined illumination in road tunnel. PMID:25587266

Direct LAMP Assay without Prior DNA Purification for Sex Determination of Papaya

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Chi-Chu Tsai

2016-09-01

Full Text Available Papaya (Carica papaya L. is an economically important tropical fruit tree with hermaphrodite, male and female sex types. Hermaphroditic plants are the major type used for papaya production because their fruits have more commercial advantages than those of female plants. Sex determination of the seedlings, or during the early growth stages, is very important for the papaya seedling industry. Thus far, the only method for determining the sex type of a papaya at the seedling stage has been DNA analysis. In this study, a molecular technique—based on DNA analysis—was developed for detecting male-hermaphrodite-specific markers to examine the papaya’s sex type. This method is based on the loop-mediated isothermal amplification (LAMP and does not require prior DNA purification. The results show that the method is an easy, efficient, and inexpensive way to determine a papaya’s sex. This is the first report on the LAMP assay, using intact plant materials-without DNA purification-as samples for the analysis of sex determination of papaya. We found that using high-efficiency DNA polymerase was essential for successful DNA amplification, using trace intact plant material as a template DNA source.

Optofluidic analysis system for amplification-free, direct detection of Ebola infection

Science.gov (United States)

Cai, H.; Parks, J. W.; Wall, T. A.; Stott, M. A.; Stambaugh, A.; Alfson, K.; Griffiths, A.; Mathies, R. A.; Carrion, R.; Patterson, J. L.; Hawkins, A. R.; Schmidt, H.

2015-09-01

The massive outbreak of highly lethal Ebola hemorrhagic fever in West Africa illustrates the urgent need for diagnostic instruments that can identify and quantify infections rapidly, accurately, and with low complexity. Here, we report on-chip sample preparation, amplification-free detection and quantification of Ebola virus on clinical samples using hybrid optofluidic integration. Sample preparation and target preconcentration are implemented on a PDMS-based microfluidic chip (automaton), followed by single nucleic acid fluorescence detection in liquid-core optical waveguides on a silicon chip in under ten minutes. We demonstrate excellent specificity, a limit of detection of 0.2 pfu/mL and a dynamic range of thirteen orders of magnitude, far outperforming other amplification-free methods. This chip-scale approach and reduced complexity compared to gold standard RT-PCR methods is ideal for portable instruments that can provide immediate diagnosis and continued monitoring of infectious diseases at the point-of-care.

Loop-mediated isothermal amplification as an emerging technology for detection of Yersinia ruckeri the causative agent of enteric red mouth disease in fish

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Soliman Hatem

2008-08-01

Full Text Available Abstract Background Enteric Redmouth (ERM disease also known as Yersiniosis is a contagious disease affecting salmonids, mainly rainbow trout. The causative agent is the gram-negative bacterium Yersinia ruckeri. The disease can be diagnosed by isolation and identification of the causative agent, or detection of the Pathogen using fluorescent antibody tests, ELISA and PCR assays. These diagnostic methods are laborious, time consuming and need well trained personnel. Results A loop-mediated isothermal amplification (LAMP assay was developed and evaluated for detection of Y. ruckeri the etiological agent of enteric red mouth (ERM disease in salmonids. The assay was optimised to amplify the yruI/yruR gene, which encodes Y. ruckeri quorum sensing system, in the presence of a specific primer set and Bst DNA polymerase at an isothermal temperature of 63°C for one hour. Amplification products were detected by visual inspection, agarose gel electrophoresis and by real-time monitoring of turbidity resulted by formation of LAMP amplicons. Digestion with HphI restriction enzyme demonstrated that the amplified product was unique. The specificity of the assay was verified by the absence of amplification products when tested against related bacteria. The assay had 10-fold higher sensitivity compared with conventional PCR and successfully detected Y. ruckeri not only in pure bacterial culture but also in tissue homogenates of infected fish. Conclusion The ERM-LAMP assay represents a practical alternative to the microbiological approach for rapid, sensitive and specific detection of Y. ruckeri in fish farms. The assay is carried out in one hour and needs only a heating block or water bath as laboratory furniture. The advantages of the ERM-LAMP assay make it a promising tool for molecular detection of enteric red mouth disease in fish farms.

Loop mediated isothermal amplification assay using hydroxy naphthol blue, conventional polymerase chain reaction and real-time PCR in the diagnosis of intraocular tuberculosis

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P K Balne

2015-01-01

Full Text Available This study is a comparative evaluation (Chi-square test of a closed tube loop mediated isothermal amplification assay using hydroxy naphthol blue dye (HNB-LAMP, real-time polymerase chain reaction (PCR and conventional PCR in the diagnosis of intraocular tuberculosis. Considering clinical presentation as the gold standard in 33 patients, the sensitivity of HNB-LAMP assay (75.8% was higher (not significant, P value 0.2 than conventional PCR (57.6% and lower than real-time PCR (90.9%. Specificity was 100% by all three methods. No amplification was observed in negative controls (n = 20 by all three methods. The cost of the HNB-LAMP assay was Rs. 500.00 and it does not require thermocycler, therefore, it can be used as an alternative to conventional PCR in resource-poor settings.

A novel method of multiple nucleic acid detection: Real-time RT-PCR coupled with probe-melting curve analysis.

Science.gov (United States)

Han, Yang; Hou, Shao-Yang; Ji, Shang-Zhi; Cheng, Juan; Zhang, Meng-Yue; He, Li-Juan; Ye, Xiang-Zhong; Li, Yi-Min; Zhang, Yi-Xuan

2017-11-15

A novel method, real-time reverse transcription PCR (real-time RT-PCR) coupled with probe-melting curve analysis, has been established to detect two kinds of samples within one fluorescence channel. Besides a conventional TaqMan probe, this method employs another specially designed melting-probe with a 5' terminus modification which meets the same label with the same fluorescent group. By using an asymmetric PCR method, the melting-probe is able to detect an extra sample in the melting stage effectively while it almost has little influence on the amplification detection. Thus, this method allows the availability of united employment of both amplification stage and melting stage for detecting samples in one reaction. The further demonstration by simultaneous detection of human immunodeficiency virus (HIV) and hepatitis C virus (HCV) in one channel as a model system is presented in this essay. The sensitivity of detection by real-time RT-PCR coupled with probe-melting analysis was proved to be equal to that detected by conventional real-time RT-PCR. Because real-time RT-PCR coupled with probe-melting analysis can double the detection throughputs within one fluorescence channel, it is expected to be a good solution for the problem of low-throughput in current real-time PCR. Copyright © 2017 Elsevier Inc. All rights reserved.

Evaluation of loop-mediated isothermal amplification assay for rapid diagnosis of Acanthamoeba keratitis

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Abhishek Mewara

2017-01-01

Full Text Available Background: The clinical features of Acanthamoeba keratitis (AK are non-specific and closely resemble bacterial, viral and fungal keratitis. Materials and Methods: We compared loop-mediated isothermal amplification (LAMP with microscopy, non-nutrient agar (NNA culture and polymerase chain reaction (PCR in clinical suspects of AK. Results: Of 52 clinical samples (42 AK suspects and 10 proven bacterial, viral or fungal keratitis, 3 were positive by direct microscopy (sensitivity 60%, confidence interval [CI]: 17%–92.7%, and 5 by NNA culture, 18S rDNA PCR and LAMP (sensitivity 100%, CI: 46.3%–100%. The limit of detection of Acanthamoeba DNA was 1 pg/μl by both LAMP and PCR. Conclusion: PCR and LAMP assays targeting 18S rDNA gene were found particularly suitable for a rapid and accurate diagnosis of AK. LAMP assay takes 2–3 h lesser than PCR, and thus offers a rapid, highly sensitive and specific, simple and affordable diagnostic modality for patients suspected of AK, especially in resource limited settings

49 CFR 393.24 - Requirements for head lamps, auxiliary driving lamps and front fog lamps.

Science.gov (United States)

2010-10-01

... Devices, and Electrical Wiring § 393.24 Requirements for head lamps, auxiliary driving lamps and front fog lamps. (a) Headlamps. Every bus, truck and truck tractor shall be equipped with headlamps as required by...

Development of a loop-mediated isothermal amplification assay for the detection of Streptococcus agalactiae in bovine milk.

Science.gov (United States)

Bosward, Katrina L; House, John K; Deveridge, Amber; Mathews, Karen; Sheehy, Paul A

2016-03-01

Streptococcus agalactiae is a well-characterized bovine mastitis pathogen that is known to be highly contagious and capable of spreading rapidly in affected dairy herds. Loop-mediated isothermal amplification (LAMP) is a novel molecular diagnostic method that has the capability to provide rapid, cost-effective screening for pathogens to support on-farm disease control and eradication programs. In the current study, a LAMP test was developed to detect S. agalactiae in milk. The assay was validated on a bank of existing clinical mastitis milk samples that had previously been identified as S. agalactiae positive via traditional microbiological culture techniques and PCR. The LAMP assay was conducted on bacterial colonies and DNA extracted from milk in tube- and plate-based formats using multiple detection platforms. The 1-h assay conducted at 64 °C exhibited repeatability (coefficient of variation) of 2.07% (tube) and 8.3% (plate), sensitivity to ~20 pg of extracted DNA/reaction, and specificity against a panel of known bacterial mastitis pathogens. Of the 109 known S. agalactiae isolates assessed by LAMP directly from bacterial cells in culture, 108 were identified as positive, in accordance with PCR analysis. The LAMP analysis from the corresponding milk samples indicated that 104 of these milks exhibited a positive amplification curve. Although exhibiting some limitations, this assay provides an opportunity for rapid screening of milk samples to facilitate on-farm management of this pathogen. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Evaluation of Two Lyophilized Molecular Assays to Rapidly Detect Foot-and-Mouth Disease Virus Directly from Clinical Samples in Field Settings.

Science.gov (United States)

Howson, E L A; Armson, B; Madi, M; Kasanga, C J; Kandusi, S; Sallu, R; Chepkwony, E; Siddle, A; Martin, P; Wood, J; Mioulet, V; King, D P; Lembo, T; Cleaveland, S; Fowler, V L

2017-06-01

Accurate, timely diagnosis is essential for the control, monitoring and eradication of foot-and-mouth disease (FMD). Clinical samples from suspect cases are normally tested at reference laboratories. However, transport of samples to these centralized facilities can be a lengthy process that can impose delays on critical decision making. These concerns have motivated work to evaluate simple-to-use technologies, including molecular-based diagnostic platforms, that can be deployed closer to suspect cases of FMD. In this context, FMD virus (FMDV)-specific reverse transcription loop-mediated isothermal amplification (RT-LAMP) and real-time RT-PCR (rRT-PCR) assays, compatible with simple sample preparation methods and in situ visualization, have been developed which share equivalent analytical sensitivity with laboratory-based rRT-PCR. However, the lack of robust 'ready-to-use kits' that utilize stabilized reagents limits the deployment of these tests into field settings. To address this gap, this study describes the performance of lyophilized rRT-PCR and RT-LAMP assays to detect FMDV. Both of these assays are compatible with the use of fluorescence to monitor amplification in real-time, and for the RT-LAMP assays end point detection could also be achieved using molecular lateral flow devices. Lyophilization of reagents did not adversely affect the performance of the assays. Importantly, when these assays were deployed into challenging laboratory and field settings within East Africa they proved to be reliable in their ability to detect FMDV in a range of clinical samples from acutely infected as well as convalescent cattle. These data support the use of highly sensitive molecular assays into field settings for simple and rapid detection of FMDV. © 2015 The Authors. Transboundary and Emerging Diseases Published by Blackwell Verlag GmbH.

LAMP assay for rapid diagnosis of cow DNA in goat milk and meat samples.

Science.gov (United States)

Deb, R; Sengar, G S; Singh, U; Kumar, S; Raja, T V; Alex, R; Alyethodi, R R; Prakash, B

2017-01-01

Animal species detection is one of the crucial steps for consumer's food analysis. In the present study we developed an in-house built loop-mediated isothermal amplification (LAMP) assay for rapid detection of adulterated cow DNA in goat milk/meat samples. The cow milk/tissue DNA in goat milk/meat samples were identified in the developed LAMP assay by either naked eye visualizing with SYBR Green I dyes or by detecting the typical ladder pattern on gel electrophoresis. This test can detect up to minimum 5% level of cow components admixed in goat milk/meat samples and can be completed within 1 h 40 min starting from DNA extraction from milk/meat samples and can be performed in a water bath. Developed LAMP methodology is simple; rapid and sensitive techniques that can detect adulterant like cow components in goat milk/meat are more accurate than other existing DNA based technologies.

Rapid and sensitive detection of canine distemper virus by real-time reverse transcription recombinase polymerase amplification.

Science.gov (United States)

Wang, Jianchang; Wang, Jinfeng; Li, Ruiwen; Liu, Libing; Yuan, Wanzhe

2017-08-15

Canine distemper, caused by Canine distemper virus (CDV), is a highly contagious and fatal systemic disease in free-living and captive carnivores worldwide. Recombinase polymerase amplification (RPA), as an isothermal gene amplification technique, has been explored for the molecular detection of diverse pathogens. A real-time reverse transcription RPA (RT-RPA) assay for the detection of canine distemper virus (CDV) using primers and exo probe targeting the CDV nucleocapsid protein gene was developed. A series of other viruses were tested by the RT-RPA.Thirty-two field samples were further tested by RT-RPA, and the resuts were compared with those obtained by the real-time RT-PCR. The RT-RPA assay was performed successfully at 40 °C, and the results were obtained within 3 min-12 min. The assay could detect CDV, but did not show cross-detection of canine parvovirus-2 (CPV-2), canine coronavirus (CCoV), canine parainfluenza virus (CPIV), pseudorabies virus (PRV) or Newcastle disease virus (NDV), demonstrating high specificity. The analytical sensitivity of RT-RPA was 31.8 copies in vitro transcribed CDV RNA, which is 10 times lower than the real-time RT-PCR. The assay performance was validated by testing 32 field samples and compared to real-time RT-PCR. The results indicated an excellent correlation between RT-RPA and a reference real-time RT-PCR method. Both assays provided the same results, and R 2 value of the positive results was 0.947. The results demonstrated that the RT-RPA assay offers an alternative tool for simple, rapid, and reliable detection of CDV both in the laboratory and point-of-care facility, especially in the resource-limited settings.

Yersinia pestis detection by loop-mediated isothermal amplification combined with magnetic bead capture of DNA

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Na Feng

Full Text Available ABSTRACT We developed a loop-mediated isothermal amplification (LAMP assay for the detection of Y. pestis by targeting the 3a sequence on chromosome. All 11 species of the genus Yersinia were used to evaluate the specificity of LAMP and PCR, demonstrating that the primers had a high level of specificity. The sensitivity of LAMP or PCR was 2.3 or 23 CFU for pure culture, whereas 2.3 × 104 or 2.3 × 106 CFU for simulated spleen and lung samples. For simulated liver samples, the sensitivity of LAMP was 2.3 × 106 CFU, but PCR was negative at the level of 2.3 × 107 CFU. After simulated spleen and lung samples were treated with magnetic beads, the sensitivity of LAMP or PCR was 2.3 × 103 or 2.3 × 106 CFU, whereas 2.3 × 105 or 2.3 × 107 CFU for magnetic bead-treated liver samples. These results indicated that some components in the tissues could inhibit LAMP and PCR, and liver tissue samples had a stronger inhibition to LAMP and PCR than spleen and lung tissue samples. LAMP has a higher sensitivity than PCR, and magnetic bead capture of DNAs could remarkably increase the sensitivity of LAMP. LAMP is a simple, rapid and sensitive assay suitable for application in the field or poverty areas.

Direct duplex real-time loop mediated isothermal amplification assay for the simultaneous detection of cow and goat species origin of milk and yogurt products for field use.

Science.gov (United States)

Kim, Mi-Ju; Kim, Hae-Yeong

2018-04-25

A multiple loop-mediated isothermal amplification (LAMP) method was developed to detect cow and goat milk in the field using a portable fluorescence device. For rapid on-site detection, this duplex LAMP assay was used in combination with direct amplification, without DNA extraction. The cow- and goat-specific LAMP primer sets were designed based on the mitochondrial cytochrome b gene, and showed specificity against 13 other animal species in the reactions. The sensitivity of the duplex LAMP assay for cow and goat was 0.1 and 1 pg, respectively. The detection limit for both target species in milk mixtures was 2%. This assay successfully amplified and identified the two target species in 24 samples of commercial milk and yogurt products, with 30 min sampling-to-result analysis time. Therefore, this direct duplex real-time LAMP assay is useful for on-site simultaneous detection of cow and goat milk in commercial products, a capability needed to confirm accurate labeling. Copyright © 2017 Elsevier Ltd. All rights reserved.

Modelling the Dynamic Interaction Power System Lamp - Application to High Pressure Mercury Gas Discharge Lamps

OpenAIRE

ZIANE, M.; MEDLES, K.; ADJOUDJ, M.; MILOUA, F.; DAMELINCOURT, J. J.; TILMATINE, A.

2007-01-01

The aim of this paper is to study the dynamic behaviour of a plant constituted by an electrical power system and a gas discharge lamp, this latter, increasingly used in street lighting, remains a nonlinear load element. Various approaches are used to represent it, one is the approximation of the discharge represented by a hot "channel", which verifies the assumption of local thermodynamic equilibrium [LTE] or the polynomial form of the conductance variation. A calculation procedure, based on ...

The role of DNA amplification and cultural growth in complicated acute appendicitis

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Francesca Tocchioni

2016-09-01

Full Text Available Bacterial growth of peritoneal fluid specimens obtained during surgical procedures for acute appendicitis may be useful to optimize further antibiotic therapy in complicated cases. DNA amplification represents a fast technique to detect microbial sequences. We aimed to compare the potential of DNA amplification versus traditional bacterial growth culture highlighting advantages and drawbacks in a surgical setting. Peritoneal fluid specimens were collected during surgery from 36 children who underwent appendectomy between May and December 2012. Real-time polymerase chain reaction (RT-PCR and cultures were performed on each sample. RT-PCR showed an amplification of 16S in 18/36 samples, Escherichia coli (in 7 cases, Pseudomonas aeruginosa (3, Fusobacterium necrophorum (3, Adenovirus (2, E.coli (1, Klebsiella pneumoniae (1, Serratia marcescens/Enterobacter cloacae (1. Bacterial growth was instead observed only in four patients (3 E.coli and 1 P.aeruginosa and Bacteroides ovatus. Preoperative C-reactive protein and inflammation degree, the most reliable indicators of bacterial translocation, were elevated as expected. DNA amplification was a quick and useful method to detect pathogens and it was even more valuable in detecting aggressive pathogens such as anaerobes, difficult to preserve in biological cultures; its drawbacks were the lack of biological growths and of antibiograms. In our pilot study RT-PCR and cultures did not influence the way patients were treated.

Development of an electronic system for signals amplification

International Nuclear Information System (INIS)

Santos, Italo S.; Tobias, Carmen C.B.

2009-01-01

This paper presents the obtained results with a spectrometer for electromagnetic radiation whose detector, a Si PIN type diode, was directly coupled to a signal amplification system developed in this project for scientific initiation. The linearity conditions and the gain operational limits, constituted of two stages of amplification based on the employment of devices from AMTEK A225 and A206, were determined using a precision pulse generator. The obtained results shown that the developed system is stable and linear in the gain range of 50-150. The spectrometric response of the electronic system coupled to the Siemens SFH-00206 type diode, were studied in view of the register of the 59.5 keV gamma ray spectra proceeding from 241 Am as function of the reversal polarization voltage. The influence pf the voltage and the electronic contribution in the energy resolution of the registered spectra under room temperature (22 degree Celsius) had also investigated considering the more adequate value of the coupling capacitance of the amplification system diode. Up to the present. the best energy resolution (FWHM = 4.85 keV) of the 59.5 keV line was obtained for the condition of the detector polarization at 16 V. This result proves that the signal amplification system developed coupled to the SFH00206 diode, besides the low cost, excellent operational condition for the detection and spectrometry or low energy electromagnetic radiation

Genetic point-of-care diagnosis of Mycoplasma pneumoniae infection using LAMP assay.

Science.gov (United States)

Kakuya, Fujio; Kinebuchi, Takahiro; Fujiyasu, Hiroaki; Tanaka, Ryosuke; Kano, Hiroki

2014-08-01

Mycoplasma pneumoniae (MP) is a major pathogen of lower respiratory tract infection (LRTI) in children. A rapid diagnostic method during the acute phase is required for the prescription of effective antibiotics. A prospective, single-centered study was conducted on community-acquired LRTI in children. We regarded the day of fever onset as the first day of illness. In part 1, we studied 191 patients with signs of LRTI. We compared diagnostic reliability using loop-mediated isothermal amplification (LAMP) assay and serological testing at the first visit. In part 2, we evaluated the clinical characteristics of 117 patients with positive LAMP assay. In part 1, 31 patients met the definite MP infection criteria. LAMP assay had a sensitivity of 96.8% and specificity of 100%, whereas enzyme immunoassay had a sensitivity of 38.7% and specificity of 76.9%, and particle agglutination test had a sensitivity of 19.4% and specificity of 93.1%. In part 2, of 106 patients with fever, 100 patients were diagnosed by the day 7 of illness. The diagnosis was made a mean of 3.5 ± 2.1 days after the onset of fever. LAMP assay had excellent sensitivity and specificity for the detection of acute MP infection at the first visit. This assay can diagnose MP infection during the very acute phase. LAMP assay is appropriate for genetic point-of-care diagnosis of MP infection in hospital laboratories. © 2014 Japan Pediatric Society.

Time Course of Detection of Human Male DNA from Stained Blood Sample on Various Surfaces by Loop Mediated Isothermal Amplification and Polymerase Chain Reaction

OpenAIRE

Panan Kanchanaphum

2018-01-01

This study explores determining the sex of humans from blood stains taken from different surfaces and compares the time course of detection with the conventional PCR, Conventional Loop Mediated Isothermal Amplification (LAMP), and LAMP-Lateral Flow Dipstick (LFD). For the DNA templates, 7 male and 7 female blood stained samples were extracted and added to LAMP and PCR reaction solution to amplify the SRY gene. The DNA samples were extracted from the following blood stained materials: cloth, w...

Fast amplification system for gamma spectroscopy

International Nuclear Information System (INIS)

Jesus, E.F.O.; Lopes, R.T.

1992-01-01

An amplification system for gamma spectroscopy with high counting rates was developed. The system was constructed with operational amplifiers, and tested and compared with ORTEC conventional system, using Iridium-192 as source of 9,25 x 10 1 0 Bq of activity and NaI (Tl) detector. The constructed system showed a better performance in relation to efficiency and resolution parameters, tested before. (C.G.C.)

A novel approach for evaluating the performance of real time quantitative loop-mediated isothermal amplification-based methods.

Science.gov (United States)

Nixon, Gavin J; Svenstrup, Helle F; Donald, Carol E; Carder, Caroline; Stephenson, Judith M; Morris-Jones, Stephen; Huggett, Jim F; Foy, Carole A

2014-12-01

Molecular diagnostic measurements are currently underpinned by the polymerase chain reaction (PCR). There are also a number of alternative nucleic acid amplification technologies, which unlike PCR, work at a single temperature. These 'isothermal' methods, reportedly offer potential advantages over PCR such as simplicity, speed and resistance to inhibitors and could also be used for quantitative molecular analysis. However there are currently limited mechanisms to evaluate their quantitative performance, which would assist assay development and study comparisons. This study uses a sexually transmitted infection diagnostic model in combination with an adapted metric termed isothermal doubling time (IDT), akin to PCR efficiency, to compare quantitative PCR and quantitative loop-mediated isothermal amplification (qLAMP) assays, and to quantify the impact of matrix interference. The performance metric described here facilitates the comparison of qLAMP assays that could assist assay development and validation activities.

Geometric Modelling of Octagonal Lamp Poles

Science.gov (United States)

Chan, T. O.; Lichti, D. D.

2014-06-01

Lamp poles are one of the most abundant highway and community components in modern cities. Their supporting parts are primarily tapered octagonal cones specifically designed for wind resistance. The geometry and the positions of the lamp poles are important information for various applications. For example, they are important to monitoring deformation of aged lamp poles, maintaining an efficient highway GIS system, and also facilitating possible feature-based calibration of mobile LiDAR systems. In this paper, we present a novel geometric model for octagonal lamp poles. The model consists of seven parameters in which a rotation about the z-axis is included, and points are constrained by the trigonometric property of 2D octagons after applying the rotations. For the geometric fitting of the lamp pole point cloud captured by a terrestrial LiDAR, accurate initial parameter values are essential. They can be estimated by first fitting the points to a circular cone model and this is followed by some basic point cloud processing techniques. The model was verified by fitting both simulated and real data. The real data includes several lamp pole point clouds captured by: (1) Faro Focus 3D and (2) Velodyne HDL-32E. The fitting results using the proposed model are promising, and up to 2.9 mm improvement in fitting accuracy was realized for the real lamp pole point clouds compared to using the conventional circular cone model. The overall result suggests that the proposed model is appropriate and rigorous.

Recombinase Polymerase Amplification Assay for Rapid Diagnostics of Dengue Infection.

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Ahmed Abd El Wahed

Full Text Available Over 2.5 billion people are exposed to the risk of contracting dengue fever (DF. Early diagnosis of DF helps to diminish its burden on public health. Real-time reverse transcription polymerase amplification assays (RT-PCR are the standard method for molecular detection of the dengue virus (DENV. Real-time RT-PCR analysis is not suitable for on-site screening since mobile devices are large, expensive, and complex. In this study, two RT-recombinase polymerase amplification (RT-RPA assays were developed to detect DENV1-4.Using two quantitative RNA molecular standards, the analytical sensitivity of a RT-RPA targeting the 3´non-translated region of DENV1-4 was found to range from 14 (DENV4 to 241 (DENV1-3 RNA molecules detected. The assay was specific and did not cross detect other Flaviviruses. The RT-RPA assay was tested in a mobile laboratory combining magnetic-bead based total nucleic acid extraction and a portable detection device in Kedougou (Senegal and in Bangkok (Thailand. In Kedougou, the RT-RPA was operated at an ambient temperature of 38 °C with auxiliary electricity tapped from a motor vehicle and yielded a clinical sensitivity and specificity of 98% (n=31 and 100% (n=23, respectively. While in the field trial in Bangkok, the clinical sensitivity and specificity were 72% (n=90 and 100%(n=41, respectively.During the first 5 days of infection, the developed DENV1-4 RT-RPA assays constitute a suitable accurate and rapid assay for DENV diagnosis. Moreover, the use of a portable fluorescence-reading device broadens its application potential to the point-of-care for outbreak investigations.

Slit Lamp-Based Ocular Scoring Systems in Toxicology and Drug Development: A Literature Survey.

Science.gov (United States)

Eaton, Joshua Seth; Miller, Paul E; Bentley, Ellison; Thomasy, Sara M; Murphy, Christopher J

2017-12-01

To present a survey of the features of published slit lamp-based scoring systems and their applicability in the context of modern ocular toxicology and drug development. References describing original or modified slit lamp-based scoring systems for human or veterinary clinical patients or in investigative or toxicologic research were collected following a comprehensive literature review using textbooks and online publication searches. Each system's indications and features were compiled to facilitate comparison. Literature review identified 138 original or modified scoring systems. Most (48%) were published for evaluation of the ocular surface, 34% for the general anterior segment, and 18% for the lens. Most systems were described for assessment of human patients (50%) and small albino laboratory species such as rabbits (19%), rats (12%), and mice (8%). Systems described for pigmented laboratory species and for larger species such as dogs, cats, pigs, and nonhuman primates (NHPs) were comparatively underrepresented. No systems described a lens scoring scheme specific to the dog, cat, pig, or NHP. Scoring schemes for aqueous and vitreous cells were infrequently described for laboratory species. Many slit lamp-based scoring systems have been published, but the features of each differ and complicate translation of findings between different species. Use and interpretation of any scoring system in toxicology and drug development must be done with awareness of the limitations of the system being used.

Development and evaluation of a real-time fluorogenic loop-mediated isothermal amplification assay integrated on a microfluidic disc chip (on-chip LAMP) for rapid and simultaneous detection of ten pathogenic bacteria in aquatic animals.

Science.gov (United States)

Zhou, Qian-Jin; Wang, Lei; Chen, Jiong; Wang, Rui-Na; Shi, Yu-Hong; Li, Chang-Hong; Zhang, De-Min; Yan, Xiao-Jun; Zhang, Yan-Jun

2014-09-01

Rapid, low-cost, and user-friendly strategies are urgently needed for early disease diagnosis and timely treatment, particularly for on-site screening of pathogens in aquaculture. In this study, we successfully developed a real-time fluorogenic loop-mediated isothermal amplification assay integrated on a microfluidic disc chip (on-chip LAMP), which was capable of simultaneously detecting 10 pathogenic bacteria in aquatic animals, i.e., Nocardia seriolae, Pseudomonas putida, Streptococcus iniae, Vibrio alginolyticus, Vibrio anguillarum, Vibrio fluvialis, Vibrio harveyi, Vibrio parahaemolyticus, Vibrio rotiferianus, and Vibrio vulnificus. The assay provided a nearly-automated approach, with only a single pipetting step per chip for sample dispensing. This technique could achieve limits of detection (LOD) ranging from 0.40 to 6.42pg per 1.414μL reaction in less than 30 min. The robust reproducibility was demonstrated by a little variation among duplications for each bacterium with the coefficient of variation (CV) for time to positive (Tp) value less than 0.10. The clinical sensitivity and specificity of this on-chip LAMP assay in detecting field samples were 96.2% and 93.8% by comparison with conventional microbiological methods. Compared with other well-known techniques, on-chip LAMP assay provides low sample and reagent consumption, ease-of-use, accelerated analysis, multiple bacteria and on-site detection, and high reproducibility, indicating that such a technique would be applicable for on-site detection and routine monitoring of multiple pathogens in aquaculture. Copyright © 2014 Elsevier B.V. All rights reserved.

Clinical value of IS6110-based loop-mediated isothermal amplification for detection of Mycobacterium tuberculosis complex in respiratory specimens.

Science.gov (United States)

Aryan, Ehsan; Makvandi, Manoochehr; Farajzadeh, Ahmad; Huygen, Kris; Alvandi, Amir-Hooshang; Gouya, Mohammad-Mehdi; Sadrizadeh, Ali; Romano, Marta

2013-06-01

A fundamental to global tuberculosis (TB) control is timely and accurate diagnosis of infectious cases of the disease. Among various methods, techniques based on nucleic acid amplification are the ones with promising prospects. The present study evaluates the diagnostic value of the recently developed IS6110-based loop-mediated isothermal amplification (LAMP) for detection of Mycobacterium tuberculosis complex (MTBC) in sputum specimens. In this cross-sectional study (2008-2009), IS6110-LAMP was evaluated on 101 sputum specimens from 93 highly suspected TB patients and compared to Amplicor MTB test and in-house IS6110-PCR and -nested PCR assays. Culture results or clinical recovery following anti-TB therapy was considered as a reference to prove the TB cases. The overall sensitivity of IS6110-LAMP, Amplicor, nPCR, and PCR were respectively 89.6% (69/77 specimens; 95% confidence interval [CI], 80.5-95.4%), 76.6% (59/77 specimens; CI, 65.6-85.5%), 79.2% (61/77 specimens; CI, 68.5-87.6%) and 59.7% (46/77 specimens; CI, 47.9-70.8%). The specificity and positive predictive value (PPV) were 100% for all the tests, and the negative predictive value (NPV) of IS6110-LAMP, Amplicor, nPCR, and PCR were respectively 75%, 57.1%, 60%, and 43.6%. There was an excellent overall agreement between LAMP and nPCR (k 0.828), and between LAMP and Amplicor (k 0.746), in addition to a better tolerance of IS6110-LAMP to inhibitors present in clinical specimens. The better diagnostic performance of IS6110-LAMP compared to Amplicor (p = 0.009), nPCR (p = 0.013) and PCR (p < 0.0001) besides its rapidity, simplicity, and cost-effectiveness makes it a valuable method for the detection of MTBC in clinical samples, particularly in resource-limited settings. Copyright © 2013 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

A novel approach for evaluating the performance of real time quantitative loop-mediated isothermal amplification-based methods

Directory of Open Access Journals (Sweden)

Gavin J. Nixon

2014-12-01

Full Text Available Molecular diagnostic measurements are currently underpinned by the polymerase chain reaction (PCR. There are also a number of alternative nucleic acid amplification technologies, which unlike PCR, work at a single temperature. These ‘isothermal’ methods, reportedly offer potential advantages over PCR such as simplicity, speed and resistance to inhibitors and could also be used for quantitative molecular analysis. However there are currently limited mechanisms to evaluate their quantitative performance, which would assist assay development and study comparisons. This study uses a sexually transmitted infection diagnostic model in combination with an adapted metric termed isothermal doubling time (IDT, akin to PCR efficiency, to compare quantitative PCR and quantitative loop-mediated isothermal amplification (qLAMP assays, and to quantify the impact of matrix interference. The performance metric described here facilitates the comparison of qLAMP assays that could assist assay development and validation activities.

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

Science.gov (United States)

Nishiuchi, Yukiko; Tamaru, Aki; Suzuki, Yasuhiko; Kitada, Seigo; Maekura, Ryoji; Tateishi, Yoshitaka; Niki, Mamiko; Ogura, Hisashi; Matsumoto, Sohkichi

2014-06-01

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.

An Analysis of Sources of Technological Change in Efficiency Improvement of Fluorescent Lamp Systems

Science.gov (United States)

Imanaka, Takeo

In Japan, energy efficient fluorescent lamp systems which use “rare-earth phosphors” and “electronic ballasts” have shown rapid diffusion since 1990s. This report investigated sources of technological change in the efficiency improvement of fluorescent lamp systems: (i) Fluorescent lamp and luminaires have been under steady technological development for getting more energy efficient lighting and the concepts to achieve high efficiency had been found in such activities; however, it took long time until they realized and become widely used; (ii) Electronic ballasts and rare-earth phosphors add fluorescent lamp systems not only energy efficiency but also various values such as compactness, lightweight, higher output, and better color rendering properties, which have also been expected and have induced research and development (R&D) (iii) Affordable electronic ballasts are realized by the new technology “power MOSFET” which is based on IC technologies and has been developed for large markets of information and communication technologies and mobile devices; and (iv) Rare-earth phosphors became available after rare-earth industries developed for the purpose of supplying rare-earth phosphors for color television. In terms of sources of technological change, (i) corresponds to “R&D” aiming at the particular purpose i.e. energy efficiency in this case, on the other hand, (ii), (iii), and (iv) correspond to “spillovers” from activities aiming at other purposes. This case exhibits an actual example in which “spillovers” were the critical sources of technological change in energy technology.

Rapid and Sensitive Detection of sFAT-1 Transgenic Pigs by Visual Loop-Mediated Isothermal Amplification.

Science.gov (United States)

Tao, Chenyu; Yang, Yalan; Li, Xunbi; Zheng, Xinmin; Ren, Hongyan; Li, Kui; Zhou, Rong

2016-07-01

Genetically modified (GM) livestock have the potential to contribute to improving the environment and human health, with consumption of fewer resources and reduced waste production. However, the transgene process also poses risks. The safety assessment and control of transgenic animal products have drawn wide attention, and the relevant regulations and technology are being developed. Quick testing technology plays a significant role in on-site and customs sampling. Nowadays, loop-mediated isothermal amplification (LAMP) was widely applied in nucleic acid analysis because of its simplicity, rapidity, high efficiency and specificity. In this study, a specific, sensitive detection system for detecting sFAT-1 transgenic pigs was designed. A set of six primers including two loop primers was designed for the target sequence. The DNA samples were amplified in less than 1 h at the optimized temperature and detecting by both Nephelometer LA-320c and unaided eyes directly adding calcein. The detection limit of sFAT-1 LAMP was as low as 1.26 ng/μL. Furthermore, blind tests of transgenic and non-transgenic DNA samples were all correctly detected. Hence, the results in this study demonstrated that LAMP is a very useful tool for transgenic detection.

Validation of a Salmonella loop-mediated isothermal amplification assay in animal food.

Science.gov (United States)

Domesle, Kelly J; Yang, Qianru; Hammack, Thomas S; Ge, Beilei

2018-01-02

Loop-mediated isothermal amplification (LAMP) has emerged as a promising alternative to PCR for pathogen detection in food testing and clinical diagnostics. This study aimed to validate a Salmonella LAMP method run on both turbidimetry (LAMP I) and fluorescence (LAMP II) platforms in representative animal food commodities. The U.S. Food and Drug Administration (FDA)'s culture-based Bacteriological Analytical Manual (BAM) method was used as the reference method and a real-time quantitative PCR (qPCR) assay was also performed. The method comparison study followed the FDA's microbiological methods validation guidelines, which align well with those from the AOAC International and ISO. Both LAMP assays were 100% specific among 300 strains (247 Salmonella of 185 serovars and 53 non-Salmonella) tested. The detection limits ranged from 1.3 to 28 cells for six Salmonella strains of various serovars. Six commodities consisting of four animal feed items (cattle feed, chicken feed, horse feed, and swine feed) and two pet food items (dry cat food and dry dog food) all yielded satisfactory results. Compared to the BAM method, the relative levels of detection (RLODs) for LAMP I ranged from 0.317 to 1 with a combined value of 0.610, while those for LAMP II ranged from 0.394 to 1.152 with a combined value of 0.783, which all fell within the acceptability limit (2.5) for an unpaired study. This also suggests that LAMP was more sensitive than the BAM method at detecting low-level Salmonella contamination in animal food and results were available 3days sooner. The performance of LAMP on both platforms was comparable to that of qPCR but notably faster, particularly LAMP II. Given the importance of Salmonella in animal food safety, the LAMP assays validated in this study holds great promise as a rapid, reliable, and robust method for routine screening of Salmonella in these commodities. Published by Elsevier B.V.

One-tube loop-mediated isothermal amplification combined with restriction endonuclease digestion and ELISA for colorimetric detection of resistance to isoniazid, ethambutol and streptomycin in Mycobacterium tuberculosis isolates.

Science.gov (United States)

Lee, Mei-Feng; Chen, Yen-Hsu; Hsu, Hui-Jine; Peng, Chien-Fang

2010-10-01

In this study, we designed a simple and rapid colorimetric detection method, a one-tube loop-mediated isothermal amplification (LAMP)-PCR-hybridization-restriction endonuclease-ELISA [one-tube LAMP-PCR-HY-RE-ELISA] system, to detect resistance to isoniazid, ethambutol and streptomycin in strains of Mycobacterium tuberculosis isolated from clinical specimens. The clinical performance of this method for detecting isoniazid-resistant, ethambutol-resistant and streptomycin-resistant isolates of M. tuberculosis showed 98.9%, 94.3% and 93.8%, respectively. This assay is rapid and convenient that can be performed within one working day. One-tube LAMP-PCR-HY-RE-ELISA system was designed based on hot spot point mutations in target drug-resistant genes, using LAMP-PCR, hybridization, digestion with restriction endonuclease and colorimetric method of ELISA. In this study, LAMP assay was used to amplify DNA from drug-resistant M. tuberculosis, and ELISA was used for colorimetrical determination. This assay will be a useful tool for rapid diagnosis of mutant codons in strains of M. tuberculosis for isoniazid at katG 315 and katG 463, ethambutol at embB 306 and embB 497, and streptomycin at rpsL 43. Crown Copyright © 2010. Published by Elsevier B.V. All rights reserved.

Rapid, simple and direct detection of Meloidogyne hapla from infected root galls using loop-mediated isothermal amplification combined with FTA technology

OpenAIRE

Peng, Huan; Long, Haibo; Huang, Wenkun; Liu, Jing; Cui, Jiangkuan; Kong, Lingan; Hu, Xianqi; Gu, Jianfeng; Peng, Deliang

2017-01-01

The northern root-knot nematode (Meloidogyne hapla) is a damaging nematode that has caused serious economic losses worldwide. In the present study, a sensitive, simple and rapid method was developed for detection of M. hapla in infested plant roots by combining a Flinders Technology Associates (FTA) card with loop-mediated isothermal amplification (LAMP). The specific primers of LAMP were designed based on the distinction of internal transcribed spacer (ITS) sequences between M. hapla and oth...

High Expression of LAMP3 Is a Novel Biomarker of Poor Prognosis in Patients with Esophageal Squamous Cell Carcinoma

Directory of Open Access Journals (Sweden)

Xiaoyu Liao

2015-07-01

Full Text Available Lysosomal-associated membrane protein 3 (LAMP3, identified as a molecular marker of mature dendritic cells, is one of the LAMP family members. Its expression was induced by hypoxia, and was associated with hypoxia mediated metastasis in breast and cervical cancers. However, epithelial expression of LAMP3 and its prognostic value in esophageal squamous cell carcinoma (ESCC is still unknown. In the current study, mRNA expression of LAMP3 in 157 ESCC tissues and 50 adjacent normal tissues was detected by quantitative real-time PCR (qRT-PCR. LAMP3 protein expression in 46 paired cancerous and normal tissues was detected by immunohistochemistry (IHC. Then, DNA copy number was examined to observe its potential correlation with mRNA expression. The results showed that both mRNA and protein expression level of LAMP3 was significantly higher in cancerous tissues compared with normal controls (p < 0.001. LAMP3 DNA copy number was amplified in 70% of ESCC tissues and positive correlated with mRNA expression (p = 0.037. Furthermore, patients with higher LAMP3 expression had worse overall survival (HR = 1.90, 95% CI = 1.17–3.09, p = 0.010 and disease-free survival (HR = 1.80, 95% CI = 1.18–2.74, p = 0.006. In conclusion, our results suggest that epithelial LAMP3 expression is an independent prognostic biomarker for ESCC.

Modified Polyadenylation-Based RT-qPCR Increases Selectivity of Amplification of 3′-MicroRNA Isoforms

Directory of Open Access Journals (Sweden)

Charlotte Nejad

2018-01-01

Full Text Available MicroRNA (miRNA detection by reverse transcription (RT quantitative real-time PCR (RT-qPCR is the most popular method currently used to measure miRNA expression. Although the majority of miRNA families are constituted of several 3′-end length variants (“isomiRs”, little attention has been paid to their differential detection by RT-qPCR. However, recent evidence indicates that 3′-end miRNA isoforms can exhibit 3′-length specific regulatory functions, underlining the need to develop strategies to differentiate 3′-isomiRs by RT-qPCR approaches. We demonstrate here that polyadenylation-based RT-qPCR strategies targeted to 20–21 nt isoforms amplify entire miRNA families, but that primers targeted to >22 nt isoforms were specific to >21 nt isoforms. Based on this observation, we developed a simple method to increase selectivity of polyadenylation-based RT-qPCR assays toward shorter isoforms, and demonstrate its capacity to help distinguish short RNAs from longer ones, using synthetic RNAs and biological samples with altered isomiR stoichiometry. Our approach can be adapted to many polyadenylation-based RT-qPCR technologies already exiting, providing a convenient way to distinguish long and short 3′-isomiRs.

Development of a loop-mediated Isothermal amplification assay for sensitive and rapid detection of Vibrio parahaemolyticus

Directory of Open Access Journals (Sweden)

Kawahara Ryuji

2008-09-01

Full Text Available Abstract Background Vibrio parahaemolyticus is a marine seafood-borne pathogen causing gastrointestinal disorders in humans. Thermostable direct hemolysin (TDH and TDH-related hemolysin (TRH are known as major virulence determinants of V. parahaemolyticus. Most V. parahaemolyticus isolates from the environment do not produce TDH or TRH. Total V. parahaemolyticus has been used as an indicator for control of seafood contamination toward prevention of infection. Detection of total V. parahaemolyticus using conventional culture- and biochemical-based assays is time-consuming and laborious, requiring more than three days. Thus, we developed a novel and highly specific loop-mediated isothermal amplification (LAMP assay for the sensitive and rapid detection of Vibrio parahaemolyticus. Results The assay provided markedly more sensitive and rapid detection of V. parahaemolyticus strains than conventional biochemical and PCR assays. The assay correctly identified 143 V. parahaemolyticus strains, but did not detect 33 non-parahaemolyticus Vibrio and 56 non-Vibrio strains. Sensitivity of the LAMP assay for direct detection of V. parahaemolyticus in pure cultures and in spiked shrimp samples was 5.3 × 102 CFU per ml/g (2.0 CFU per reaction. The sensitivity of the LAMP assay was 10-fold more sensitive than that of the conventional PCR assay. The LAMP assay was markedly faster, requiring for amplification 13–22 min in a single colony on TCBS agar from each of 143 V. parahaemolyticus strains and less than 35 min in spiked shrimp samples. The LAMP assay for detection of V. parahaemolyticus required less than 40 min in a single colony on thiosulfate citrate bile salt sucrose (TCBS agar and 60 min in spiked shrimp samples from the beginning of DNA extraction to final determination. Conclusion The LAMP assay is a sensitive, rapid and simple tool for the detection of V. parahaemolyticus and will facilitate the surveillance for control of contamination of V

RT-PCR amplification of RNA extracted from formalin-fixed, paraffin-embedded oral cancer sections: analysis of p53 pathway.

Science.gov (United States)

Tachibana, Masatsugu; Shinagawa, Yasuhiro; Kawamata, Hitoshi; Omotehara, Fumie; Horiuchi, Hideki; Ohkura, Yasuo; Kubota, Keiichi; Imai, Yutaka; Fujibayashi, Takashi; Fujimori, Takahiro

2003-01-01

We present a new approach towards the detection of the mRNAs in formalin-fixed, paraffin-embedded samples using a reverse transcriptase (RT)-polymerase chain reaction (PCR). The total RNAs were extracted from 10-micron-thick sections and were reverse-transcribed, then the RT-products were subjected to PCR amplification of GAPDH mRNA for screening the mRNA degradation. Next, nested PCR was performed for examining the expression of p53-related genes, p21WAF1, MDM2, p33ING1 and p14ARF. GAPDH mRNA expression was detectable in 12 out of 21 oral squamous cell carcinoma (SCC) samples. p21WAF1 mRNA expression was detectable in 5 out of 12 SCC samples, MDM2 mRNA expression was detectable in 5 our of 12 SCC samples and p33ING1 mRNA expression was detectable in 6 out of 12 SCC samples. However, the expression of p14ARF mRNA was not detectable in any of the samples. Seven out of 12 oral SCC samples showed abnormal nuclear accumulation of p53 protein by immunohistochemical staining, whereas 5 out of 12 oral SCCs showed negative staining for p53 protein. Of of p33ING1 mRNA. One of these was a verrucous carcinoma in which the p53 gene products might be inactivated by the oncoprotein E6 of human papilloma virus. Thus, the p53 tumor suppressor pathway was disrupted in most oral SCCs at the cellular levels, due to either an abnormality in p53 itself or loss of expression of p53 regulatory factors. This method would assist in making diagnosis, determining therapeutic strategy and predicting the prognosis of various cancers including oral SCCs.

Amplification through chaotic synchronization in spatially extended beam-plasma systems

Science.gov (United States)

Moskalenko, Olga I.; Frolov, Nikita S.; Koronovskii, Alexey A.; Hramov, Alexander E.

2017-12-01

In this paper, we have studied the relationship between chaotic synchronization and microwave signal amplification in coupled beam-plasma systems. We have considered a 1D particle-in-cell numerical model of unidirectionally coupled beam-plasma oscillatory media being in the regime of electron pattern formation. We have shown the significant gain of microwave oscillation power in coupled beam-plasma media being in the different regimes of generation. The discovered effect has a close connection with the chaotic synchronization phenomenon, so we have observed that amplification appears after the onset of the complete time scale synchronization regime in the analyzed coupled spatially extended systems. We have also provided the numerical study of physical processes in the chain of beam-plasma systems leading to the chaotic synchronization and the amplification of microwave oscillations power, respectively.

Quantitative real-time RT-PCR and chromogenic in situ hybridization: precise methods to detect HER-2 status in breast carcinoma

International Nuclear Information System (INIS)

Rosa, Fabíola E; Silveira, Sara M; Silveira, Cássia GT; Bérgamo, Nádia A; Neto, Francisco A Moraes; Domingues, Maria AC; Soares, Fernando A; Caldeira, José RF; Rogatto, Silvia R

2009-01-01

HER-2 gene testing has become an integral part of breast cancer patient diagnosis. The most commonly used assay in the clinical setting for evaluating HER-2 status is immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). These procedures permit correlation between HER-2 expression and morphological features. However, FISH signals are labile and fade over time, making post-revision of the tumor difficult. CISH (chromogenic in situ hybridization) is an alternative procedure, with certain advantages, although still limited as a diagnostic tool in breast carcinomas. To elucidate the molecular profile of HER-2 status, mRNA and protein expression in 75 invasive breast carcinomas were analyzed by real time quantitative RT-PCR (qRT-PCR) and IHC, respectively. Amplifications were evaluated in 43 of these cases by CISH and in 11 by FISH. The concordance rate between IHC and qRT-PCR results was 78.9%, and 94.6% for qRT-PCR and CISH. Intratumoral heterogeneity of HER-2 status was identified in three cases by CISH. The results of the three procedures were compared and showed a concordance rate of 83.8%; higher discordances were observed in 0 or 1+ immunostaining cases, which showed high-level amplification (15.4%) and HER-2 transcript overexpression (20%). Moreover, 2+ immunostaining cases presented nonamplified status (50%) by CISH and HER-2 downexpression (38.5%) by qRT-PCR. In general, concordance occurred between qRT-PCR and CISH results. A high concordance was observed between CISH/qRT-PCR and FISH. Comparisons with clinicopathological data revealed a significant association between HER-2 downexpression and the involvement of less than four lymph nodes (P = 0.0350). Based on these findings, qRT-PCR was more precise and reproducible than IHC. Furthermore, CISH was revealed as an alternative and useful procedure for investigating amplifications involving the HER-2 gene

An enzymatic signal amplification system for calorimetric studies of cellobiohydrolases

DEFF Research Database (Denmark)

Murphy, Leigh; Baumann, Martin Johannes; Borch, Kim

2010-01-01

amplification method has been developed to measure even slow hydrolytically active enzymes such as cellobiohydrolases. This method is explained in detail for the amplification of the heat signal by more than 130 times by using glucose oxidase and catalase. The kinetics of this complex coupled reaction system...

Lamp-Ballast Compatibility Index for Efficient Ceramic Metal Halide Lamp Operation

OpenAIRE

Sourish Chatterjee

2013-01-01

Development of energy efficient products and exploration of energy saving potential are major challenges for present day’s technology. Ceramic Metal Halide lamp is the latest improved version of metal halide lamp that finds its wide applications in indoor commercial lighting especially in retail shop lighting. This lamp shows better performance in terms of higher lumen per watt and colour constancy in comparison to conventional metal halide lamp. The inherent negative incremental impedance of...

Analysis of the performance of domestic lighting lamps

International Nuclear Information System (INIS)

Aman, M.M.; Jasmon, G.B.; Mokhlis, H.; Bakar, A.H.A.

2013-01-01

The power crisis problem is getting worse in the developing countries. Measures are being taken to overcome the power shortage problem by efficiently utilizing the available power. Replacement of high-power consumption lamps with energy efficient lamps is also among these steps. This paper presents a detailed comparative analysis between domestic lighting lamps (DLLs) use for producing artificial light. DLLs include incandescent lamp (IL), fluorescent lamp (FL) and compact fluorescent lamp (CFL). Light emitting diodes (LED) based lamp technology is relatively new in comparison with conventional incandescent and discharge lamps. However, the present study will also cover the LED lamps. Power quality based experiments have been conducted on DLLs in Power System Laboratory and power consumption based calculations are carried out using the lighting design software DIALux. The result shows that with the current technology, the use of FL and LED lamp is beneficial for utility as well as for consumer. However, with the current pace in the development of LED technology, it is possible LED lamps will lead the lighting market in the near future. The paper has also presented the uncertainties that exist in lighting market and proposed the guidelines that will help in making future energy policy. - Highlights: ► Performances of domestic lighting lamps are compared. ► Power quality and power consumption based case study results are presented. ► For future energy policies, recommendations are also given.

A loop-mediated isothermal amplification assay and sample preparation procedure for sensitive detection of Xanthomonas fragariae in strawberry

Science.gov (United States)

Xanthomonas fragariae is a bacterium that causes angular leaf spot of strawberry. Asymptomatic infections are common and contribute to the difficulties in disease management. The aim of this study was to develop a loop-mediated isothermal amplification (LAMP) assay with a bacterial enrichment proced...

Development of a toxR-based loop-mediated isothermal amplification assay for detecting Vibrio parahaemolyticus

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Ge Beilei

2010-02-01

Full Text Available Abstract Background Vibrio parahaemolyticus is a leading cause of seafood-related bacterial gastroenteritis and outbreaks worldwide. Sensitive and specific detection methods are needed to better control V. parahaemolyticus infections. This study aimed at developing a highly specific and sensitive loop-mediated isothermal amplification (LAMP assay for detecting V. parahaemolyticus in oysters. A set of five LAMP primers, two outer, two inner, and one loop were designed based on the published V. parahaemolyticus toxR sequence. Specificity of the assay was evaluated using a panel of 36 V. parahaemolyticus and 39 other strains. The assay sensitivity was determined using serial dilutions of V. parahaemolyticus ATCC 27969 culture ranging from 108 CFU/ml to extinction. The assay was also tested in experimentally inoculated oyster samples. Results The toxR-based LAMP assay was able to specifically detect all of the 36 V. parahaemolyticus strains without amplification from 39 other strains. The detection limit was 47-470 cells per reaction in pure culture, up to 100-fold more sensitive than that of toxR-PCR. When applied in spiked oysters, the assay was able to detect 1.1 × 105 V. parahaemolyticus cells per gram of oyster without enrichment, up to 100-fold more sensitive than that of toxR-PCR. Standard curves generated for detecting V. parahaemolyticus in both pure culture and spiked oyster samples showed good linear relationship between cell numbers and the fluorescence or turbidity signals. Conclusions The toxR-based LAMP assay developed in this study was sensitive, specific, and quantitative, holding great potential for future field detection of V. parahaemolyticus in raw oysters.

Gluing for Raman lidar systems using the lamp mapping technique.

Science.gov (United States)

Walker, Monique; Venable, Demetrius; Whiteman, David N

2014-12-20

In the context of combined analog and photon counting (PC) data acquisition in a Lidar system, glue coefficients are defined as constants used for converting an analog signal into a virtual PC signal. The coefficients are typically calculated using Lidar profile data taken under clear, nighttime conditions since, in the presence of clouds or high solar background, it is difficult to obtain accurate glue coefficients from Lidar backscattered data. Here we introduce a new method in which we use the lamp mapping technique (LMT) to determine glue coefficients in a manner that does not require atmospheric profiles to be acquired and permits accurate glue coefficients to be calculated when adequate Lidar profile data are not available. The LMT involves scanning a halogen lamp over the aperture of a Lidar receiver telescope such that the optical efficiency of the entire detection system is characterized. The studies shown here involve two Raman lidar systems; the first from Howard University and the second from NASA/Goddard Space Flight Center. The glue coefficients determined using the LMT and the Lidar backscattered method agreed within 1.2% for the water vapor channel and within 2.5% for the nitrogen channel for both Lidar systems. We believe this to be the first instance of the use of laboratory techniques for determining the glue coefficients for Lidar data analysis.

Electrodeless discharge lamp is easily started, has high stability

Science.gov (United States)

Bell, W. E.; Bloom, A. L.

1966-01-01

Electrodeless discharge borosilicate glass lamp is used in various high-resolution optical systems. It is partially charged with krypton, contains small amounts of rubidium, and is enclosed in a hermetically sealed envelope that maintains the lamp at an optimum temperature during discharge. The lamp is quickly started by its excitation coil.

Loop-Mediated Isothermal Amplification Assay Targeting the MOMP Gene for Rapid Detection of Chlamydia psittaci Abortus Strain

Directory of Open Access Journals (Sweden)

Guo-Zhen Lin, Fu-Ying Zheng, Ji-Zhang Zhou, Guang-Hua Wang, Xiao-An Cao, Xiao-Wei Gong and Chang-Qing Qiu*

2012-05-01

Full Text Available For rapid detection of the Chlamydia psittaci abortus strain, a loop-mediated isothermal amplification (LAMP assay was developed and evaluated in this study. The primers for the LAMP assay were designed on the basis of the main outer membrane protein (MOMP gene sequence of C. psittaci. Analysis showed that the assay could detect the abortus strain of C. psittaci with adequate specificity. The sensitivity of the test was the same as that of the nested-conventional PCR and higher than that of chick embryo isolation. Testing of 153 samples indicated that the LAMP assay could detect the genome of the C. psittaci abortus strain effectively in clinical samples. This assay is a useful tool for rapid diagnosis of C. psittaci infection in sheep, swine and cattle.

Hollow-Core Fiber Lamp

Science.gov (United States)

Yi, Lin (Inventor); Tjoelker, Robert L. (Inventor); Burt, Eric A. (Inventor); Huang, Shouhua (Inventor)

2016-01-01

Hollow-core capillary discharge lamps on the millimeter or sub-millimeter scale are provided. The hollow-core capillary discharge lamps achieve an increased light intensity ratio between 194 millimeters (useful) and 254 millimeters (useless) light than conventional lamps. The capillary discharge lamps may include a cone to increase light output. Hollow-core photonic crystal fiber (HCPCF) may also be used.

UV lamp for photoelectron spectroscopy

International Nuclear Information System (INIS)

Cardoso, M.J.B.; Landers, R.; Sundaram, V.S.

1983-01-01

An UV lamp and a differential pumping system which enables to couple the lamp to an ultra-high vacuum chamber (10 -9 torr) without using windows, are described. The differential between the pressure inside the discharge chamber and the one in de UHV region, which is of 10 8 -10 9 , is achieved with two pumping states separated by pyrex capillaries having an internal diameter of 0.6 mm. In the first stage, a mechanical pump (10 -3 torr) is used; in the second stage, a diffusor pump with a cryogenic trap (N 2 liq - 10 -7 torr) is employed. The lamp produces, when used with high purity He, narrow lines almost clear at 21.2 eV and 40.8 eV, depending on the discharge chamber pressure, thus eliminating the need of a monochromator. As a high voltage source (3 KV), a commercial unit with a good current control was used, ensuring UV beam stability - an essential characteristic for this lamp if it is employed for photoelectron excitation of crystalline samples. (C.L.B.) [pt

Dirac's æther in curved spacetime-II: the geometric amplification of the cosmic magnetic induction

Directory of Open Access Journals (Sweden)

Marcelo Carvalho

2004-03-01

Full Text Available We search for an amplification mechanism of the seed cosmic magnetic induction by studying a new version of the Dirac's æther in a curved cosmological background. We find that the variation of the scale factor R(t with cosmic time brings to the magnetic field the desired effect of amplification, that we call geometric amplification.Procuramos por um mecanismo de amplificação das sementes do campo de indução magnética cósmico através do estudo de uma nova versão do æ ther de Dirac em uma arena cosmológica curva. Encontramos uma amplificação que ocorre se o fator de escala R(t varia com o tempo cósmico, acarretando um efeito o qual chamamos de amplificação geométrica do campo magnético.

Integrity Monitoring of Mercury Discharge Lamps

Science.gov (United States)

Tjoelker, Robert L.

2010-01-01

Mercury discharge lamps are critical in many trapped ion frequency standard applications. An integrity monitoring system can be implemented using end-of-life signatures observed in operational mercury discharge lamps, making it possible to forecast imminent failure and to take action to mitigate the consequences (such as switching to a redundant system). Mercury lamps are used as a source of 194-nm ultraviolet radiation for optical pumping and state selection of mercury trapped ion frequency standards. Lamps are typically fabricated using 202Hg distilled into high-purity quartz, or other 194-nm transmitting material (e.g., sapphire). A buffer gas is also placed into the bulb, typically a noble gas such as argon, neon, or krypton. The bulbs are driven by strong RF fields oscillating at .200 MHz. The lamp output may age over time by two internal mechanisms: (1) the darkening of the bulb that attenuates light transmission and (2) the loss of mercury due to migration or chemical interactions with the bulb surface. During fabrication, excess mercury is placed into a bulb, so that the loss rate is compensated with new mercury emanating from a cool tip or adjacent reservoir. The light output is nearly constant or varies slightly at a constant rate for many months/years until the mercury source is depleted. At this point, the vapor pressure abruptly falls and the total light output and atomic clock SNR (signal-to-noise ratio) decrease. After several days to weeks, the light levels decrease to a point where the atomic clock SNR is no longer sufficient to stay in lock, or the lamp self-extinguishes. This signature has been observed in four separate end-of-life lamp failures while operating in the Deep Space Network (DSN). A simple integrator circuit can observe and document steady-state lamp behavior. When the light levels drop over a predetermined time interval by a specified amount (e.g., 20 percent), an alarm is set. For critical operational applications, such as the DSN

Effects of Battery Voltage in Fluorescent Lamps for Use in Photovoltaic Systems; Efectos de la Tension de Bateria en Lamparas Fluorescentes para Uso en Instalaciones Fotovoltaicas

Energy Technology Data Exchange (ETDEWEB)

Garcia Rosillo, F; Egido Aguilera, M A

2008-07-01

It is known the importance of lighting systems with fluorescent lamps and electronic ballast in isolated photo-voltaic systems. The positive characteristics of the fluorescent lamps sometimes are not developed in full, due to the incorrect choice of ballast The failure of a lamp in the photovoltaic system can have great significance in the subjective perception of the user for the complete system. The purpose of this study is to analyze the behavior of two very similar fluorescent lamps, fed with different electronic ballasts, showing the great influence of the electronic ballast topology in the operation of the fluorescent lamp. In particular, this work aims to analyze the effect of variations of the input voltage to the ballast in the electrical and luminous characteristics of the lamp, as well as to understand the influence of electrical parameters in the life time of the fluorescent lamp. It is also analyzed the operating temperature of the electrode and the time necessary for electrode cooling after switching off the lamp. (Author) 23 refs.

49 CFR 393.25 - Requirements for lamps other than head lamps.

Science.gov (United States)

2010-10-01

... listed in paragraph (c) of this section. If motor vehicle equipment (e.g., mirrors, snow plows, wrecker...: J586—Stop Lamps for Use on Motor Vehicles Less Than 2032 mm in Overall Width, March 2000; J2261 Stop Lamps and Front- and Rear-Turn Signal Lamps for Use on Motor Vehicles 2032 mm or More in Overall Width...

The development of a loop-mediated isothermal amplification assay for rapid and sensitive detection of abalone herpesvirus DNA.

Science.gov (United States)

Chen, M H; Kuo, S T; Renault, T; Chang, P H

2014-02-01

A loop-mediated isothermal amplification (LAMP) assay was developed for the detection of abalone herpesvirus DNA. Two pairs of primers were designed, based on the sequence of the DNA polymerase gene of abalone herpesvirus. The reaction temperature and time were optimized to 63°C and 60min, respectively. LAMP amplicons were analyzed by 2% agarose gel electrophoresis or by visual inspection of a colour change emitted by fluorescent dye. The method developed was specific for the detection of abalone herpesvirus, without cross-reactions with other tested herpesviruses including ostreid herpesvirus 1 (OsHV-1), European eel herpesvirus, koi herpesvirus (KHV) and an avian herpesvirus. The LAMP assay was 100 folds more sensitive than a conventional PCR and 10 folds less sensitive than a SYBR Green PCR. These results indicate that the developed LAMP assay is a simple, rapid, sensitive, specific and reliable technique for the detection of abalone herpesvirus. Copyright © 2013 Elsevier B.V. All rights reserved.

Development of a multiplex amplification refractory mutation system reverse transcription polymerase chain reaction assay for the differential diagnosis of Feline leukemia virus vaccine and wild strains.

Science.gov (United States)

Ho, Chia-Fang; Chan, Kun-Wei; Yang, Wei-Cheng; Chiang, Yu-Chung; Chung, Yang-Tsung; Kuo, James; Wang, Chi-Young

2014-07-01

A multiplex amplification refractory mutation system reverse transcription polymerase chain reaction (ARMS RT-PCR) was developed for the differential diagnosis of Feline leukemia virus (FeLV) vaccine and wild-type strains based on a point mutation between the vaccine strain (S) and the wild-type strain (T) located in the p27 gene. This system was further upgraded to obtain a real-time ARMS RT-PCR (ARMS qRT-PCR) with a high-resolution melt analysis (HRMA) platform. The genotyping of various strains of FeLV was determined by comparing the HRMA curves with the defined wild-type FeLV (strain TW1), and the results were expressed as a percentage confidence. The detection limits of ARMS RT-PCR and ARMS qRT-PCR combined with HRMA were 100 and 1 copies of transcribed FeLV RNA per 0.5 ml of sample, respectively. No false-positive results were obtained with 6 unrelated pathogens and 1 feline cell line. Twelve FeLV Taiwan strains were correctly identified using ARMS qRT-PCR combined with HRMA. The genotypes of the strains matched the defined FeLV wild-type strain genotype with at least 91.17% confidence. A higher degree of sequence polymorphism was found throughout the p27 gene compared with the long terminal repeat region. In conclusion, the current study describes the phylogenetic relationship of the FeLV Taiwan strains and demonstrates that the developed ARMS RT-PCR assay is able to be used to detect the replication of a vaccine strain that has not been properly inactivated, thus acting as a safety check for the quality of FeLV vaccines.

Use of alternative alkali chlorides in RT and PCR of polynucleotides containing G quadruplex structures.

Science.gov (United States)

Ramos-Alemán, Fabiola; González-Jasso, Eva; Pless, Reynaldo C

2018-02-15

Several alkali chlorides were compared for their use in reverse transcription (RT) and PCR of different types of nucleic acid templates. On a test region of biological DNA incapable of forming G quadruplex (G4) structures, Taq DNA polymerase showed similar PCR performance with 50 mM KCl, CsCl, LiCl, and NaCl. In contrast, on a synthetic model polydeoxyribonucleotide prone to G4 formation, good PCR amplification was obtained with 50 mM CsCl, but little or none with LiCl or KCl. Similarly, in RT of a G4-prone model polyribonucleotide, MMLV reverse transcriptase produced a good yield with 50 mM CsCl, mediocre yields with LiCl or without added alkali chloride, and a poor yield with 50 mM KCl. The full RT-PCR assay starting from the G4-prone polyribonucleotide, showed good results with CsCl in both stages, poor results with LiCl, and no product formation with KCl. The model polynucleotides showed fast G quadruplex formation under PCR or RT conditions with 50 mM KCl, but not with CsCl or LiCl. The results argue for the use of CsCl instead of KCl for RT and PCR of G4-prone sequences. No advantage was observed when using the 7-deaza type nucleotide analog c 7 dGTP in PCR amplification of the G4-prone polydeoxyribonucleotide. Copyright © 2017 Elsevier Inc. All rights reserved.

Development of Nested PCR, Multiplex PCR, and Loop-Mediated Isothermal Amplification Assays for Rapid Detection of Cylindrocladium scoparium on Eucalyptus

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Tian-Min Qiao

2016-10-01

Full Text Available Eucalyptus dieback disease, caused by Cylindrocladium scoparium, has occurred in last few years in large Eucalyptus planting areas in China and other countries. Rapid, simple, and reliable diagnostic techniques are desired for the early detection of Eucalyptus dieback of C. scoparium prior to formulation of efficient control plan. For this purpose, three PCR-based methods of nested PCR, multiplex PCR, loop-mediated isothermal amplification (LAMP were developed for detection of C. scoparium based on factor 1-alpha (tef1 and beta-tubulin gene in this study. All of the three methods showed highly specific to C. scoparium. The sensitivities of the nested PCR and LAMP were much higher than the multiplex PCR. The sensitivity of multiplex PCR was also higher than regular PCR. C. scoparium could be detected within 60 min from infected Eucalyptus plants by LAMP, while at least 2 h was needed by the rest two methods. Using different Eucalyptus tissues as samples for C. scoparium detection, all of the three PCR-based methods showed much better detection results than regular PCR. Base on the results from this study, we concluded that any of the three PCR-based methods could be used as diagnostic technology for the development of efficient strategies of Eucalyptus dieback disease control. Particularly, LAMP was the most practical method in field application because of its one-step and rapid reaction, simple operation, single-tube utilization, and simple visualization of amplification products.

Development of Nested PCR, Multiplex PCR, and Loop-Mediated Isothermal Amplification Assays for Rapid Detection of Cylindrocladium scoparium on Eucalyptus

Science.gov (United States)

Qiao, Tian-Min; Zhang, Jing; Li, Shu-Jiang; Han, Shan; Zhu, Tian-Hui

2016-01-01

Eucalyptus dieback disease, caused by Cylindrocladium scoparium, has occurred in last few years in large Eucalyptus planting areas in China and other countries. Rapid, simple, and reliable diagnostic techniques are desired for the early detection of Eucalyptus dieback of C. scoparium prior to formulation of efficient control plan. For this purpose, three PCR-based methods of nested PCR, multiplex PCR, loop-mediated isothermal amplification (LAMP) were developed for detection of C. scoparium based on factor 1-alpha (tef1) and beta-tubulin gene in this study. All of the three methods showed highly specific to C. scoparium. The sensitivities of the nested PCR and LAMP were much higher than the multiplex PCR. The sensitivity of multiplex PCR was also higher than regular PCR. C. scoparium could be detected within 60 min from infected Eucalyptus plants by LAMP, while at least 2 h was needed by the rest two methods. Using different Eucalyptus tissues as samples for C. scoparium detection, all of the three PCR-based methods showed much better detection results than regular PCR. Base on the results from this study, we concluded that any of the three PCR-based methods could be used as diagnostic technology for the development of efficient strategies of Eucalyptus dieback disease control. Particularly, LAMP was the most practical method in field application because of its one-step and rapid reaction, simple operation, single-tube utilization, and simple visualization of amplification products. PMID:27721691

Development of Nested PCR, Multiplex PCR, and Loop-Mediated Isothermal Amplification Assays for Rapid Detection of Cylindrocladium scoparium on Eucalyptus.

Science.gov (United States)

Qiao, Tian-Min; Zhang, Jing; Li, Shu-Jiang; Han, Shan; Zhu, Tian-Hui

2016-10-01

Eucalyptus dieback disease, caused by Cylindrocladium scoparium , has occurred in last few years in large Eucalyptus planting areas in China and other countries. Rapid, simple, and reliable diagnostic techniques are desired for the early detection of Eucalyptus dieback of C. scoparium prior to formulation of efficient control plan. For this purpose, three PCR-based methods of nested PCR, multiplex PCR, loop-mediated isothermal amplification (LAMP) were developed for detection of C. scoparium based on factor 1-alpha (tef1) and beta-tubulin gene in this study. All of the three methods showed highly specific to C. scoparium . The sensitivities of the nested PCR and LAMP were much higher than the multiplex PCR. The sensitivity of multiplex PCR was also higher than regular PCR. C. scoparium could be detected within 60 min from infected Eucalyptus plants by LAMP, while at least 2 h was needed by the rest two methods. Using different Eucalyptus tissues as samples for C. scoparium detection, all of the three PCR-based methods showed much better detection results than regular PCR. Base on the results from this study, we concluded that any of the three PCR-based methods could be used as diagnostic technology for the development of efficient strategies of Eucalyptus dieback disease control. Particularly, LAMP was the most practical method in field application because of its one-step and rapid reaction, simple operation, single-tube utilization, and simple visualization of amplification products.

The enhanced pneumococcal LAMP assay: a clinical tool for the diagnosis of meningitis due to Streptococcus pneumoniae.

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Dong Wook Kim

Full Text Available BACKGROUND: Streptococcus pneumoniae is a leading cause of invasive bacterial disease in developed and developing countries. We studied the loop-mediated isothermal amplification (LAMP technique to assess its suitability for detecting S. pneumoniae nucleic acid in cerebrospinal fluid (CSF. METHODOLOGY/PRINCIPAL FINDINGS: We established an improved LAMP assay targeting the lytA gene (Streptococcus pneumoniae [Sp] LAMP. The analytical specificity of the primers was validated by using 32 reference strains (10 Streptococcus and seven non-Streptococcus species plus 25 clinical alpha-hemolytic streptococcal strains, including four S. pneumoniae strains and 21 other strains (3 S. oralis, 17 S. mitis, and one Streptococcus species harboring virulence factor-encoding genes (lytA or ply. Within 30 minutes, the assay could detect as few as 10 copies of both purified DNA and spiked CSF specimens with greater sensitivity than conventional polymerase chain reaction (PCR. The linear determination range for this assay is 10 to 1,000,000 microorganisms per reaction mixture using real-time turbidimetry. We evaluated the clinical sensitivity and specificity of the Sp LAMP assay using 106 randomly selected CSF specimens from children with suspected meningitis in Korea, China and Vietnam. For comparison, CSF specimens were also tested against conventional PCR and culture tests. The detection rate of the LAMP method was substantially higher than the rates of PCR and culture tests. In this small sample, relative to the LAMP assay, the clinical sensitivity of PCR and culture tests was 54.5% and 33.3%, respectively, while clinical specificity of the two tests was 100%. CONCLUSIONS/SIGNIFICANCE: Compared to PCR, Sp LAMP detected S. pneumoniae with higher analytical and clinical sensitivity. This specific and sensitive LAMP method offers significant advantages for screening patients on a population basis and for diagnosis in clinical settings.

Development of a Loop Mediated Isothermal Amplification for Diagnosis of Ascaris lumbricoides in Fecal Samples

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Esther A. Shiraho

2016-01-01

Full Text Available Ascaris lumbricoides is a nematode parasite that causes the common tropical infection ascariasis in humans. It is also considered among the neglected tropical diseases. Diagnosis relies mainly on microscopy-based methods which are laborious, are limited by low sensitivity, and require high expertise. We have developed a loop mediated isothermal amplification (LAMP for diagnosis of ascariasis in fecal samples, based on the first internal transcribed (ITS-1 spacer region of the ribosomal DNA. We used Primer Explorer V4 software to design primers. Ascaris adult and ova were obtained from naturally infected school children, whose parents/guardians gave consent for their participation in the study. Genomic DNA was extracted using alkaline lysis method and amplified by LAMP at 63°C for 45 minutes. LAMP products were visualized by naked eyes after adding SYBR Green dye and also on agarose gel. LAMP successfully and reliably detected Ascaris DNA from a single egg and in fecal samples. The assay specifically detected Ascaris DNA without amplifying DNA from ova of other parasites which commonly coexist with A. lumbricoides in feces. The developed LAMP assay has great potential for use in ascariasis diagnosis at the point of care and in low infection intensity situation that characterize control and elimination campaigns.

Rapid and Quantitative Detection of Vibrio parahemolyticus by the Mixed-Dye-Based Loop-Mediated Isothermal Amplification Assay on a Self-Priming Compartmentalization Microfluidic Chip.

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Pang, Bo; Ding, Xiong; Wang, Guoping; Zhao, Chao; Xu, Yanan; Fu, Kaiyue; Sun, Jingjing; Song, Xiuling; Wu, Wenshuai; Liu, Yushen; Song, Qi; Hu, Jiumei; Li, Juan; Mu, Ying

2017-12-27

Vibrio parahemolyticus (VP) mostly isolated from aquatic products is one of the major causes of bacterial food-poisoning events worldwide, which could be reduced using a promising on-site detection method. Herein, a rapid and quantitative method for VP detection was developed by applying a mixed-dye-loaded loop-mediated isothermal amplification (LAMP) assay on a self-priming compartmentalization (SPC) microfluidic chip, termed on-chip mixed-dye-based LAMP (CMD-LAMP). In comparison to conventional approaches, CMD-LAMP was advantageous on the limit of detection, which reached down to 1 × 10 3 CFU/mL in food-contaminated samples without the pre-enrichment of bacteria. Additionally, as a result of the use of a mixed dye and SPC chip, the quantitative result could be easily acquired, avoiding the requirement of sophisticated instruments and tedious operation. Also, CMD-LAMP was rapid and cost-effective. Conclusively, CMD-LAMP has great potential in realizing the on-site quantitative analysis of VP for food safety.

Discharge lamp technologies

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Dakin, James

1994-01-01

This talk is an overview of discharge lamp technology commonly employed in general lighting, with emphasis on issues pertinent to lighting for plant growth. Since the audience is primarily from the plant growth community, and this begins the light source part of the program, we will start with a brief description of the discharge lamps. Challenges of economics and of thermal management make lamp efficiency a prime concern in controlled environment agriculture, so we will emphasize science considerations relating to discharge lamp efficiency. We will then look at the spectra and ratings of some representative lighting products, and conclude with a discussion of technological advances.

A high-throughput method to detect RNA profiling by integration of RT-MLPA with next generation sequencing technology.

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Wang, Jing; Yang, Xue; Chen, Haofeng; Wang, Xuewei; Wang, Xiangyu; Fang, Yi; Jia, Zhenyu; Gao, Jidong

2017-07-11

RNA in formalin-fixed and paraffin-embedded (FFPE) tissues provides large amount of information indicating disease stages, histological tumor types and grades, as well as clinical outcomes. However, Detection of RNA expression levels in formalin-fixed and paraffin-embedded samples is extremely difficult due to poor RNA quality. Here we developed a high-throughput method, Reverse Transcription-Multiple Ligation-dependent Probe Sequencing (RT-MLPSeq), to determine expression levels of multiple transcripts in FFPE samples. By combining Reverse Transcription-Multiple Ligation-dependent Amplification method and next generation sequencing technology, RT-MLPSeq overcomes the limit of probe length in multiplex ligation-dependent probe amplification assay and thus could detect expression levels of transcripts without quantitative limitations. We proved that different RT-MLPSeq probes targeting on the same transcripts have highly consistent results and the starting RNA/cDNA input could be as little as 1 ng. RT-MLPSeq also presented consistent relative RNA levels of selected 13 genes with reverse transcription quantitative PCR. Finally, we demonstrated the application of the new RT-MLPSeq method by measuring the mRNA expression levels of 21 genes which can be used for accurate calculation of the breast cancer recurrence score - an index that has been widely used for managing breast cancer patients.

Quantitative real-time RT-PCR and chromogenic in situ hybridization: precise methods to detect HER-2 status in breast carcinoma

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Soares Fernando A

2009-03-01

Full Text Available Abstract Background HER-2 gene testing has become an integral part of breast cancer patient diagnosis. The most commonly used assay in the clinical setting for evaluating HER-2 status is immunohistochemistry (IHC and fluorescence in situ hybridization (FISH. These procedures permit correlation between HER-2 expression and morphological features. However, FISH signals are labile and fade over time, making post-revision of the tumor difficult. CISH (chromogenic in situ hybridization is an alternative procedure, with certain advantages, although still limited as a diagnostic tool in breast carcinomas. Methods To elucidate the molecular profile of HER-2 status, mRNA and protein expression in 75 invasive breast carcinomas were analyzed by real time quantitative RT-PCR (qRT-PCR and IHC, respectively. Amplifications were evaluated in 43 of these cases by CISH and in 11 by FISH. Results The concordance rate between IHC and qRT-PCR results was 78.9%, and 94.6% for qRT-PCR and CISH. Intratumoral heterogeneity of HER-2 status was identified in three cases by CISH. The results of the three procedures were compared and showed a concordance rate of 83.8%; higher discordances were observed in 0 or 1+ immunostaining cases, which showed high-level amplification (15.4% and HER-2 transcript overexpression (20%. Moreover, 2+ immunostaining cases presented nonamplified status (50% by CISH and HER-2 downexpression (38.5% by qRT-PCR. In general, concordance occurred between qRT-PCR and CISH results. A high concordance was observed between CISH/qRT-PCR and FISH. Comparisons with clinicopathological data revealed a significant association between HER-2 downexpression and the involvement of less than four lymph nodes (P = 0.0350. Conclusion Based on these findings, qRT-PCR was more precise and reproducible than IHC. Furthermore, CISH was revealed as an alternative and useful procedure for investigating amplifications involving the HER-2 gene.

Loop-mediated isothermal amplification assays for screening of bacterial integrons

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Guangchao Yu

2014-01-01

Full Text Available BACKGROUND: The occurrence and prevalence of integrons in clinical microorganisms and their role played in antimicrobial resistance have been well studied recently. As screening and detection of integrons are concerned, current diagnostic methodologies are restricted by significant drawbacks and novel methods are required for integrons detection. RESULTS: In this study, three loop-mediated isothermal amplification (LAMP assays targeting on class 1, 2 and 3 integrons were implemented and evaluated. Optimization of these detection assays were performed, including studing on the reaction temperature, volume, time, sensitivity and specificity (both primers and targets. Application of the established LAMP assays were further verified on a total of 1082 isolates (previously identified to be 397 integron-positive and 685 integron-negative strains. According to the results, the indispensability of each primer had been confirmed and the optimal reaction temperature, volume and time were found to be 65°C, 45 min and 25 µL, respectively. As application was concerned, 361, 28 and 8 isolates carrying intI1, intI2 and intI3 yielded positive amplicons, respectively. Other 685 integron-negative bacteria were negative for the integron-screening LAMP assays, totaling the detection rate and specificity to be 100%. CONCLUSIONS: The intI1-, intI2- and intI3-LAMP assays established in this study were demonstrated to be the valid and rapid detection methodologies for the screening of bacterial integrons.

Rapid, simple and direct detection of Meloidogyne hapla from infected root galls using loop-mediated isothermal amplification combined with FTA technology.

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Peng, Huan; Long, Haibo; Huang, Wenkun; Liu, Jing; Cui, Jiangkuan; Kong, Lingan; Hu, Xianqi; Gu, Jianfeng; Peng, Deliang

2017-04-03

The northern root-knot nematode (Meloidogyne hapla) is a damaging nematode that has caused serious economic losses worldwide. In the present study, a sensitive, simple and rapid method was developed for detection of M. hapla in infested plant roots by combining a Flinders Technology Associates (FTA) card with loop-mediated isothermal amplification (LAMP). The specific primers of LAMP were designed based on the distinction of internal transcribed spacer (ITS) sequences between M. hapla and other Meloidogyne spp. The LAMP assay can detect nematode genomic DNA at concentrations low to 1/200 000, which is 100 times more sensitive than conventional PCR. The LAMP was able to highly specifically distinguish M. hapla from other closely related nematode species. Furthermore, the advantages of the FTA-LAMP assay to detect M. hapla were demonstrated by assaying infected root galls that were artificially inoculated. In addition, M. hapla was successfully detected from six of forty-two field samples using FTA-LAMP technology. This study was the first to provide a simple diagnostic assay for M. hapla using the LAMP assay combined with FTA technology. In conclusion, the new FTA-LAMP assay has the potential for diagnosing infestation in the field and managing the pathogen M. hapla.

Development of Primer Sets for Loop-Mediated Isothermal Amplification that Enables Rapid and Specific Detection of Streptococcus dysgalactiae, Streptococcus uberis and Streptococcus agalactiae

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Deguo Wang

2015-05-01

Full Text Available Streptococcus dysgalactiae, Streptococcus uberis and Streptococcus agalactiae are the three main pathogens causing bovine mastitis, with great losses to the dairy industry. Rapid and specific loop-mediated isothermal amplification methods (LAMP for identification and differentiation of these three pathogens are not available. With the 16S rRNA gene and 16S-23S rRNA intergenic spacers as targets, four sets of LAMP primers were designed for identification and differentiation of S. dysgalactiae, S. uberis and S. agalactiae. The detection limit of all four LAMP primer sets were 0.1 pg DNA template per reaction, the LAMP method with 16S rRNA gene and 16S-23S rRNA intergenic spacers as the targets can differentiate the three pathogens, which is potentially useful in epidemiological studies.

Development of Primer Sets for Loop-Mediated Isothermal Amplification that Enables Rapid and Specific Detection of Streptococcus dysgalactiae, Streptococcus uberis and Streptococcus agalactiae.

Science.gov (United States)

Wang, Deguo; Liu, Yanhong

2015-05-26

Streptococcus dysgalactiae, Streptococcus uberis and Streptococcus agalactiae are the three main pathogens causing bovine mastitis, with great losses to the dairy industry. Rapid and specific loop-mediated isothermal amplification methods (LAMP) for identification and differentiation of these three pathogens are not available. With the 16S rRNA gene and 16S-23S rRNA intergenic spacers as targets, four sets of LAMP primers were designed for identification and differentiation of S. dysgalactiae, S. uberis and S. agalactiae. The detection limit of all four LAMP primer sets were 0.1 pg DNA template per reaction, the LAMP method with 16S rRNA gene and 16S-23S rRNA intergenic spacers as the targets can differentiate the three pathogens, which is potentially useful in epidemiological studies.

Breakdown of hot-spot model in determining convective amplification in large homogeneous systems

International Nuclear Information System (INIS)

Mounaix, Philippe; Divol, Laurent

2004-01-01

Convective amplification in large homogeneous systems is studied, both analytically and numerically, in the case of a linear diffraction-free stochastic amplifier. Overall amplification does not result from successive amplifications in small scale high intensity hot spots, but from a single amplification in a delocalized mode of the driver field spreading over the whole interaction length. For this model, the hot-spot approach is found to systematically underestimate the gain factor by more than 50%

Performance Evaluation of Irbene RT-16 Radio Telescope Receiving System

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Bleiders M.

2017-12-01

Full Text Available In the present paper, recent measurement results of refurbished Irbene RT-16 radio telescope receiving system performance are presented. The aim of the research is to evaluate characteristics of RT-16, which will allow carrying out necessary amplitude calibration in both single dish and VLBI observations, to improve the performance of existing system as well as to monitor, control and compare performance if possible changes in the receiving system will occur in future. The evaluated receiving system is 16 m Cassegrain antenna equipped with a cryogenic receiver with frequency range from 4.5 to 8.8 GHz, which is divided into four sub-bands. Multiple calibration sessions have been carried out by observing stable astronomical sources with known flux density by using in-house made total power registration backend. First, pointing offset calibration has been carried out and pointing model coefficients calculated and applied. Then, amplitude calibration, namely antenna sensitivity, calibration diode equivalent flux density and gain curve measurements have been carried out by observing calibration sources at different antenna elevations at each of the receiver sub-bands. Beam patterns have also been evaluated at different frequency bands. As a whole, acquired data will serve as a reference point for comparison in future performance evaluation of RT-16.

Max Tech and Beyond: Fluorescent Lamps

Energy Technology Data Exchange (ETDEWEB)

Scholand, Michael

2012-04-01

contains less material (i.e., glass, fill gas and phosphor), and has a higher luminance, enabling fixtures to take advantage of the smaller lamp size to improve the optics and provide more efficient overall system illuminance. In addition to offering the market a high-quality efficacious light source, another strong value proposition of fluorescent lighting is its long operating life. In today's market, one manufacturer is offering fluorescent lamps that have a rated life of 79,000 hours - which represents 18 years of service at 12 hours per day, 365 days per year. These lamps, operated using a long-life ballast specified by the manufacturer, take advantage of improvements in cathode coatings, fill gas chemistry and pressure to extend service life by a factor of four over conventional fluorescent lamps. It should be noted that this service life is also longer (approximately twice as long) as today's high-quality LED products. The fluorescent market is currently focused on the T5 and T8 lamp diameters, and it is not expected that other diameters would be introduced. Although T8 is a more optimal diameter from an efficacy perspective, the premium efficiency and optimization effort has been focused on T5 lamps because they are 40% smaller than T8, and are designed to operate at a higher temperature using high-frequency electronic ballasts. The T5 lamp offers savings in terms of materials, packaging and shipping, as well as smaller fixtures with improved optical performance. Manufacturers are actively researching improvements in four critical areas that are expected to yield additional efficacy improvements of approximately 10 to 14 percent over the next five years, ultimately achieving approximately 130 lumens per watt by 2015. The active areas of research where these improvements are anticipated include: (1) Improved phosphors which continue to be developed and patented, enabling higher efficacies as well as better color rendering and lumen maintenance; (2

Property of lysosomal storage disease associated with midbrain pathology in the central nervous system of Lamp-2-deficient mice.

Science.gov (United States)

Furuta, Akiko; Kikuchi, Hisae; Fujita, Hiromi; Yamada, Daisuke; Fujiwara, Yuuki; Kabuta, Tomohiro; Nishino, Ichizo; Wada, Keiji; Uchiyama, Yasuo

2015-06-01

Lysosome-associated membrane protein-2 (LAMP-2) is the gene responsible for Danon disease, which is characterized by cardiomyopathy, autophagic vacuolar myopathy, and variable mental retardation. To elucidate the function of LAMP-2 in the central nervous system, we investigated the neuropathological changes in Lamp-2-deficient mice. Immunohistochemical observations revealed that Lamp-1 and cathepsin D-positive lysosomal structures increased in the large neurons of the mouse brain. Ubiquitin-immunoreactive aggregates and concanavalin A-positive materials were detected in these neurons. By means of ultrastructural studies, we found various-shaped accumulations, including lipofuscin, glycolipid-like materials, and membranous structures, in the neurons and glial cells of Lamp-2-deficient brains. In deficient mice, glycogen granules accumulated in hepatocyte lysosomes but were not observed in neurons. These pathological features indicate lysosomal storage disease; however, the findings are unlikely a consequence of deficiency of a single lysosomal enzyme. Although previous study results have shown a large amount of autophagic vacuoles in parenchymal cells of the visceral organs, these findings were rarely detected in the brain tissue except for some axons in the substantia nigra, in which abundant activated microglial cells with increased lipid peroxidation were observed. Thus, LAMP-2 in the central nervous system has a possible role in the degradation of the various macromolecules in lysosomes and an additional function concerning protection from oxidative stress, especially in the substantia nigra. Copyright © 2015 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Deuterium lamps as transfer standards for spectral radiance measurements

International Nuclear Information System (INIS)

Key, P.J.; Nettleton, D.H.

1985-01-01

This report describes the work carried out at NPL and PTB to improve the performance of a low pressure deuterium discharge lamp, so that it can be used as a transfer standard in the spectral range 120 to 350 nm. To this end it was necessary: - to replace the original quartz windows by magnesium fluoride single crystal plates, which were cut perpendicular to the c-axis of the crystal and which had to be free of impurities, - to construct the lamps in that way that the directional uniformity of the emitted radiation is within the demands, - to age the lamps and to preselect only those of which the irradiance was stable within ± 1% during a thirty minute period after warm-up, - to improve the commercially available electrical power supply to meet the operational needs of the lamps. Thus, the deuterium lamps drifted by about 3% over a period of 100 h at all wavelengths except at 250 nm, where the ageing increased to 4.5%. A liquid nitrogen trap has been developed which can be installed between the vacuum system and the lamp. This reduced to about 2% the decrease of the window's transmission during the first hour of operation, caused by the deposition of oil from the vacuum system

Rapid detection of Puccinia triticina causing leaf rust of wheat by PCR and loop mediated isothermal amplification.

Science.gov (United States)

Manjunatha, C; Sharma, Sapna; Kulshreshtha, Deepika; Gupta, Sangeeta; Singh, Kartar; Bhardwaj, Subhash C; Aggarwal, Rashmi

2018-01-01

Leaf rust of wheat caused by Puccinia triticina has significant impact on wheat production worldwide. Effective and quick detection methodologies are required to mitigate yield loss and time constraints associated with monitoring and management of leaf rust of wheat. In the present study, detection of P. triticina has been simplified by developing a rapid, reliable, efficient and visual colorimetric method i.e., loop mediated isothermal amplification of DNA (LAMP). Based on in silico analysis of P. triticina genome, PTS68, a simple sequence repeat was found highly specific to leaf rust fungus. A marker (PtRA68) was developed and its specificity was validated through PCR technique which gave a unique and sharp band of 919 bp in P. triticina pathotypes only. A novel gene amplification method LAMP which enables visual detection of pathogen by naked eye was developed for leaf rust pathogen. A set of six primers was designed from specific region of P. triticina and conditions were optimised to complete the observation process in 60 minutes at 65o C. The assay developed in the study could detect presence of P. triticina on wheat at 24 hpi (pre-symptomatic stage) which was much earlier than PCR without requiring thermal cycler. Sensitivity of LAMP assay developed in the study was 100 fg which was more sensitive than conventional PCR (50 pg) and equivalent to qPCR (100 fg). The protocol developed in the study was utilized for detection of leaf rust infected samples collected from different wheat fields. LAMP based colorimetric detection assay showed sky blue color in positive reaction and violet color in negative reaction after addition of 120 μM hydroxyl napthol blue (HNB) solution to reaction mixture. Similarly, 0.6 mg Ethidium bromide/ml was added to LAMP products, placed on transilluminator to witness full brightness in positive reaction and no such brightness could be seen in negative reaction mixture. Further, LAMP products spread in a ladder like banding pattern in

Wood's lamp examination

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... this page: //medlineplus.gov/ency/article/003386.htm Wood lamp examination To use the sharing features on this page, please enable JavaScript. A Wood lamp examination is a test that uses ultraviolet ( ...

Rapid Detection of Prunus Necrotic Ringspot Virus by Reverse Transcription-cross-priming Amplification Coupled with Nucleic Acid Test Strip Cassette.

Science.gov (United States)

Huo, Ya-Yun; Li, Gui-Fen; Qiu, Yan-Hong; Li, Wei-Min; Zhang, Yong-Jiang

2017-11-23

Prunus necrotic ringspot virus (PNRSV) is one of the most devastating viruses to Prunus spp. In this study, we developed a diagnostic system RT-CPA-NATSC, wherein reverse transcription-cross-priming amplification (RT-CPA) is coupled with nucleic acid test strip cassette (NATSC), a vertical flow (VF) visualization, for PNRSV detection. The RT-CPA-NATSC assay targets the encoding gene of the PNRSV coat protein with a limit of detection of 72 copies per reaction and no cross-reaction with the known Prunus pathogenic viruses and viroids, demonstrating high sensitivity and specificity. The reaction is performed on 60 °C and can be completed less than 90 min with the prepared template RNA. Field sample test confirmed the reliability of RT-CPA-NATSC, indicating the potential application of this simple and rapid detection method in routine test of PNRSV.

Detection and differentiation of Fusarium oxysporum f. sp. lycopersici race 1 using loop-mediated isothermal amplification with three primer sets.

Science.gov (United States)

Ayukawa, Y; Komatsu, K; Kashiwa, T; Akai, K; Yamada, M; Teraoka, T; Arie, T

2016-09-01

Fusarium oxysporum f. sp. lycopersici (Fol) causes tomato wilt. Based on the difference in pathogenicity towards tomato cultivars, Fol is classified into three races. In this study, a rapid method is developed for the detection and discrimination of Fol race 1 using a loop-mediated isothermal amplification (LAMP) assay with two primer sets targeting a region of the nucleotide sequence of the SIX4 gene specific for race 1 and a primer set targeting the SIX5 gene, conserved in all known Fol isolates. Upon LAMP reaction, amplification using all three primer sets was observed only when DNA of Fol race 1 was used as a template, and not when DNA of other Fol races or other fungal species was used. This method could detect 300 fg of Fol race 1 DNA, a 100-fold higher sensitivity than that obtained by conventional PCR. The method can also detect DNA extracted from soil artificially infested with Fol race 1. It is now possible to detect Fol race 1 in colonies and infected tomato stems without DNA isolation. This method is a rapid and simple tool for discrimination of Fol race 1. This study developed a loop-mediated isothermal amplification (LAMP) assay for detection and differentiation of Fusarium oxysporum f. sp. lycopersici (Fol) race 1 by using three primer sets targeting for the SIX4 and SIX5 genes. These genes are present together only in Fol race 1. This method can detect Fol race 1 in infected tomato stems without DNA extraction, affording an efficient diagnosis of Fusarium wilt on tomatoes in the field. © 2016 The Society for Applied Microbiology.

Sensitive Detection of Plasmodium vivax Using a High-Throughput, Colourimetric Loop Mediated Isothermal Amplification (HtLAMP Platform: A Potential Novel Tool for Malaria Elimination.

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Sumudu Britton

2016-02-01

Full Text Available Plasmodium vivax malaria has a wide geographic distribution and poses challenges to malaria elimination that are likely to be greater than those of P. falciparum. Diagnostic tools for P. vivax infection in non-reference laboratory settings are limited to microscopy and rapid diagnostic tests but these are unreliable at low parasitemia. The development and validation of a high-throughput and sensitive assay for P. vivax is a priority.A high-throughput LAMP assay targeting a P. vivax mitochondrial gene and deploying colorimetric detection in a 96-well plate format was developed and evaluated in the laboratory. Diagnostic accuracy was compared against microscopy, antigen detection tests and PCR and validated in samples from malaria patients and community controls in a district hospital setting in Sabah, Malaysia.The high throughput LAMP-P. vivax assay (HtLAMP-Pv performed with an estimated limit of detection of 1.4 parasites/ μL. Assay primers demonstrated cross-reactivity with P. knowlesi but not with other Plasmodium spp. Field testing of HtLAMP-Pv was conducted using 149 samples from symptomatic malaria patients (64 P. vivax, 17 P. falciparum, 56 P. knowlesi, 7 P. malariae, 1 mixed P. knowlesi/P. vivax, with 4 excluded. When compared against multiplex PCR, HtLAMP-Pv demonstrated a sensitivity for P. vivax of 95% (95% CI 87-99%; 61/64, and specificity of 100% (95% CI 86-100%; 25/25 when P. knowlesi samples were excluded. HtLAMP-Pv testing of 112 samples from asymptomatic community controls, 7 of which had submicroscopic P. vivax infections by PCR, showed a sensitivity of 71% (95% CI 29-96%; 5/7 and specificity of 93% (95% CI87-97%; 98/105.This novel HtLAMP-P. vivax assay has the potential to be a useful field applicable molecular diagnostic test for P. vivax infection in elimination settings.

Influence of Voltage on Main Characteristics of Electric Lighting Lamps

Directory of Open Access Journals (Sweden)

V. B. Kozlovskaya

2009-01-01

Full Text Available An analysis and systemization of data on influence of voltage value on main lighting engineering, electric and economic characteristics of incandescent lamps, gaseous-discharge lamps of low and high pressure have been made in the paper.Analytical and graphical dependences have been obtained that ensure to evaluate quantitative changes of corresponding lamp characteristics at voltage deviation from nominal value.

Retail Lamps Study 3.1: Dimming, Flicker, and Power Quality Characteristics of LED A Lamps

Energy Technology Data Exchange (ETDEWEB)

Royer, Michael P. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Poplawski, Michael E. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Brown, Charles C. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

2014-12-01

To date, all three reports in the retail lamps series have focused on basic performance parameters, such as lumen output, efficacy, and color quality. This report goes a step further, examining the photoelectric characteristics (i.e., dimming and flicker) of a subset of lamps from CALiPER Retails Lamps Study 3. Specifically, this report focuses on the dimming, power quality, and flicker characteristics of 14 LED A lamps, as controlled by four different retail-available dimmers. The results demonstrate notable variation across the various lamps, but little variation between the four dimmers. Overall, the LED lamps: ~tended to have higher relative light output compared to the incandescent and halogen benchmark at the same dimmer output signal (RMS voltage). The lamps’ dimming curves (i.e., the relationship between control signal and relative light output) ranged from linear to very similar to the square-law curve typical of an incandescent lamp. ~generally exhibited symmetrical behavior—the same dimming curve—when measured proceeding from maximum to minimum or minimum to maximum control signal. ~mostly dimmed below 10% of full light output, with some exceptions for specific lamp and dimmer combinations ~exhibited a range of flicker characteristics, with many comparing favorably to the level typical of a magnetically-ballasted fluorescent lamp through at least a majority of the dimming range. ~ always exceeded the relative (normalized) efficacy over the dimming range of the benchmark lamps, which rapidly decline in efficacy when they are dimmed. This report generally does not attempt to rank the performance of one product compared to another, but instead focuses on the collective performance of the group versus conventional incandescent or halogen lamps, the performance of which is likely to be the baseline for a majority of consumers. Undoubtedly, some LED lamps perform better—or more similar to conventional lamps—than others. Some perform desirably for one

Comparative study of energy-efficiency and conservation systems for ceramic metal-halide discharge lamps

International Nuclear Information System (INIS)

Hermoso Orzáez, Manuel Jesús; Andrés Díaz, José Ramón de

2013-01-01

Interest in energy savings in urban lighting is gaining traction and has become a priority for municipal administrations. LED (light-emitting diode) technology appears to be the clear future lighting choice. However, this technology is still rapidly developing and has not been sufficiently tested. As an intermediate step, alternative proposals for energy-saving equipment for traditional discharge lamps are desirable so that the current technologies can coexist with the new LED counterparts for the short and medium term. This article provides a comparative study between two efficiency and energy-saving systems for discharge lamps with metal-halide and ceramic technologies, i.e., a lighting flow dimmer-stabilizer and a double-level electronic ballast. - Highlights: ► It has been demonstrated the possibility of regulating ceramic metal-halide lamps with lighting flow dimmer-stabilizer. ► Electronic ballasts can save approximately double quantity of energy than lighting flow dimmer-stabilizers. ► The use of lighting flow dimmer-stabilizer is more profitable than electronic ballasts due to costs and reliability

A real-time loop-mediated isothermal amplification assay for rapid detection of Shigella species.

Science.gov (United States)

Liew, P S; Teh, C S J; Lau, Y L; Thong, K L

2014-12-01

Shigellosis is a foodborne illness caused by the genus Shigella and is an important global health issue. The development of effective techniques for rapid detection of this pathogen is essential for breaking the chain of transmission. Therefore, we have developed a novel loop-mediated isothermal amplification (LAMP) assay targeting the invasion plasmid antigen H (ipaH) gene to rapidly detect Shigella species. This assay could be performed in 90 min at an optimal temperature of 64ºC, with endpoint results visualized directly. Notably, the method was found to be more sensitive than conventional PCR. Indeed, the detection limit for the LAMP assay on pure bacterial cultures was 5.9 x 10(5) CFU/ml, while PCR displayed a limit of 5.9 x 10(7) CFU/ml. In spiked lettuce samples, the sensitivity of the LAMP assay was 3.6 x 10(4) CFU/g, whereas PCR was 3.6 x 10(5) CFU/g. Overall, the assay accurately identified 32 Shigella spp. with one enteroinvasive Escherichia coli displaying positive reaction while the remaining 32 non-Shigella strains tested were negative.

Discharge lamp technologies

Energy Technology Data Exchange (ETDEWEB)

Dakin, J. [GE Lighting, Cleveland, OH (United States)

1994-12-31

This talk is an overview of discharge lamp technology commonly employed in general lighting, with emphasis on issues pertinent to lighting for plant growth. Since the audience is primarily from the plant growth community, and this begins the light source part of the program, we will start with a brief description of the discharge lamps. Challenges of economics and of thermal management make lamp efficiency a prime concern in controlled environment agriculture, so we will emphasize science considerations relating to discharge lamp efficiency. We will then look at the spectra and ratings of some representative lighting products, and conclude with a discussion of technological advance. A general overview of discharge lighting technology can be found in the book of Waymouth (1971). A recent review of low pressure lighting discharge science is found in Dakin (1991). The pioneering paper of Reiling (1964) provides a good introduction to metal halide discharges. Particularly relevant to lighting for plant growth, a recent and thorough treatment of high pressure Na lamps is found in the book by deGroot and vanVliet (1986). Broad practical aspects of lighting application are thoroughly covered in the IES Lighting Handbook edited by Kaufman (1984).

Detection and characterization of Newcastle disease virus in clinical samples using real time RT-PCR and melting curve analysis based on matrix and fusion genes amplification

Directory of Open Access Journals (Sweden)

Saad Sharawi

2013-10-01

Full Text Available Aim: Newcastle disease is still one of the major threats for poultry industry allover the world. Therefore, attempt was made in this study to use the SYBR Green I real-time PCR with melting curves analysis as for detection and differentiation of NDV strains in suspected infected birds. Materials and Methods: Two sets of primers were used to amplify matrix and fusion genes in samples collected from suspectly infected birds (chickens and pigeons. Melting curve analysis in conjunction with real time PCR was conducted for identifying different pathotypes of the isolated NDVs. Clinical samples were propagated on specific pathogen free ECE and tested for MDT and ICIP. Results: The velogenic NDVs isolated from chickens and pigeons were distinguished with mean T 85.03±0.341 and m 83.78±0.237 respectively for M-gene amplification and for F-gene amplification the mean T were 84.04±0.037 and m 84.53±0.223. On the other hand the lentogenic NDV isolates including the vaccinal strains (HB1 and LaSota have a higher mean T (86.99±0.021 for M-gene amplification and 86.50±0.063 for F-gene amplification. The test showed no reaction with m unrelated RNA samples. In addition, the results were in good agreement with both virus isolation and biological pathotyping (MDT and ICIP. The assay offers an attractive alternative method for the diagnosis of NDV that can be easily applied in laboratory diagnosis as a screening test for the detection and differentiation of NDV infections. Conclusion: As was shown by the successful rapid detection and pathotyping of 15 NDV strains in clinical samples representing velogenic and lentogenic NDV strains, and the agreement with the results of virus isolation , biological pathotyping and pathogenicity indices. The results of this report suggests that the described SybrGreen I real-time RT-PCR assay in conjunction with Melting curve analysis used as a rapid, specific and simple diagnostic tools for detection and pathotyping of

A rapid and specific detection of pathogenic serovar Salmonella typhimurium by loop-mediated isothermal amplification method (LAMP

Directory of Open Access Journals (Sweden)

Hadi Ravan

2017-09-01

Discussion and conclusion: As a result of a high sensitivity and specificity of the method as well as its low cost per assay, it could be concluded that the present LAMP assay is a powerful, accurate, and efficient method for detecting pathogenic serovar Salmonella typhimurium in food-processing industries and diagnostic laboratories.

Optimization of an Optical Parametric Chirped Pulse Amplification System for the OMEGA EP Laser System

International Nuclear Information System (INIS)

Begishev, I.; Bagnoud, V.; Guardalben, M.; Waxer, L.; Puth, J.; Zuegel, J.

2003-01-01

OAK B204 We report on the experimental achievements of the optical parametric chirped-pulse amplification (OPCPA) system, including 29% pump-to-signal conversion efficiency and 107 gain using two LBO crystals configured as a single amplification stage. Temporal and spatial shaping of the pump laser pulse is required to achieve both high-gain and high-conversion efficiency

DaRT: A CALL System to Help Students Practice and Develop Reasoning Skills in Choosing English Articles.

Science.gov (United States)

Yoshii, Rika; Milne, Alastair

1998-01-01

Describes DaRT, a computer assisted language-learning system for helping English-as-a-Second-Language students master English articles. DaRT uses a diagrammatic reasoning tool to present communicative contexts for exercises in choosing appropriate articles. This paper describes the development of DaRT and DaRT's system components and concludes…

Loop-mediated isothermal amplification as a reliable assay for Toxocara canis infection in pet dogs.

Science.gov (United States)

Khoshakhlagh, Paria; Spotin, Adel; Mahami-Oskouei, Mahmoud; Shahbazi, Abbas; Ozlati, Maryam

2017-09-01

Keeping of infected dogs as pet results in the potential transmission risk factors for shedding helminthic infections such as toxocariasis. Lack of accurate identification of Toxocara canis eggs in non-dewormed infected pet dogs remains a diagnostic concern among researchers. In this study, dog owners were asked to fill up a questionnaire regarding their pets and their attitude towards the deworming regimen. One hundred faecal samples were collected from pet dogs (Northwest Iran) and were subsequently identified by the ZnSo4 flotation technique, PCR and loop-mediated isothermal amplification (LAMP) assays. The DNA of the recovered T. canis eggs was then extracted and amplified by LAMP and PCR. Furthermore, ITS2 amplicons were sequenced for appraisal of the phylogenetic analysis. Nine, 5 and 11% of T. canis infections were identified by microscopy, PCR and LAMP, respectively. It was detected that LAMP was 10 times (10 -10 to 10 -13  g/μl) more sensitive than PCR (10 -10 to 10 -12  g/μl). The kappa value between LAMP and PCR indicated a faint concurrence (0.463). The kappa coefficient between LAMP and flotation technique indicated a strong agreement (0.667). The highest infection rate (n = 11) was detected in non-dewormed pet dogs, particularly those less than 3 months old (P canis eggs in infected pet dogs. It was proposed that the dog holder's awareness is insufficient to implement regular deworming schedules. Additionally, regional policymakers should broadly revise anthelmintic treatment guidelines.

Automatic Lamp and Fan Control Based on Microcontroller

Science.gov (United States)

Widyaningrum, V. T.; Pramudita, Y. D.

2018-01-01

In general, automation can be described as a process following pre-determined sequential steps with a little or without any human exertion. Automation is provided with the use of various sensors suitable to observe the production processes, actuators and different techniques and devices. In this research, the automation system developed is an automatic lamp and an automatic fan on the smart home. Both of these systems will be processed using an Arduino Mega 2560 microcontroller. A microcontroller is used to obtain values of physical conditions through sensors connected to it. In the automatic lamp system required sensors to detect the light of the LDR (Light Dependent Resistor) sensor. While the automatic fan system required sensors to detect the temperature of the DHT11 sensor. In tests that have been done lamps and fans can work properly. The lamp can turn on automatically when the light begins to darken, and the lamp can also turn off automatically when the light begins to bright again. In addition, it can concluded also that the readings of LDR sensors are placed outside the room is different from the readings of LDR sensors placed in the room. This is because the light intensity received by the existing LDR sensor in the room is blocked by the wall of the house or by other objects. Then for the fan, it can also turn on automatically when the temperature is greater than 25°C, and the fan speed can also be adjusted. The fan may also turn off automatically when the temperature is less than equal to 25°C.

Raman lidar characterization using a reference lamp

Science.gov (United States)

Landulfo, Eduardo; da Costa, Renata F.; Rodrigues, Patricia F.; da Silva Lopes, Fábio J.

2014-10-01

The determination of the amount of water vapor in the atmosphere using lidar is a calibration dependent technique. Different collocated instruments are used for this purpose, like radiossoundings and microwave radiometers. When there are no collocated instruments available, an independente lamp mapping calibration technique can be used. Aiming to stabilish an independ technique for the calibration of the six channels Nd-YAG Raman lidar system located at the Center for Lasers and Applications (CLA), S˜ao Paulo, Brazil, an optical characterization of the system was first performed using a reference tungsten lamp. This characterization is useful to identify any possible distortions in the interference filters, telescope mirror and stray light contamination. In this paper we show three lamp mapping caracterizations (01/16/2014, 01/22/2014, 04/09/2014). The first day is used to demostrate how the tecnique is useful to detect stray light, the second one how it is sensible to the position of the filters and the third one demostrates a well optimized optical system.

Analysis of compact and portable goniospectrometer system for test of LED lamps

DEFF Research Database (Denmark)

Dam-Hansen, Carsten; Amdemeskel, Mekbib Wubishet; Thorseth, Anders

2015-01-01

measurements in a near-field goniophotometer. A collection of six different types of directional and non-directional integrated LED lamps with three samples of each were used as test devices. It is shown that the main uncertainty comes from the inadequate thermal stabilisation of the LED lamps. With pre...

Utility of loop-mediated isothermal amplification assay, polymerase chain reaction, and elisa for diagnosis of leptospirosis in South Indian patients

Directory of Open Access Journals (Sweden)

Mallika Sengupta

2017-01-01

Full Text Available Background: Leptospirosis is a zoonotic disease which requires laboratory diagnosis for confirmation. Materials and Methods: In this study serum samples from adults with acute undifferentiated fever (duration ≤15 days were tested for IgM antibodies to Leptospira by ELISA, PCR for rrs gene and loop-mediated isothermal amplification (LAMP assay for LipL32 and LipL41. Results: Among the 150 sera tested, three were positive by PCR, LAMP and IgM ELISA/modified Faines' criteria, two by only PCR; seven only by LAMP assay and forty fulfilled modified Faine's criteria (illness clinically compatible and IgM ELISA positive for leptospirosis. Clinical correlation revealed renal compromise, low platelet count and severe jaundice were significantly related to leptospirosis (P < 0.05. Conclusion: This study suggests that LAMP assay could be useful for diagnosis of leptospirosis during the 1st week of illness whereas IgM ELISA forms the mainstay of diagnosis from the 2nd week onward. Further studies especially community based, comparing ELISA, PCR, LAMP, culture and microscopic agglutination test are required to evaluate the veracity of these findings.

Application Summary Report 22: LED MR16 Lamps

Energy Technology Data Exchange (ETDEWEB)

Royer, Michael P.

2014-07-23

, or the approximate equivalent of a 50 W halogen MR16 lamp. Many factors beyond photometric performance should be considered during specification. For example, performance over time, transformer and dimmer compatibility, and total system performance are all critical to a successful installation. Subsequent CALiPER reports will investigate more complex issues.

Accuracy of loop-mediated isothermal amplification for diagnosis of human leptospirosis in Thailand.

Science.gov (United States)

Sonthayanon, Piengchan; Chierakul, Wirongrong; Wuthiekanun, Vanaporn; Thaipadungpanit, Janjira; Kalambaheti, Thareerat; Boonsilp, Siriphan; Amornchai, Premjit; Smythe, Lee D; Limmathurotsakul, Direk; Day, Nicholas P; Peacock, Sharon J

2011-04-01

There is a lack of diagnostic tests for leptospirosis in technology-restricted settings. We developed loop-mediated isothermal amplification (LAMP) specific for the 16S ribosomal RNA gene (rrs) of pathogenic and intermediate group Leptospira species. The lower limit of detection was 10 genomic equivalents/reaction, and analytical specificity was high; we observed positive reactions for pathogenic/intermediate groups and negative reactions for non-pathogenic Leptospira species and other bacterial species. We evaluated this assay in Thailand by using a case-control study of 133 patients with laboratory-proven leptospirosis and 133 patients with other febrile illnesses. Using admission blood, we found that the rrs LAMP showed positive results in 58 of 133 cases (diagnostic sensitivity = 43.6, 95% confidence interval [CI] = 35.0-52.5) and in 22 of 133 controls (diagnostic specificity = 83.5, 95% CI = 76.0-89.3). Sensitivity was high for 39 patients who were culture positive for Leptospira spp. (84.6, 95% CI = 69.5-94.1). The rrs LAMP can provide an admission diagnosis in approximately half of patients with leptospirosis, but its clinical utility is reduced by a lower specificity.

Marker lamps

International Nuclear Information System (INIS)

Watkins, D.V.

1980-01-01

A marker lamp is described which consists of a block of transparent plastics material encapsulated in which is a radioactive light source. These lights comprise a small sealed glass capsule, the hollow inside surface of which is coated with phosphor and which contains tritium or similar radioactive gas. The use of such lamps for identification marking of routes, for example roads, and for identification of underwater oil pipelines is envisaged. (U.K.)

An ion quencher operated lamp for multiplexed fluorescent bioassays.

Science.gov (United States)

Qing, Taiping; Sun, Huanhuan; He, Xiaoxiao; Huang, Xiaoqin; He, Dinggeng; Bu, Hongchang; Qiao, Zhenzhen; Wang, Kemin

2018-02-01

A novel and adjustable lamp based on competitive interaction among dsDNA-SYBR Green I (SGI), ion quencher, and analyte was designed for bioanalysis. The "filament" and switch of the lamp could be customized by employing different dsDNA and ion quencher. The poly(AT/TA) dsDNA was successfully screened as the most effective filament of the lamp. Two common ions, Hg 2+ and Fe 3+ , were selected as the model switch, and the corresponding ligand molecules cysteine (Cys) and pyrophosphate ions (PPi) were selected as the targets. When the fluorescence-quenched dsDNA/SGI-ion complex was introduced into a target-containing system, ions could be bound by competitive molecules and separate from the complex, thereby lighting the lamp. However, no light was observed if the biomolecule could not snatch the metal ions from the complex. Under the optimal conditions, sensitive and selective detection of Cys and PPi was achieved by the lamp, with practical applications in fetal bovine serum and human urine. This ion quencher regulated lamp for fluorescent bioassays is simple in design, fast in operation, and is more convenient than other methods. Significantly, as many molecules could form stable complexes with metal ions selectively, this ion quencher operated lamp has potential for the detection of a wide spectrum of analytes. Graphical abstract A novel and adjustable lamp on the basis of competitive interaction among dsDNA-SYBR Green I, ions quencher and analyte was designed for bioanalysis. The filament and switch of lamp could be customized by employing different dsDNA and ions quencher.

CALiPER Report 22.1: Photoelectric Performance of LED MR16 Lamps

Energy Technology Data Exchange (ETDEWEB)

None, None

2015-08-31

This report looks at the photoelectric performance of the same set of lamps assessed in Report 22, using commercially available transformers and dimmers as well as laboratory power supplies providing either AC or DC. The investigation explores several issues related to the testing and use of MR16 lamps in lighting systems and examines the range of performance that is possible for a given lamp model, based on the system to which it is connected.

Transparent ceramic lamp envelope materials

Energy Technology Data Exchange (ETDEWEB)

Wei, G C [OSRAM SYLVANIA, 71 Cherry Hill Drive, Beverly, MA 01915 (United States)

2005-09-07

Transparent ceramic materials with optical qualities comparable to single crystals of similar compositions have been developed in recent years, as a result of the improved understanding of powder-processing-fabrication- sintering-property inter-relationships. These high-temperature materials with a range of thermal and mechanical properties are candidate envelopes for focused-beam, short-arc lamps containing various fills operating at temperatures higher than quartz. This paper reviews the composition, structure and properties of transparent ceramic lamp envelope materials including sapphire, small-grained polycrystalline alumina, aluminium oxynitride, yttrium aluminate garnet, magnesium aluminate spinel and yttria-lanthana. A satisfactory thermal shock resistance is required for the ceramic tube to withstand the rapid heating and cooling cycles encountered in lamps. Thermophysical properties, along with the geometry, size and thickness of a transparent ceramic tube, are important parameters in the assessment of its resistance to fracture arising from thermal stresses in lamps during service. The corrosive nature of lamp-fill liquid and vapour at high temperatures requires that all lamp components be carefully chosen to meet the target life. The wide range of new transparent ceramics represents flexibility in pushing the limit of envelope materials for improved beamer lamps.

Intelligent software systems and SMiRT: Potentials, actual results, expectations, trends

International Nuclear Information System (INIS)

Jovanovic, A.

1993-01-01

The paper gives a survey of recent development trends in the area of knowledge-based systems, hypermedia, neural networks and other similar technologies which are on the baseline of modern 'intelligent' software systems, applied in the areas relevant for SMiRT: power plant operation, design and analysis of structural components, materials, and many others. The paper highlights the historical background of these trends, as well as the methodologies and technologies which made such a development possible ('enabling methodologies/technologies'). Examples from several, SMiRT characteristic application areas are mentioned, in order to give an illustration what the deployment of an intelligent software system can mean in practice. Finally, a summary of these result is made and future perspectives indicated. (author)

A PCA3 gene-based transcriptional amplification system targeting primary prostate cancer

OpenAIRE

Neveu, Bertrand; Jain, Pallavi; T?tu, Bernard; Wu, Lily; Fradet, Yves; Pouliot, Fr?d?ric

2015-01-01

Targeting specifically primary prostate cancer (PCa) cells for immune therapy, gene therapy or molecular imaging is of high importance. The PCA3 long non-coding RNA is a unique PCa biomarker and oncogene that has been widely studied. This gene has been mainly exploited as an accurate diagnostic urine biomarker for PCa detection. In this study, the PCA3 promoter was introduced into a new transcriptional amplification system named the 3-Step Transcriptional Amplification System (PCA3-3STA) and ...

On the square arc voltage waveform model in magnetic discharge lamp studies

OpenAIRE

Molina, Julio; Sainz Sapera, Luis; Mesas García, Juan José

2011-01-01

The current number of magnetic and electronic ballast discharge lamps in power distribution systems is increasing because they perform better than incandescent lamps. This paper studies the magnetic discharge lamp modeling. In particular, the arc voltage waveform is analyzed and the limitations of the square waveform model are revealed from experimental measurements.

Design of LED lamps | Ashryatov | Journal of Fundamental and ...

African Journals Online (AJOL)

Design of LED lamps. ... In this paper, we study the effect of LED high brightness on the brightness of a luminaire. The nomenclature of diffusers used in the production of ... The variant of the lighting system energy efficiency increase with luminaires and linear fluorescent lamps is considered. In the proposed variant, the ...

DNA detection of Schistosoma japonicum: diagnostic validity of a LAMP assay for low-intensity infection and effects of chemotherapy in humans.

Science.gov (United States)

Xu, Jing; Guan, Zhi-Xun; Zhao, Bo; Wang, Yan-Yan; Cao, Yun; Zhang, Hui-Qin; Zhu, Xing-Quan; He, Yong-Kang; Xia, Chao-Ming

2015-04-01

Schistosomiasis has decreased significantly in prevalence and intensity of infection in China, thus more accurate and sensitive methods are desperately needed for the further control of schistosomiasis. The present work aimed to assess the utility of the loop-mediated isothermal amplification (LAMP) for detection of light intensity infection or false-negative patients and patients post-treatment, targeting the highly repetitive retrotransposon SjR2 of Schistosoma japonicum. LAMP was first assessed in rabbits with low intensity infection (EPGLAMP. Meanwhile, 42 sera from healthy individuals in a non-endemic area, and 60 sera from "healthy" residents who were identified as being negative for feces examination and immuno-methods in an endemic area were also examined. The results showed that LAMP could detect S. japonicum DNA in sera from rabbits at 3rd day post-infection. Following administration of praziquantel, the S. japonicum DNA in rabbit sera became negative at 10 weeks post-treatment. Of 110 sera from patients, LAMP showed 95.5% sensitivity, and even for 41 patients with less than 10 EPG, the sensitivity of LAMP still reached to 95.1%. For 47 patients after treatment, the negative conversion rate of S. japonicum DNA in patient sera increased from 23.4%, 61.7% to 83.0% at 3 months, 6 months and 9 months post-treatment, respectively. No false-positive result was obtained for 42 human sera from non-endemic area, while for the 60 "healthy" individuals from endemic area, 10 (16.7%) individuals were positive by LAMP, which suggested that these individuals might be false-negative patients. The present study demonstrated that the LAMP assay is sensitive, specific, and affordable, which would help reduce schistosomiasis transmission through targeted treatment of individuals, particularly for those with negative stool examinations who may yet remain infected. The LAMP assay may provide a potential tool to support schistosomiasis control and elimination strategies.

Use of Malachite Green-Loop Mediated Isothermal Amplification for Detection of Plasmodium spp. Parasites

Science.gov (United States)

Lucchi, Naomi W.; Ljolje, Dragan; Silva-Flannery, Luciana; Udhayakumar, Venkatachalam

2016-01-01

Malaria elimination efforts are hampered by the lack of sensitive tools to detect infections with low-level parasitemia, usually below the threshold of standard diagnostic methods, microscopy and rapid diagnostic tests. Isothermal nucleic acid amplification assays such as the loop-mediated isothermal amplification (LAMP), are well suited for field use as they do not require thermal cyclers to run the test. However, the use of specialized equipment, as described by many groups, reduces the versatility of the LAMP technique as a simple tool for use in endemic countries. In this study, the use of the malachite green (MG) dye, as a visual endpoint readout, together with a simple mini heat block was evaluated for the detection of malaria parasites. The assay was performed for 1 hour at 63°C and the results scored by 3 independent human readers. The limit of detection of the assay was determined using well-quantified Plasmodium spp. infected reference samples and its utility in testing clinical samples was determined using 190 pre-treatment specimens submitted for reference diagnosis of imported malaria in the United States. Use of a simplified boil and spin methods of DNA extraction from whole blood and filter paper was also investigated. We demonstrate the accurate and sensitive detection of malaria parasites using this assay with a detection limit ranging between 1–8 parasites/μL, supporting its applicability for the detection of infections with low parasite burden. This assay is compatible with the use of a simple boil and spin sample preparation method from both whole blood and filter papers without a loss of sensitivity. The MG-LAMP assay described here has great potential to extend the reach of molecular tools to settings where they are needed. PMID:26967908

Use of Malachite Green-Loop Mediated Isothermal Amplification for Detection of Plasmodium spp. Parasites.

Directory of Open Access Journals (Sweden)

Naomi W Lucchi

Full Text Available Malaria elimination efforts are hampered by the lack of sensitive tools to detect infections with low-level parasitemia, usually below the threshold of standard diagnostic methods, microscopy and rapid diagnostic tests. Isothermal nucleic acid amplification assays such as the loop-mediated isothermal amplification (LAMP, are well suited for field use as they do not require thermal cyclers to run the test. However, the use of specialized equipment, as described by many groups, reduces the versatility of the LAMP technique as a simple tool for use in endemic countries. In this study, the use of the malachite green (MG dye, as a visual endpoint readout, together with a simple mini heat block was evaluated for the detection of malaria parasites. The assay was performed for 1 hour at 63°C and the results scored by 3 independent human readers. The limit of detection of the assay was determined using well-quantified Plasmodium spp. infected reference samples and its utility in testing clinical samples was determined using 190 pre-treatment specimens submitted for reference diagnosis of imported malaria in the United States. Use of a simplified boil and spin methods of DNA extraction from whole blood and filter paper was also investigated. We demonstrate the accurate and sensitive detection of malaria parasites using this assay with a detection limit ranging between 1-8 parasites/μL, supporting its applicability for the detection of infections with low parasite burden. This assay is compatible with the use of a simple boil and spin sample preparation method from both whole blood and filter papers without a loss of sensitivity. The MG-LAMP assay described here has great potential to extend the reach of molecular tools to settings where they are needed.

Detection of Panton-Valentine Leukocidin DNA from methicillin-resistant Staphylococcus aureus by resistive pulse sensing and loop-mediated isothermal amplification with gold nanoparticles

International Nuclear Information System (INIS)

Yang, Alice Kar Lai; Lu, Haifei; Wu, Shu Yuen; Kwok, Ho Chin; Ho, Ho Pui; Yu, Samuel; Cheung, Anthony Ka Lun; Kong, Siu Kai

2013-01-01

Graphical abstract: -- Highlights: •A novel diagnostic assay is developed to detect the MRSA's Panton-Valentine Leukocidin toxin. •Detection is based on target DNA amplification at one single temperature at 65 °C by LAMP. •Amplicons are then hybridized with 2 Au-nanoparticles with specific DNA probes for sensing. •The supra-assemblies are subsequently sensed by resistive pulse sensing. •Detection limit: ∼200 copies of DNA; time for detection: completed within 2 h. -- Abstract: This report describes a novel diagnostic assay for rapid detection of the Panton-Valentine Leukocidin (PVL) toxin of methicillin-resistant Staphylococcus aureus (MRSA) utilizing resistive pulse sensing (RPS), loop-mediated isothermal DNA amplification (LAMP) in combination with gold nanoparticles (AuNPs). The PVL DNA from MRSA was specifically amplified by LAMP using four primers at one temperature (65 °C). The DNA products with biotin were then conjugated to a first AuNP1 (55 ± 2 nm) through biotin–avidin binding. A second AuNP2 (30 ± 1.5 nm) coated with a specific DNA probe hybridized with the LAMP DNA products at the loop region to enhance assay sensitivity and specificity, to generate supra-AuNP1-DNA-AuNP2 assemblies. Scanning electron microscopy confirmed the presence of these supra-assemblies. Using RPS, detection and quantitation of the agglomerated AuNPs were performed by a tunable fluidic nanopore sensor. The results demonstrate that the LAMP-based RPS sensor is sensitive and rapid for detecting the PVL DNA. This technique could achieve a limit of detection (LOD) up to about 500 copies of genomic DNA from the bacteria MRSA MW2 and the detection can be completed within two hours with a straightforward signal-to-readout setup. It is anticipated that this LAMP-based AuNP RPS may become an effective tool for MRSA detection and a potential platform in clinical laboratory to report the presence or absence of other types of infectious agents

Detection of Tomato black ring virus by real-time one-step RT-PCR.

Science.gov (United States)

Harper, Scott J; Delmiglio, Catia; Ward, Lisa I; Clover, Gerard R G

2011-01-01

A TaqMan-based real-time one-step RT-PCR assay was developed for the rapid detection of Tomato black ring virus (TBRV), a significant plant pathogen which infects a wide range of economically important crops. Primers and a probe were designed against existing genomic sequences to amplify a 72 bp fragment from RNA-2. The assay amplified all isolates of TBRV tested, but no amplification was observed from the RNA of other nepovirus species or healthy host plants. The detection limit of the assay was estimated to be around nine copies of the TBRV target region in total RNA. A comparison with conventional RT-PCR and ELISA, indicated that ELISA, the current standard test method, lacked specificity and reacted to all nepovirus species tested, while conventional RT-PCR was approximately ten-fold less sensitive than the real-time RT-PCR assay. Finally, the real-time RT-PCR assay was tested using five different RT-PCR reagent kits and was found to be robust and reliable, with no significant differences in sensitivity being found. The development of this rapid assay should aid in quarantine and post-border surveys for regulatory agencies. Copyright © 2010 Elsevier B.V. All rights reserved.

A new Cassegrain calibration lamp unit for the Blanco Telescope

Science.gov (United States)

Points, S. D.; James, D. J.; Tighe, R.; Montané, A.; David, N.; Martínez, M.

2016-08-01

The f/8 RC-Cassegrain Focus of the Blanco Telescope at Cerro Tololo Inter-American Observatory, hosts two new instruments: COSMOS, a multi-object spectrograph in the visible wavelength range (350 - 1030nm), and ARCoIRIS, a NIR cross-dispersed spectrograph featuring 6 spectral orders spanning 0.8 - 2.45μm. Here we describe a calibration lamp unit designed to deliver the required illumination at the telescope focal plane for both instruments. These requirements are: (1) an f/8 beam of light covering a spot of 92mm diameter (or 10 arcmin) for a wavelength range of 0.35μm through 2.5μm and (2) no saturation of flat-field calibrations for the minimal exposure times permitted by each instrument, and (3) few saturated spectral lines when using the wavelength calibration lamps for the instruments. To meet these requirements this unit contains an adjustable quartz halogen lamp for flat-field calibrations, and one hollow cathode lamp and four penray lamps for wavelength calibrations. The wavelength calibration lamps are selected to provide optimal spectral coverage for the instrument mounted and can be used individually or in sets. The device designed is based on an 8-inch diameter integrating sphere, the output of which is optimized to match the f/8 calibration input delivery system which is a refractive system based on fused-silica lenses. We describe the optical design, the opto-mechanical design, the electronic control and give results of the performance of the system.

Optimisation of the RT-PCR detection of immunomagnetically enriched carcinoma cells

International Nuclear Information System (INIS)

Raynor, Michael; Stephenson, Sally-Anne; Walsh, David CA; Pittman, Kenneth B; Dobrovic, Alexander

2002-01-01

Immunomagnetic enrichment followed by RT-PCR (immunobead RT-PCR) is an efficient methodology to identify disseminated carcinoma cells in the blood and bone marrow. The RT-PCR assays must be both specific for the tumor cells and sufficiently sensitive to enable detection of single tumor cells. We have developed a method to test RT-PCR assays for any cancer. This has been investigated using a panel of RT-PCR markers suitable for the detection of breast cancer cells. In the assay, a single cell line-derived tumor cell is added to 100 peripheral blood mononuclear cells (PBMNCs) after which mRNA is isolated and reverse transcribed for RT-PCR analysis. PBMNCs without added tumor cells are used as specificity controls. The previously studied markers epidermal growth factor receptor (EGFR), mammaglobin 1 (MGB1), epithelial cell adhesion molecule (EpCAM/TACSTD1), mucin 1 (MUC1), carcinoembryonic antigen (CEA) were tested. Two new epithelial-specific markers ELF3 and EphB4 were also tested. MUC1 was unsuitable as strong amplification was detected in 100 cell PBMNC controls. Expression of ELF3, EphB4, EpCAM, EGFR, CEA and MGB1 was found to be both specific for the tumor cell, as demonstrated by the absence of a signal in most 100 cell PBMNC controls, and sensitive enough to detect a single tumor cell in 100 PBMNCs using a single round of RT-PCR. ELF3, EphB4, EpCAM, EGFR, CEA and MGB1 are appropriate RT-PCR markers for use in a marker panel to detect disseminated breast cancer cells after immunomagnetic enrichment

Feasibility of small animal cranial irradiation with the microRT system

International Nuclear Information System (INIS)

Kiehl, Erich L.; Stojadinovic, Strahinja; Malinowski, Kathleen T.; Limbrick, David; Jost, Sarah C.; Garbow, Joel R.; Rubin, Joshua B.; Deasy, Joseph O.; Khullar, Divya; Izaguirre, Enrique W.; Parikh, Parag J.; Low, Daniel A.; Hope, Andrew J.

2008-01-01

Purpose: To develop and validate methods for small-animal CNS radiotherapy using the microRT system. Materials and Methods: A custom head immobilizer was designed and built to integrate with a pre-existing microRT animal couch. The Delrin couch-immobilizer assembly, compatible with multiple imaging modalities (CT, microCT, microMR, microPET, microSPECT, optical), was first imaged via CT in order to verify the safety and reproducibility of the immobilization method. Once verified, the subject animals were CT-scanned while positioned within the couch-immobilizer assembly for treatment planning purposes. The resultant images were then imported into CERR, an in-house-developed research treatment planning system, and registered to the microRTP treatment planning space using rigid registration. The targeted brain was then contoured and conformal radiotherapy plans were constructed for two separate studies: (1) a whole-brain irradiation comprised of two lateral beams at the 90 degree sign and 270 degree sign microRT treatment positions and (2) a hemispheric (left-brain) irradiation comprised of a single A-P vertex beam at the 0 degree sign microRT treatment position. During treatment, subject animals (n=48) were positioned to the CERR-generated treatment coordinates using the three-axis microRT motor positioning system and were irradiated using a clinical Ir-192 high-dose-rate remote after-loading system. The radiation treatment course consisted of 5 Gy fractions, 3 days per week. 90% of the subjects received a total dose of 30 Gy and 10% received a dose of 60 Gy. Results: Image analysis verified the safety and reproducibility of the immobilizer. CT scans generated from repeated reloading and repositioning of the same subject animal in the couch-immobilizer assembly were fused to a baseline CT. The resultant analysis revealed a 0.09 mm average, center-of-mass translocation and negligible volumetric error in the contoured, murine brain. The experimental use of the head

Design and testing of a separate-type lighting system using solar energy and cold-cathode fluorescent lamps

Energy Technology Data Exchange (ETDEWEB)

Yang, J.-P. [Department of Electrical Engineering, China Institute of Technology, Taipei, Taiwan 115, Taiwan (China)]. E-mail: april4120@tp.edu.tw; Hsiao, H.-C. [Department of Electrical Engineering, National Taiwan University of Science and Technology, Taipei, Taiwan 106, Taiwan (China)]. E-mail: hsiao@mouse.ee.ntust.edu.tw

2007-01-15

This paper presents a solar-powered lighting system, using cold-cathode fluorescent-lamps (CCFLs), with its battery-charging circuit and lamp-ignition circuit being separated so that its solar panels can be installed at any distance deemed necessary away from the lighting site in order to receive the maximum solar energy available. This system adopts the maximum-power point tracking (MPPT) method to control the power output of the solar panels and uses the zero-voltage switching (ZVS) DC-DC converter, as the charging circuit, to increase the panels' power generation efficiency and the charging circuit's conversion efficiency. The electronic ballast circuit for the CCFL is constructed with a half-bridge inverter, a resonant inductor, and a Rosen-type piezoelectric transformer, which forms a piezoelectric resonant-type inverter: to simplify the circuitry and to improve the power conversion efficiency, the ballast circuit is designed to directly step up the battery voltage in igniting the lamp. We also establish the transmission-parameter model for the piezoelectric resonant-type inverter to provide the base for the electric-power circuit design. Our experimental results indicate that the proposed system possesses some advantages, such as greater energy efficiency, circuitry simplicity, and so on, and is suitable for night lighting in house yards, parks and advertising panels.

Design and testing of a separate-type lighting system using solar energy and cold-cathode fluorescent lamps

International Nuclear Information System (INIS)

Yang, J.-P.; Hsiao, H.-C.

2007-01-01

This paper presents a solar-powered lighting system, using cold-cathode fluorescent-lamps (CCFLs), with its battery-charging circuit and lamp-ignition circuit being separated so that its solar panels can be installed at any distance deemed necessary away from the lighting site in order to receive the maximum solar energy available. This system adopts the maximum-power point tracking (MPPT) method to control the power output of the solar panels and uses the zero-voltage switching (ZVS) DC-DC converter, as the charging circuit, to increase the panels' power generation efficiency and the charging circuit's conversion efficiency. The electronic ballast circuit for the CCFL is constructed with a half-bridge inverter, a resonant inductor, and a Rosen-type piezoelectric transformer, which forms a piezoelectric resonant-type inverter: to simplify the circuitry and to improve the power conversion efficiency, the ballast circuit is designed to directly step up the battery voltage in igniting the lamp. We also establish the transmission-parameter model for the piezoelectric resonant-type inverter to provide the base for the electric-power circuit design. Our experimental results indicate that the proposed system possesses some advantages, such as greater energy efficiency, circuitry simplicity, and so on, and is suitable for night lighting in house yards, parks and advertising panels

Modeling qRT-PCR dynamics with application to cancer biomarker quantification.

Science.gov (United States)

Chervoneva, Inna; Freydin, Boris; Hyslop, Terry; Waldman, Scott A

2017-01-01

Quantitative reverse transcription polymerase chain reaction (qRT-PCR) is widely used for molecular diagnostics and evaluating prognosis in cancer. The utility of mRNA expression biomarkers relies heavily on the accuracy and precision of quantification, which is still challenging for low abundance transcripts. The critical step for quantification is accurate estimation of efficiency needed for computing a relative qRT-PCR expression. We propose a new approach to estimating qRT-PCR efficiency based on modeling dynamics of polymerase chain reaction amplification. In contrast, only models for fluorescence intensity as a function of polymerase chain reaction cycle have been used so far for quantification. The dynamics of qRT-PCR efficiency is modeled using an ordinary differential equation model, and the fitted ordinary differential equation model is used to obtain effective polymerase chain reaction efficiency estimates needed for efficiency-adjusted quantification. The proposed new qRT-PCR efficiency estimates were used to quantify GUCY2C (Guanylate Cyclase 2C) mRNA expression in the blood of colorectal cancer patients. Time to recurrence and GUCY2C expression ratios were analyzed in a joint model for survival and longitudinal outcomes. The joint model with GUCY2C quantified using the proposed polymerase chain reaction efficiency estimates provided clinically meaningful results for association between time to recurrence and longitudinal trends in GUCY2C expression.

Wood's lamp illumination (image)

Science.gov (United States)

A Wood's lamp emits ultraviolet light and can be a diagnostic aid in determining if someone has a fungal ... is an infection on the area where the Wood's lamp is illuminating, the area will fluoresce. Normally ...

Design of Solar Street Lamp Control System Based on MPPT

Science.gov (United States)

Cui, Fengying

This paper proposes a new solar street lamp control system which is composed of photovoltaic cell, controller, battery and load. In this system controller as the key part applies the microchip to achieve many functions. According to the nonlinear output characteristics of solar cell and the influence of environment, it uses the perturbation and observation (P&O) method to realize the maximum power point tracking (MPPT) and promotes the efficiency. In order to prolong the battery life the pulse width modulation (PWM) charge mode is selected to control the battery capacity and provent the battery from the state of over-charge and over-discharge. Meanwhile the function of temperature compensation, charge and discharge protection are set to improve the running safety and stability.

Impact of asymmetric lamp positioning on the performance of a closed-conduit UV reactor

Directory of Open Access Journals (Sweden)

Tipu Sultan

2017-06-01

Full Text Available Computational fluid dynamics (CFD analyses for the performance improvement of a closed-conduit ultraviolet (UV reactor were performed by changing the lamp positions from symmetric to asymmetric. The asymmetric lamp positioning can be useful for UV reactor design and optimization. This goal was achieved by incorporating the two performance factors, namely reduction equivalent dose (RED and system dose performance. Four cases were carried out for asymmetric lamp positioning within the UV reactor chamber and each case consisted of four UV lamps that were simulated once symmetrically and four times asymmetrically. The results of the four asymmetric cases were compared with the symmetric one. Moreover, these results were evaluated by using CFD simulations of a closed-conduit UV reactor. The fluence rate model, UVCalc3D was employed to validate the simulations results. The simulation results provide detailed information about the dose distribution, pathogen track modeling and RED. The RED value was increased by approximately 15% by using UVCalc3D fluence rate model. Additionally, the asymmetric lamp positioning of the UV lamps had more than 50% of the pathogens received a better and a higher UV dose than in the symmetric case. Consequently, the system dose performance was improved by asymmetric lamp positioning. It was concluded that the performance parameters (higher RED and system dose performance were improved by using asymmetric lamp positioning.

Lysosome-Associated Membrane Proteins (LAMP Maintain Pancreatic Acinar Cell Homeostasis: LAMP-2–Deficient Mice Develop PancreatitisSummary

Directory of Open Access Journals (Sweden)

Olga A. Mareninova

2015-11-01

Full Text Available Background & Aims: The pathogenic mechanism of pancreatitis is poorly understood. Recent evidence implicates defective autophagy in pancreatitis responses; however, the pathways mediating impaired autophagy in pancreas remain largely unknown. Here, we investigate the role of lysosome associated membrane proteins (LAMPs in pancreatitis. Methods: We analyzed changes in LAMPs in experimental models and human pancreatitis, and the underlying mechanisms: LAMP deglycosylation and degradation. LAMP cleavage by cathepsin B (CatB was analyzed by mass spectrometry. We used mice deficient in LAMP-2 to assess its role in pancreatitis. Results: Pancreatic levels of LAMP-1 and LAMP-2 greatly decrease across various pancreatitis models and in human disease. Pancreatitis does not trigger the LAMPs’ bulk deglycosylation but induces their degradation via CatB-mediated cleavage of the LAMP molecule close to the boundary between luminal and transmembrane domains. LAMP-2 null mice spontaneously develop pancreatitis that begins with acinar cell vacuolization due to impaired autophagic flux, and progresses to severe pancreas damage characterized by trypsinogen activation, macrophage-driven inflammation, and acinar cell death. LAMP-2 deficiency causes a decrease in pancreatic digestive enzymes content, and stimulates the basal and inhibits cholecystokinin-induced amylase secretion by acinar cells. The effects of LAMP-2 knockout and acute cerulein pancreatitis overlap, which corroborates the pathogenic role of LAMP decrease in experimental pancreatitis models. Conclusions: The results indicate a critical role for LAMPs, particularly LAMP-2, in maintaining pancreatic acinar cell homeostasis and provide evidence that defective lysosomal function, resulting in impaired autophagy, leads to pancreatitis. Mice with LAMP-2 deficiency present a novel genetic model of human pancreatitis caused by lysosomal/autophagic dysfunction. Keywords: Amylase Secretion, Autophagy

Real-Time Fluorescence Loop Mediated Isothermal Amplification for the Detection of Acinetobacter baumannii

Science.gov (United States)

Wang, Qinqin; Zhou, Yanbin; Li, Shaoli; Zhuo, Chao; Xu, Siqi; Huang, Lixia; Yang, Ling; Liao, Kang

2013-01-01

Background Detection of Acinetobacter baumannii has been relying primarily on bacterial culture that often fails to return useful results in time. Although DNA-based assays are more sensitive than bacterial culture in detecting the pathogen, the molecular results are often inconsistent and challenged by doubts on false positives, such as those due to system- and environment-derived contaminations. In addition, these molecular tools require expensive laboratory instruments. Therefore, establishing molecular tools for field use require simpler molecular platforms. The loop-mediated isothermal amplification method is relatively simple and can be improved for better use in a routine clinical bacteriology laboratory. A simple and portable device capable of performing both the amplification and detection (by fluorescence) of LAMP in the same platform has been developed in recent years. This method is referred to as real-time loop-mediated isothermal amplification. In this study, we attempted to utilize this method for rapid detection of A. baumannii. Methodology and Significant Findings Species-specific primers were designed to test the utility of this method. Clinical samples of A. baumannii were used to determine the sensitivity and specificity of this system compared to bacterial culture and a polymerase chain reaction method. All positive samples isolated from sputum were confirmed to be the species of Acinetobacter by 16S rRNA gene sequencing. The RealAmp method was found to be simpler and allowed real-time detection of DNA amplification, and could distinguish A. baumannii from Acinetobacter calcoaceticus and Acinetobacter genomic species 3. DNA was extracted by simple boiling method. Compared to bacterial culture, the sensitivity and specificity of RealAmp in detecting A. baumannii was 98.9% and 75.0%, respectively. Conclusion The RealAmp assay only requires a single unit, and the assay positivity can be verified by visual inspection. Therefore, this assay has

DNA detection of Schistosoma japonicum: diagnostic validity of a LAMP assay for low-intensity infection and effects of chemotherapy in humans.

Directory of Open Access Journals (Sweden)

Jing Xu

2015-04-01

Full Text Available Schistosomiasis has decreased significantly in prevalence and intensity of infection in China, thus more accurate and sensitive methods are desperately needed for the further control of schistosomiasis. The present work aimed to assess the utility of the loop-mediated isothermal amplification (LAMP for detection of light intensity infection or false-negative patients and patients post-treatment, targeting the highly repetitive retrotransposon SjR2 of Schistosoma japonicum.LAMP was first assessed in rabbits with low intensity infection (EPG<10. Then 110 patient sera from Hunan Province, China, and 47 sera after treatment by praziquantel were used to evaluate the diagnostic validity of LAMP. Meanwhile, 42 sera from healthy individuals in a non-endemic area, and 60 sera from "healthy" residents who were identified as being negative for feces examination and immuno-methods in an endemic area were also examined. The results showed that LAMP could detect S. japonicum DNA in sera from rabbits at 3rd day post-infection. Following administration of praziquantel, the S. japonicum DNA in rabbit sera became negative at 10 weeks post-treatment. Of 110 sera from patients, LAMP showed 95.5% sensitivity, and even for 41 patients with less than 10 EPG, the sensitivity of LAMP still reached to 95.1%. For 47 patients after treatment, the negative conversion rate of S. japonicum DNA in patient sera increased from 23.4%, 61.7% to 83.0% at 3 months, 6 months and 9 months post-treatment, respectively. No false-positive result was obtained for 42 human sera from non-endemic area, while for the 60 "healthy" individuals from endemic area, 10 (16.7% individuals were positive by LAMP, which suggested that these individuals might be false-negative patients.The present study demonstrated that the LAMP assay is sensitive, specific, and affordable, which would help reduce schistosomiasis transmission through targeted treatment of individuals, particularly for those with

The Effect of Operating Lamps on the Protected Area of a Unidirectional Down Flow (UDF) System.

Science.gov (United States)

Traversari, A A L; Bottenheft, C; Louman, R; van Heumen, S P M; Böggemann, J

2017-04-01

Operating lamps are often seen as the most disruptive factors within the protective area in the operating theater (OT). The effect of the operation lamps (with different shapes) should be demonstrated in an OT by trial, since research on the effects of the lamps is still limited. The main aim of this study was to determine the effects of a skirt, different lamps, and the position of the lamp on the protected area. The concentration of airborne particles was measured under different circumstances, in order to determine the size and quality of the protected area. This entrainment/segregation test is based on the deliberate and controlled emission of particles outside the zone that is protected. The degree of protection (DP) at the center of the protected area was higher for the case with the skirt. This skirt stimulates more down flow and prevents the early entry of particles into the protected area. It can also be concluded that Lamp Y, due to its open shape, has the most positive effect on the DP at the center. It has also been shown that the position of the lamp has an effect on the protected area.

The design and implementation of device operation and management system based on LAMP

International Nuclear Information System (INIS)

Liu Shuzhen; Ma Lanxin; Qi Fazhi

2010-01-01

In order to manage devices with high efficiency and high quality and make it more standardized, more scientific and more modern, we use the platform of LAMP (Linux-Apache-Mysql-PHP) to develop a web-based equipment operation and management system. It uses the Browser/Server mode as the system mode and Mysql as the database to complete the function, such as query, modify, add, delete, batch import, batch export user information and device information, automatic alarm, user reviews and so on. (authors)

Signal amplification of microRNAs with modified strand displacement-based cycling probe technology.

Science.gov (United States)

Jia, Huning; Bu, Ying; Zou, Bingjie; Wang, Jianping; Kumar, Shalen; Pitman, Janet L; Zhou, Guohua; Song, Qinxin

2016-10-24

Micro ribose nucleic acids (miRNAs) play an important role in biological processes such as cell differentiation, proliferation and apoptosis. Therefore, miRNAs are potentially a powerful marker for monitoring cancer and diagnosis. Here, we present sensitive signal amplification for miRNAs based on modified cycling probe technology with strand displacement amplification. miRNA was captured by the template coupled with beads, and then the first cycle based on SDA was repeatedly extended to the nicking end, which was produced by the extension reaction of miRNA. The products generated by SDA are captured by a molecular beacon (MB), which is designed to initiate the second amplification cycle, with a similar principle to the cycling probe technology (CPT), which is based on repeated digestion of the DNA-RNA hybrid by the RNase H. After one sample enrichment and two steps of signal amplification, 0.1 pM of let-7a can be detected. The miRNA assay exhibits a great dynamic range of over 100 orders of magnitude and high specificity to clearly discriminate a single base difference in miRNA sequences. This isothermal amplification does not require any special temperature control instrument. The assay is also about signal amplification rather than template amplification, therefore minimising contamination issues. In addition, there is no need for the reverse transcription (RT) process. Thus the amplification is suitable for miRNA detection.

Model of discharge lamps with magnetic ballast

OpenAIRE

Molina, Julio; Sainz Sapera, Luis; Mesas García, Juan José; Bergas Jané, Joan Gabriel

2013-01-01

Magnetic ballast discharge lamp modeling has been extensively studied because these lamps can be an important source of harmonics. Discharge lamp models usually represent the arc voltage by a square waveform. However, this waveform can be far from actual arc voltages, which affects the accuracy of the lamp models. This paper investigates the actual arc voltage behavior of discharge lamps from laboratory measurements and proposes a novel characterization of these voltages to reformulate the co...

[Investigation of RNA viral genome amplification by multiple displacement amplification technique].

Science.gov (United States)

Pang, Zheng; Li, Jian-Dong; Li, Chuan; Liang, Mi-Fang; Li, De-Xin

2013-06-01

In order to facilitate the detection of newly emerging or rare viral infectious diseases, a negative-strand RNA virus-severe fever with thrombocytopenia syndrome bunyavirus, and a positive-strand RNA virus-dengue virus, were used to investigate RNA viral genome unspecific amplification by multiple displacement amplification technique from clinical samples. Series of 10-fold diluted purified viral RNA were utilized as analog samples with different pathogen loads, after a series of reactions were sequentially processed, single-strand cDNA, double-strand cDNA, double-strand cDNA treated with ligation without or with supplemental RNA were generated, then a Phi29 DNA polymerase depended isothermal amplification was employed, and finally the target gene copies were detected by real time PCR assays to evaluate the amplification efficiencies of various methods. The results showed that multiple displacement amplification effects of single-strand or double-strand cDNA templates were limited, while the fold increases of double-strand cDNA templates treated with ligation could be up to 6 X 10(3), even 2 X 10(5) when supplemental RNA existed, and better results were obtained when viral RNA loads were lower. A RNA viral genome amplification system using multiple displacement amplification technique was established in this study and effective amplification of RNA viral genome with low load was achieved, which could provide a tool to synthesize adequate viral genome for multiplex pathogens detection.

The effect of operating lamps on the protected area of a unidirectional down flow (UDF) system

NARCIS (Netherlands)

Traversari, A.A.L.; Bottenheft, C.; Louman, R.; Heumen, S.P.M. van; Böggeman, J.

2016-01-01

Background: Operating lamps are often seen as the most disruptive factors within the protective area in the operating theater (OT). The effect of the operation lamps (with different shapes) should be demonstrated in an OT by trial, since research on the effects of the lamps is still limited.

A new LED lamp for the collection of nocturnal Lepidoptera and a spectral comparison of light-trapping lamps

Directory of Open Access Journals (Sweden)

Gunnar Brehm

2017-04-01

Full Text Available Most nocturnal Lepidoptera can be attracted to artificial light sources, particularly to those that emit a high proportion of ultraviolet radiation. Here, I describe a newly developed LED lamp set for the use in the field that is lightweight, handy, robust, and energy efficient. The emitted electromagnetic spectrum corresponds to the peak sensitivity in most Lepidoptera eye receptors (ultraviolet, blue and green. Power LEDs with peaks at 368 nm (ultraviolet, 450 nm (blue, 530 nm (green, and 550 nm (cool white are used. I compared the irradiance (Ee of many commonly used light-trapping lamps at a distance of 50 cm. Between wavelengths of 300 and 1000 nm, irradiance from the new lamp was 1.43 W m-2. The new lamp proved to be the most energy efficient, and it emitted more radiation in the range between 300 and 400 nm than any other lamp tested. Cold cathodes are the second most energy-efficient lamps. Irradiation from fluorescent actinic tubes is higher than from fluorescent blacklight-blue tubes. High-wattage incandescent lamps and self-ballasted mercury vapour lamps have highest irradiance, but they mainly emit in the long wave spectrum. The use of gauze and sheets decreases the proportion of UV radiation and increases the share of blue light, probably due to optical brighteners. Compared with sunlight, UV irradiance is low at a distance of 50 cm from the lamp, but (safety glasses as well as keeping sufficient distance from the lamp are recommended. In field tests, the new LED lamp attracted large numbers of Lepidoptera in both the Italian Alps and in the Peruvian Andes.

Isothermal amplification of environmental DNA (eDNA for direct field-based monitoring and laboratory confirmation of Dreissena sp.

Directory of Open Access Journals (Sweden)

Maggie R Williams

Full Text Available Loop-mediated isothermal amplification (LAMP of aquatic invasive species environmental DNA (AIS eDNA was used for rapid, sensitive, and specific detection of Dreissena sp. relevant to the Great Lakes (USA basin. The method was validated for two uses including i direct amplification of eDNA using a hand filtration system and ii confirmation of the results after DNA extraction using a conventional thermal cycler run at isothermal temperatures. Direct amplification eliminated the need for DNA extraction and purification and allowed detection of target invasive species in grab or concentrated surface water samples, containing both free DNA as well as larger cells and particulates, such as veligers, eggs, or seeds. The direct amplification method validation was conducted using Dreissena polymorpha and Dreissena bugensis and uses up to 1 L grab water samples for high target abundance (e.g., greater than 10 veligers (larval mussels per L for Dreissena sp. or 20 L samples concentrated through 35 μm nylon screens for low target abundance, at less than 10 veligers per liter water. Surface water concentrate samples were collected over a period of three years, mostly from inland lakes in Michigan with the help of a network of volunteers. Field samples collected from 318 surface water locations included i filtered concentrate for direct amplification validation and ii 1 L grab water sample for eDNA extraction and confirmation. Though the extraction-based protocol was more sensitive (resulting in more positive detections than direct amplification, direct amplification could be used for rapid screening, allowing for quicker action times. For samples collected between May and August, results of eDNA direct amplification were consistent with known presence/absence of selected invasive species. A cross-platform smartphone application was also developed to disseminate the analyzed results to volunteers. Field tests of the direct amplification protocol using a

Capacitive discharge exciplex lamps

Energy Technology Data Exchange (ETDEWEB)

Sosnin, E A; Erofeev, M V; Tarasenko, V F [High Current Electronics Institute, 2/3, Akademichesky Ave., Tomsk 634055 (Russian Federation)

2005-09-07

Simple-geometry exciplex lamps of a novel type excited by a capacitive discharge (CD-excilamps) have been investigated. An efficient radiation has been obtained on KrBr*, KrCl*, XeBr*, XeCl* molecules and I* atom. The highest values of efficiency of various working molecules are approximately 10-18%. The lifetime of the operating gas mixture in KrCl*, XeCl*, I* and XeBr* exciplex lamps excited by a CD is above 1000 h. Owing to the above-mentioned characteristics, the exciplex lamps excited by a CD are supposed to be very promising for various applications.

Capacitive discharge exciplex lamps

International Nuclear Information System (INIS)

Sosnin, E A; Erofeev, M V; Tarasenko, V F

2005-01-01

Simple-geometry exciplex lamps of a novel type excited by a capacitive discharge (CD-excilamps) have been investigated. An efficient radiation has been obtained on KrBr*, KrCl*, XeBr*, XeCl* molecules and I* atom. The highest values of efficiency of various working molecules are approximately 10-18%. The lifetime of the operating gas mixture in KrCl*, XeCl*, I* and XeBr* exciplex lamps excited by a CD is above 1000 h. Owing to the above-mentioned characteristics, the exciplex lamps excited by a CD are supposed to be very promising for various applications

49 CFR 234.221 - Lamp voltage.

Science.gov (United States)

2010-10-01

... 49 Transportation 4 2010-10-01 2010-10-01 false Lamp voltage. 234.221 Section 234.221 Transportation Other Regulations Relating to Transportation (Continued) FEDERAL RAILROAD ADMINISTRATION..., Inspection, and Testing Maintenance Standards § 234.221 Lamp voltage. The voltage at each lamp shall be...

Realization of quantum state privacy amplification in a nuclear magnetic resonance quantum system

International Nuclear Information System (INIS)

Hao, Liang; Wang, Chuan; Long, Gui Lu

2010-01-01

Quantum state privacy amplification (QSPA) is the quantum analogue of classical privacy amplification. If the state information of a series of single-particle states has some leakage, QSPA reduces this leakage by condensing the state information of two particles into the state of one particle. Recursive applications of the operations will eliminate the quantum state information leakage to a required minimum level. In this paper, we report the experimental implementation of a quantum state privacy amplification protocol in a nuclear magnetic resonance system. The density matrices of the states are constructed in the experiment, and the experimental results agree well with theory.

30 CFR 57.17010 - Electric lamps.

Science.gov (United States)

2010-07-01

... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Electric lamps. 57.17010 Section 57.17010 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND NONMETAL MINE....17010 Electric lamps. Individual electric lamps shall be carried for illumination by all persons...

[Study on the discharge properties of xeon flash lamp and experimental measurement].

Science.gov (United States)

Zhao, You-Quan; Miao, Pei-Liang; He, Feng; Gu, Jian; Zhai, Rui-Wei

2014-07-01

The Xenon flash lamp is a new type of light source for analytical instrument. The present paper analyzed the discharge process of xenon flash lamp, presented the discharge test system, and conducted experimental measurement of the voltage, current and optical pulse signal in the process of discharge. The results show that in the preliminary discharge, the free electron concentration was at a low level, so the energy was at a low level, then following the gas discharge, numerous free electrons formed in the lamp, resultin in the increase in the concentration of free electrons, therefore discharge current rised rapidly and voltage reduced. The lamp released photons to generate light pulse in the moment of ionic recombination, The pulse xenon lamp light energy output and spectral characteristic is related to electron energy in recombination and combination level of xenon, if the input energy and the energy consumption of the xenon lamp is inconsistent, it will lead to repeated capacitor charging and discharging and produce oscillation waveform. This paper is very useful for understanding the process of xenon lamp discharge, optimizing the driver circuit and the production of xenon flash lamp.

A Sweet Spot for Molecular Diagnostics: Coupling Isothermal Amplification and Strand Exchange Circuits to Glucometers

Science.gov (United States)

Du, Yan; Hughes, Randall A.; Bhadra, Sanchita; Jiang, Yu Sherry; Ellington, Andrew D.; Li, Bingling

2015-06-01

Strand exchange nucleic acid circuitry can be used to transduce isothermal nucleic acid amplification products into signals that can be readable on an off-the-shelf glucometer. Loop-mediated isothermal amplification (LAMP) is limited by the accumulation of non-specific products, but nucleic acid circuitry can be used to probe and distinguish specific amplicons. By combining this high temperature isothermal amplification method with a thermostable invertase, we can directly transduce Middle-East respiratory syndrome coronavirus and Zaire Ebolavirus templates into glucose signals, with a sensitivity as low as 20-100 copies/μl, equating to atto-molar (or low zepto-mole). Virus from cell lysates and synthetic templates could be readily amplified and detected even in sputum or saliva. An OR gate that coordinately triggered on viral amplicons further guaranteed fail-safe virus detection. The method describes has potential for accelerating point-of-care applications, in that biological samples could be applied to a transducer that would then directly interface with an off-the-shelf, approved medical device.

THE APPARATUS FOR ALIGNMENT OF THE PHOTOMETRIC LAMP FILAMENT

Directory of Open Access Journals (Sweden)

V. A. Dlugunovich

2015-01-01

Full Text Available During photometric measurements involving the use of photometric lamps it is necessary that the filament of lamp takes a strictly predetermined position with respect to the photodetector and the optical axis of the photometric setup. The errors in positioning of alignment filament with respect to the optical axis of the measuring system lead to increase the uncertainty of measurement of the photometric characteristics of the light sources. A typical method for alignment of filament of photometric lamps is based on the use a diopter tubes (telescopes. Using this method, the mounting of filament to the required position is carried out by successive approximations, which requires special concentration and a lot of time. The aim of this work is to develop an apparatus for alignment which allows simultaneous alignment of the filament of lamps in two mutually perpendicular planes. The method and apparatus for alignment of the photometric lamp filament during measurements of the photometric characteristics of light sources based on two digital video cameras is described in this paper. The apparatus allows to simultaneously displaying the image of lamps filament on the computer screen in two mutually perpendicular planes. The apparatus eliminates a large number of functional units requiring elementwise alignment and reduces the time required to carry out the alignment. The apparatus also provides the imaging of lamps filament with opaque coated on the bulb. The apparatus is used at the National standard of light intensity and illuminance units of the Republic of Belarus. 

Lamps recycling aiming at the environment preservation

International Nuclear Information System (INIS)

Yamachita, Roberto Akira; Gama, Paulo Henrique R. Pereira; Haddad, Jamil; Santos, Afonso H. Moreira; Guardia, Eduardo C.

1999-01-01

The article discusses the following issues of lamps recycling in Brazil: mercury lamps recycling, recycling potential, energy conservation and environmental impacts, enterprises lamps recycling, and incentives policy

Loop-mediated isothermal amplification: rapid detection of Angiostrongylus cantonensis infection in Pomacea canaliculata

Directory of Open Access Journals (Sweden)

Zhuo MingMing

2011-10-01

Full Text Available Abstract Background Angiostrongylus cantonensis is a zoonotic parasite that causes eosinophilic meningitis in humans. The most common source of infection with A. cantonensis is the consumption of raw or undercooked mollusks (e.g., snails and slugs harbouring infectious third-stage larvae (L3. However, the parasite is difficult to identify in snails. The purpose of this study was to develop a quick, simple molecular method to survey for A. cantonensis in intermediate host snails. Findings We used a loop-mediated isothermal amplification (LAMP assay, which was performed using Bst DNA polymerase. Reactions amplified the A. cantonensis 18S rRNA gene and demonstrated high sensitivity; as little as 1 fg of DNA was detected in the samples. Furthermore, no cross-reactivity was found with other parasites such as Toxoplasma gondii, Plasmodium falciparum, Schistosoma japonicum, Clonorchis sinensis, Paragonimus westermani and Anisakis. Pomacea canaliculata snails were exposed to A. cantonensis first-stage larvae (L1 in the laboratory, and L3 were observed in the snails thirty-five days after infection. All nine samples were positive as determined by the LAMP assay for A. cantonensis, which was identified as positive by using PCR and microscopy, this demonstrates that LAMP is sensitive and effective for diagnosis. Conclusions LAMP is an appropriate diagnostic method for the routine identification of A. cantonensis within its intermediate host snail P. canaliculata because of its simplicity, sensitivity, and specificity. It holds great promise as a useful monitoring tool for A. cantonensis in endemic regions.

Loop-Mediated Isothermal Amplification Targeting Actin DNA of Trichomonas vaginalis.

Science.gov (United States)

Goo, Youn-Kyoung; Shin, Won-Sik; Yang, Hye-Won; Joo, So-Young; Song, Su-Min; Ryu, Jae-Sook; Kong, Hyun-Hee; Lee, Won-Ki; Chung, Dong-Il; Hong, Yeonchul

2016-06-01

Trichomoniasis caused by Trichomonas vaginalis is a common sexually transmitted disease. Its association with several health problems, including preterm birth, pelvic inflammatory disease, cervical cancer, and transmission of human immunodeficiency virus, emphasizes the importance of improved access to early and accurate detection of T. vaginalis. In this study, a rapid and efficient loop-mediated isothermal amplification-based method for the detection of T. vaginalis was developed and validated, using vaginal swab specimens from subjects suspected to have trichomoniasis. The LAMP assay targeting the actin gene was highly sensitive with detection limits of 1 trichomonad and 1 pg of T. vaginalis DNA per reaction, and specifically amplified the target gene only from T. vaginalis. Validation of this assay showed that it had the highest sensitivity and better agreement with PCR (used as the gold standard) compared to microscopy and multiplex PCR. This study showed that the LAMP assay, targeting the actin gene, could be used to diagnose early infections of T. vaginalis. Thus, we have provided an alternative molecular diagnostic tool and a point-of-care test that may help to prevent trichomoniasis transmission and associated complications.

Graphene oxide based fluorescence resonance energy transfer and loop-mediated isothermal amplification for white spot syndrome virus detection.

Science.gov (United States)

Waiwijit, U; Phokaratkul, D; Kampeera, J; Lomas, T; Wisitsoraat, A; Kiatpathomchai, W; Tuantranont, A

2015-10-20

Graphene oxide (GO) is attractived for biological or medical applications due to its unique electrical, physical, optical and biological properties. In particular, GO can adsorb DNA via π-π stacking or non-covalent interactions, leading to fluorescence quenching phenomenon applicable for bio-molecular detection. In this work, a new method for white spot syndrome virus (WSSV)-DNA detection is developed based on loop-mediated isothermal amplification (LAMP) combined with fluorescence resonance energy transfer (FRET) between GO and fluorescein isothiocyanate-labeled probe (FITC-probe). The fluorescence quenching efficiency of FITC-probe was found to increase with increasing GO concentration and reached 98.7% at a GO concentration of 50 μg/ml. The fluorescence intensity of FITC-probe was recovered after hybridization with WSSV LAMP product with an optimal hybridization time of 10 min and increased accordingly with increasing amount of LAMP products. The detection limit was estimated to be as low as 10 copies of WSSV plasmid DNA or 0.6 fg of the total DNA extracted from shrimp infected with WSSV. In addition, no cross reaction was observed with other common shrimp viral pathogens. Therefore, the GO-FRET-LAMP technique is promising for fast, sensitive and specific detection of DNAs. Copyright © 2015 Elsevier B.V. All rights reserved.

CALiPER Retail Lamps Study 3.1: Dimming, Flicker, and Power Quality Characteristics of LED A Lamps

Energy Technology Data Exchange (ETDEWEB)

none,

2014-12-31

This CALiPER report examines the characteristics of a subset of lamps from CALiPER Retail Lamps Study 3 in more detail. Specifically, it focuses on the dimming, power quality, and flicker characteristics of 14 LED A lamps, as controlled by four different retail-available dimmers.

Thomson scattering on a low-pressure, inductively-coupled gas discharge lamp

International Nuclear Information System (INIS)

Sande, M.J. van de; Mullen, J.J.A.M. van der

2002-01-01

Excitation and light production processes in gas discharge lamps are the result of inelastic collisions between atoms and free electrons in the plasma. Therefore, knowledge of the electron density n e and temperature T e is essential for a proper understanding of such plasmas. In this paper, an experimental system for laser Thomson scattering on a low-pressure, inductively-coupled gas discharge lamp and measurements of n e and T e in this lamp are presented. The experimental system is suitable for low electron temperatures (down to below 0.2 eV) and employs a triple grating spectrograph for a high stray light rejection, or equivalently a low stray light redistribution (R eff approximately 7x10 -9 nm -1 at 0.5 nm from the laser wavelength). The electron density detection limit of the system is n e approximately 10 16 m -3 . The modifications to the lamp that were necessary for the measurements are described, and results are presented and compared to previous work and trends expected from the electron particle and energy balances. The electron density and temperature are about n e approximately 10 19 m -3 and T e approximately 1 eV in the most active part of the plasma; the exact values depend on the argon filling pressure, the mercury pressure and the position in the lamp. (author)

Portable nucleic acid thermocyclers.

Science.gov (United States)

Almassian, David R; Cockrell, Lisa M; Nelson, William M

2013-11-21

A nucleic acid thermal cycler is considered to be portable if it is under ten pounds, easily carried by one individual, and battery powered. Nucleic acid amplification includes both polymerase chain reaction (e.g. PCR, RT-PCR) and isothermal amplification (e.g. RPA, HDA, LAMP, NASBA, RCA, ICAN, SMART, SDA). There are valuable applications for portable nucleic acid thermocyclers in fields that include clinical diagnostics, biothreat detection, and veterinary testing. A system that is portable allows for the distributed detection of targets at the point of care and a reduction of the time from sample to answer. The designer of a portable nucleic acid thermocycler must carefully consider both thermal control and the detection of amplification. In addition to thermal control and detection, the designer may consider the integration of a sample preparation subsystem with the nucleic acid thermocycler. There are a variety of technologies that can achieve accurate thermal control and the detection of nucleic acid amplification. Important evaluation criteria for each technology include maturity, power requirements, cost, sensitivity, speed, and manufacturability. Ultimately the needs of a particular market will lead to user requirements that drive the decision between available technologies.

Spectroscopy on metal-halide lamps under varying gravity conditions

NARCIS (Netherlands)

Flikweert, A.J.

2008-01-01

Worldwide, 20% of all electricity is used for lighting. For this reason, efficient lamps are economically and ecologically important. High intensity discharge (HID) lamps are efficient lamps. The most common HID lamp these days is the metal-halide (MH) lamp. MH lamps have a good colour rendering

Endpoint visual detection of three genetically modified rice events by loop-mediated isothermal amplification.

Science.gov (United States)

Chen, Xiaoyun; Wang, Xiaofu; Jin, Nuo; Zhou, Yu; Huang, Sainan; Miao, Qingmei; Zhu, Qing; Xu, Junfeng

2012-11-07

Genetically modified (GM) rice KMD1, TT51-1, and KF6 are three of the most well known transgenic Bt rice lines in China. A rapid and sensitive molecular assay for risk assessment of GM rice is needed. Polymerase chain reaction (PCR), currently the most common method for detecting genetically modified organisms, requires temperature cycling and relatively complex procedures. Here we developed a visual and rapid loop-mediated isothermal amplification (LAMP) method to amplify three GM rice event-specific junction sequences. Target DNA was amplified and visualized by two indicators (SYBR green or hydroxy naphthol blue [HNB]) within 60 min at an isothermal temperature of 63 °C. Different kinds of plants were selected to ensure the specificity of detection and the results of the non-target samples were negative, indicating that the primer sets for the three GM rice varieties had good levels of specificity. The sensitivity of LAMP, with detection limits at low concentration levels (0.01%−0.005% GM), was 10- to 100-fold greater than that of conventional PCR. Additionally, the LAMP assay coupled with an indicator (SYBR green or HNB) facilitated analysis. These findings revealed that the rapid detection method was suitable as a simple field-based test to determine the status of GM crops.

Endpoint Visual Detection of Three Genetically Modified Rice Events by Loop-Mediated Isothermal Amplification

Directory of Open Access Journals (Sweden)

Qing Zhu

2012-11-01

Full Text Available Genetically modified (GM rice KMD1, TT51-1, and KF6 are three of the most well known transgenic Bt rice lines in China. A rapid and sensitive molecular assay for risk assessment of GM rice is needed. Polymerase chain reaction (PCR, currently the most common method for detecting genetically modified organisms, requires temperature cycling and relatively complex procedures. Here we developed a visual and rapid loop-mediated isothermal amplification (LAMP method to amplify three GM rice event-specific junction sequences. Target DNA was amplified and visualized by two indicators (SYBR green or hydroxy naphthol blue [HNB] within 60 min at an isothermal temperature of 63 °C. Different kinds of plants were selected to ensure the specificity of detection and the results of the non-target samples were negative, indicating that the primer sets for the three GM rice varieties had good levels of specificity. The sensitivity of LAMP, with detection limits at low concentration levels (0.01%–0.005% GM, was 10- to 100-fold greater than that of conventional PCR. Additionally, the LAMP assay coupled with an indicator (SYBR green or HNB facilitated analysis. These findings revealed that the rapid detection method was suitable as a simple field-based test to determine the status of GM crops.

Loop-Mediated Isothermal Amplification on Crude DNA as a Point-of-Care Test for the Diagnosis of Mycoplasma-Related Vaginitis During Early Pregnancy.

Science.gov (United States)

Wang, Yichao; Zhang, Bumei; Sun, Yan; Liu, Yunde; Gu, Yajun

2017-12-20

Mycoplasma-related vaginitis gradually has been growing as a threat in adults-genitourinary infection contributes to funisitis, spontaneous abortion, and low birth weight. Until now, use of loop-mediated isothermal amplification (LAMP) to detect Ureaplasma urealyticum (UU), Mycoplasma hominis (MH), or Mycoplasma genitalium (MG) has been reported by some researchers. However, previous studies focused on purified DNA as the template for LAMP assay, which is usually extracted via commercial kit. We developed a LAMP assay for rapid detection of UU, MH, and MG genital mycoplasmas using a simple boiling method for DNA extraction, in a cohort of pregnant women with mycoplasma-related vaginitis. We monitored amplicons with the naked eye using SYBR Green I. The cohort in our study showed a prevalence of 22.6% in pregnant women, as detected by UU-LAMP assay. Compared to the polymerase chain reaction (PCR) test with purified DNA, the sensitivity of the UU-LAMP in clinical specimens with crude DNA was 87.5% (95% confidence interval [CI], 64.6%->99.9). For crude DNA specimens, UU-LAMP was more sensitive and reliable than PCR, with a higher agreement rate (96.8%) and Youden index value (0.88). As a point-of-care test, LAMP is a useful, specific, and efficient way to detect genital mycoplasmas in resource-limited settings, especially for crude DNA. © American Society for Clinical Pathology 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com

CALiPER Retail Lamps Study 3

Energy Technology Data Exchange (ETDEWEB)

Royer, Michael P. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States); Beeson, Tracy A. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States)

2014-02-01

The CALiPER program first began investigating LED lamps sold at retail stores in 2010, purchasing 33 products from eight retailers and covering six product categories. The findings revealed a fragmented marketplace, with large disparities in performance of different products, accuracy of manufacturer claims, and offerings from different retail outlets. Although there were some good products, looking back many would not be considered viable competitors to other available options, with too little lumen output, not high enough efficacy, or poor color quality. CALiPER took another look in late 2011purchasing 38 products of five different types from nine retailers and the improvement was marked. Performance was up; retailer claims were more accurate; and the price per lumen and price per unit efficacy were down, although the price per product had not changed much. Nonetheless, there was still plenty of room for improvement, with the performance of LED lamps not yet reaching that of well-established classes of conventional lamps (e.g., 75 W incandescent A19 lamps). Since the second retail lamp study was published in early 2012, there has been substantial progress in all aspects of LED lamps available from retailers. To document this progress, CALiPER again purchased a sample of lamps from retail stores 46 products in total, focusing on A19, PAR30, and MR16 lamps but instead of a random sample, sought to select products to answer specific hypotheses about performance. These hypotheses focused on expanding ranges of LED equivalency, the accuracy of lifetime claims, efficacy and price trends, as well as changes to product designs. Among other results, key findings include: There are now very good LED options to compete with 60 W, 75 W, and 100 W incandescent A19 lamps, and 75 W halogen PAR30 lamps. MR16 lamps have shown less progress, but there are now acceptable alternatives to 35 W, 12 V halogen MR16 lamps and 50 W, 120 V halogen MR16 lamps for some applications. Other

System of programming units for the K556RT4 and K556RT5 fixed programmed memory devices

International Nuclear Information System (INIS)

Bobkov, S.G.; Ermolin, Yu.V.; Kantserov, V.A.; Strigin, V.B.

1983-01-01

The programming system of constant programmable memory devices K556RT4 and K556RT5 that consist of two units (a programming device and an electrothermotraining unit) is described. The modules are made in the KAMAK standard. The programming device takes up 2 normal places, while the electrothermotraining block takes up 1 place. As information recording is done using a computer the time for programming is reduced and the possibility of errors is limited as compared with the manual method. The computer introduces the whole word to be recorded, not the separate parts, in the programming device. The transition to a new digit of a given word in the programming device is done automatically. This reduces the expense of computer time and accelerates the programming of microdiagrams

Computational method and system for modeling, analyzing, and optimizing DNA amplification and synthesis

Science.gov (United States)

Vandersall, Jennifer A.; Gardner, Shea N.; Clague, David S.

2010-05-04

A computational method and computer-based system of modeling DNA synthesis for the design and interpretation of PCR amplification, parallel DNA synthesis, and microarray chip analysis. The method and system include modules that address the bioinformatics, kinetics, and thermodynamics of DNA amplification and synthesis. Specifically, the steps of DNA selection, as well as the kinetics and thermodynamics of DNA hybridization and extensions, are addressed, which enable the optimization of the processing and the prediction of the products as a function of DNA sequence, mixing protocol, time, temperature and concentration of species.

Detection of Toxoplasma gondii oocysts in different water resources by Loop Mediated Isothermal Amplification (LAMP).

Science.gov (United States)

Gallas-Lindemann, Carmen; Sotiriadou, Isaia; Mahmoodi, Mohammad Reza; Karanis, Panagiotis

2013-02-01

Human toxoplasmosis is potentially contracted due to consumption of contaminated drinking water and represents an increasing public health risk worldwide. Toxoplasma gondii oocysts can be resistant to standard disinfection processes, including UV radiation. Increased awareness of the risk of waterborne toxoplasmosis outbreaks has led to an increase in research interest in the detection of oocysts in environmental water systems. Ninety-five environmental water samples from the Lower Rhine area in Germany have been included in the study and examined for the presence of Toxoplasma. Water samples were filtered or flocculated by aluminum sulfate and purified by sucrose density gradient. DNA was then extracted, and the DNA samples were then examined by LAMP analysis. T. gondii DNA was detected in eight out of 83 (9.6%) influent and effluent samples obtained from wastewater treatment plants. All samples (n=12) from the surface, ground, raw and tap waters tested negative. The purpose of this work was to investigate the occurrence and distribution of Toxoplasma oocysts on the Lower Rhine in Germany. Our study provides evidence that the assay is a sensitive, specific, rapid and cost effective method for the detection of T. gondii and is useful for both the investigations of cases of waterborne outbreaks and for identifying the source of contamination. Copyright © 2012 Elsevier B.V. All rights reserved.

Rapid identification of drug-type strains in Cannabis sativa using loop-mediated isothermal amplification assay.

Science.gov (United States)

Kitamura, Masashi; Aragane, Masako; Nakamura, Kou; Watanabe, Kazuhito; Sasaki, Yohei

2017-01-01

In Cannabis sativa L., tetrahydrocannabinol (THC) is the primary psychoactive compound and exists as the carboxylated form, tetrahydrocannabinolic acid (THCA). C. sativa is divided into two strains based on THCA content-THCA-rich (drug-type) strains and THCA-poor (fiber-type) strains. Both strains are prohibited by law in many countries including Japan, whereas the drug-type strains are regulated in Canada and some European countries. As the two strains cannot be discriminated by morphological analysis, a simple method for identifying the drug-type strains is required for quality control in legal cultivation and forensic investigation. We have developed a novel loop-mediated isothermal amplification (LAMP) assay for identifying the drug-type strains of C. sativa. We designed two selective LAMP primer sets for on-site or laboratory use, which target the drug-type THCA synthase gene. The LAMP assay was accomplished within approximately 40 min. The assay showed high specificity for the drug-type strains and its sensitivity was the same as or higher than that of conventional polymerase chain reaction. We also showed the effectiveness of melting curve analysis that was conducted after the LAMP assay. The melting temperature values of the drug-type strains corresponded to those of the cloned drug-type THCA synthase gene, and were clearly different from those of the cloned fiber-type THCA synthase gene. Moreover, the LAMP assay with simple sample preparation could be accomplished within 1 h from sample treatment to identification without the need for special devices or techniques. Our rapid, sensitive, specific, and simple assay is expected to be applicable to laboratory and on-site detection.

Inductive tuners for microwave driven discharge lamps

Science.gov (United States)

Simpson, James E.

1999-01-01

An RF powered electrodeless lamp utilizing an inductive tuner in the waveguide which couples the RF power to the lamp cavity, for reducing reflected RF power and causing the lamp to operate efficiently.

An Open Source "Smart Lamp" for the Optimization of Plant Systems and Thermal Comfort of Offices.

Science.gov (United States)

Salamone, Francesco; Belussi, Lorenzo; Danza, Ludovico; Ghellere, Matteo; Meroni, Italo

2016-03-07

The article describes the design phase, development and practical application of a smart object integrated in a desk lamp and called "Smart Lamp", useful to optimize the indoor thermal comfort and energy savings that are two important workplace issues where the comfort of the workers and the consumption of the building strongly affect the economic balance of a company. The Smart Lamp was built using a microcontroller, an integrated temperature and relative humidity sensor, some other modules and a 3D printer. This smart device is similar to the desk lamps that are usually found in offices but it allows one to adjust the indoor thermal comfort, by interacting directly with the air conditioner. After the construction phase, the Smart Lamp was installed in an office normally occupied by four workers to evaluate the indoor thermal comfort and the cooling consumption in summer. The results showed how the application of the Smart Lamp effectively reduced the energy consumption, optimizing the thermal comfort. The use of DIY approach combined with read-write functionality of websites, blog and social platforms, also allowed to customize, improve, share, reproduce and interconnect technologies so that anybody could use them in any occupied environment.

Precision charge amplification and digitization system for a scintillating and lead glass array

Energy Technology Data Exchange (ETDEWEB)

Delchamps, S.W.; Rameika, R.; Arenton, M.; Chen, T.Y.; Conetti, S.; Cox, B.; Etemadi, B.; Fortney, L.; Guffey, K.; Haire, M.

1989-01-01

A 544-channel low-noise, high-rate, precision charge amplification and ADC system was constructed for the Fermilab Experiment 705 electromagnetic calorimeter, which employs SCG1-C scintillating glass and SF5 lead glass instrumented with photo-multiplier tubes. A general discussion of the system is given, and the charge amplification, fast trigger pulse generation, and analog to digital conversion aspects of the system are presented in more detail. Performance is evaluated using data from Experiment 705 and from off-line tests. Short and long term pedestal stability, baseline recovery and rate capability, linearity of response, and crosstalk between channels are discussed. 8 refs., 2 tabs.

Precision charge amplification and digitization system for a scintillating and lead glass array

International Nuclear Information System (INIS)

Delchamps, S.W.; Rameika, R.; Arenton, M.

1989-01-01

A 544-channel low-noise, high-rate, precision charge amplification and ADC system was constructed for the Fermilab Experiment 705 electromagnetic calorimeter, which employs SCG1-C scintillating glass and SF5 lead glass instrumented with photo-multiplier tubes. A general discussion of the system is given, and the charge amplification, fast trigger pulse generation, and analog to digital conversion aspects of the system are presented in more detail. Performance is evaluated using data from Experiment 705 and from off-line tests. Short and long term pedestal stability, baseline recovery and rate capability, linearity of response, and crosstalk between channels are discussed. 8 refs., 2 tabs

Development of an Automated Microfluidic System for DNA Collection, Amplification, and Detection of Pathogens

Energy Technology Data Exchange (ETDEWEB)

Hagan, Bethany S.; Bruckner-Lea, Cynthia J.

2002-12-01

This project was focused on developing and testing automated routines for a microfluidic Pathogen Detection System. The basic pathogen detection routine has three primary components; cell concentration, DNA amplification, and detection. In cell concentration, magnetic beads are held in a flow cell by an electromagnet. Sample liquid is passed through the flow cell and bacterial cells attach to the beads. These beads are then released into a small volume of fluid and delivered to the peltier device for cell lysis and DNA amplification. The cells are lysed during initial heating in the peltier device, and the released DNA is amplified using polymerase chain reaction (PCR) or strand displacement amplification (SDA). Once amplified, the DNA is then delivered to a laser induced fluorescence detection unit in which the sample is detected. These three components create a flexible platform that can be used for pathogen detection in liquid and sediment samples. Future developments of the system will include on-line DNA detection during DNA amplification and improved capture and release methods for the magnetic beads during cell concentration.

Evaluation of two loop-mediated isothermal amplification methods for the detection of Salmonella Enteritidis and Listeria monocytogenes in artificially contaminated ready-to-eat fresh products

Directory of Open Access Journals (Sweden)

Angeliki Birmpa

2015-08-01

Full Text Available In the present study, the effectiveness of two loop-mediated isothermal amplification (LAMP assays was evaluated. Samples of romaine lettuce, strawberries, cherry tomatoes, green onions and sour berries were inoculated with known dilutions (100-108 CFU/g of produce of S. Enteritidis and L. monocytogenes. With LAMP assay, pathogens can be detected in less than 60 min. The limits of detection of S. Enteritidis and L. monocytogenes depended on the food sample tested and on the presence of enrichment step. After enrichment steps, all food samples were found positive even at low initial pathogen levels. The developed LAMP, assays, are expected to become a valuable, robust, innovative, powerful, cheap and fast monitoring tool, which can be extensively used for routine analysis, and screening of contaminated foods by the food industry and the Public Food Health Authorities.

The instantaneous light-intensity function of a fluorescent lamp

Energy Technology Data Exchange (ETDEWEB)

Gluskin, Emanuel [Holon Institute of Technology, 52 Golomb St., Holon 58102 (Israel): Electrical Engineering Department, Ben-Gurion University, Beer-Sheva 84105 (Israel)]. E-mail: gluskin@ee.bgu.ac.il; Topalis, Frangiskos V. [Technical University of Athens, School of Electrical and Computer Engineering, 9 Iroon Politechniou St., 15780 Athens (Greece); Kateri, Ifigenia [Technical University of Athens, School of Electrical and Computer Engineering, 9 Iroon Politechniou St., 15780 Athens (Greece); Bisketzis, Nikolas [Technical University of Athens, School of Electrical and Computer Engineering, 9 Iroon Politechniou St., 15780 Athens (Greece)

2006-05-08

Using some simple physics and 'system' considerations, the instantaneous light intensity function {psi}(t) of a fluorescent lamp fed via a regular ballast from the 50-60 Hz line is argued to be {psi}(t)={psi}{sub min}+bp(t), where p(t) is the instantaneous power function of the lamp, and b is a constant, and experiment confirms this formula well. The main frequency of {psi}(t), the very significant singularity of its waveform, and the relative intensity of the ripple, i.e., the depth of the modulation, are the focus. The results are important for research into the vision problem that some humans (autistic, but others, too) experience regarding fluorescent light. The inertia of the processes in the lamp which are responsible for the light emission, provides some nonzero emission at the instants when p(t) has zeros. The smaller the volume of the tube and the mass of the gas are, the more weakly the inertia of the processes is expressed, and the relatively smaller is {psi}{sub min}. However, it should be very difficult to theoretically obtain {psi}(t), in particular {psi}{sub min}, from the very complicated physics of the low-pressure discharge in the tube. We conclude that {psi}{sub min} has to be connected with the (also easily measured) lamp's inductance. The work should attract more attention of the physicists to the properties of the common fluorescent lamps. escent lamps.

Loop-mediated isothermal DNA amplification for asymptomatic malaria detection in challenging field settings: Technical performance and pilot implementation in the Peruvian Amazon.

Directory of Open Access Journals (Sweden)

Elisa Serra-Casas

Full Text Available Loop-mediated isothermal DNA amplification (LAMP methodology offers an opportunity for point-of-care (POC molecular detection of asymptomatic malaria infections. However, there is still little evidence on the feasibility of implementing this technique for population screenings in isolated field settings.Overall, we recruited 1167 individuals from terrestrial ('road' and hydric ('riverine' communities of the Peruvian Amazon for a cross-sectional survey to detect asymptomatic malaria infections. The technical performance of LAMP was evaluated in a subgroup of 503 samples, using real-time Polymerase Chain Reaction (qPCR as reference standard. The operational feasibility of introducing LAMP testing in the mobile screening campaigns was assessed based on field-suitability parameters, along with a pilot POC-LAMP assay in a riverine community without laboratory infrastructure.LAMP had a sensitivity of 91.8% (87.7-94.9 and specificity of 91.9% (87.8-95.0, and the overall accuracy was significantly better among samples collected during road screenings than riverine communities (p≤0.004. LAMP-based diagnostic strategy was successfully implemented within the field-team logistics and the POC-LAMP pilot in the riverine community allowed for a reduction in the turnaround time for case management, from 12-24 hours to less than 5 hours. Specimens with haemolytic appearance were regularly observed in riverine screenings and could help explaining the hindered performance/interpretation of the LAMP reaction in these communities.LAMP-based molecular malaria diagnosis can be deployed outside of reference laboratories, providing similar performance as qPCR. However, scale-up in remote field settings such as riverine communities needs to consider a number of logistical challenges (e.g. environmental conditions, labour-intensiveness in large population screenings that can influence its optimal implementation.

Electronic Dimmable Ballasts for High-Intensity Discharge Sodium Vapor and Metal Halide Lamps

Science.gov (United States)

Boulanger, Richard

2002-01-01

Two types of high-intensity discharge lamps were tested using dimmable ballasts. The main purpose for evaluating this lighting system was to determine its efficacy for saving power. Whereas previous variable level lighting systems for HID lamps in Advanced Life Support applications were adjustable in two or three steps using capacitive switching, this system allows for continuously adjustable lamp output. This type of lighting system when used as part of an Advanced Life Support biomass production system would provide only the amount of light energy a crop needed at any particular point in its growth cycle. Since most of the equivalent system mass in an ALS system is from the light energy required to grow the crops, controlling that light energy dynamically over a continuous range of operation would dramatically reduce the power consumption and reduce system mass.

The effects of a sound-field amplification system on managerial time in middle school physical education settings.

Science.gov (United States)

Ryan, Stu

2009-04-01

The focus of this research effort was to examine the effect of a sound-field amplification system on managerial time in the beginning of class in a physical education setting. A multiple baseline design across participants was used to measure change in the managerial time of 2 middle school female physical education teachers using a portable sound-field amplification system. Managerial time is defined as the cumulative amount of time that students spend on organizational, transitional, and nonsubject matter tasks in a lesson. The findings showed that the amount of managerial time at the beginning of class clearly decreased when the teacher used sound-field amplification feedback to physical education students. Findings indicate an immediate need for administrators to determine the most appropriate, cost-effective procedure to support sound-field amplification systems in existing physical education settings.

Visual Detection of Canine Parvovirus Based on Loop-Mediated Isothermal Amplification Combined with Enzyme-Linked Immunosorbent Assay and with Lateral Flow Dipstick

Science.gov (United States)

SUN, Yu-Ling; YEN, Chon-Ho; TU, Ching-Fu

2013-01-01

ABSTRACT Loop-mediated isothermal amplification (LAMP) combined with enzyme-linked immunosorbent assay (LAMP–ELISA) and with lateral flow dipstick (LAMP–LFD) are rapid, sensitive and specific methods for the visual detection of clinical pathogens. In this study, LAMP–ELISA and LAMP–LFD were developed for the visual detection of canine parvovirus (CPV). For LAMP, a set of four primers (biotin-labeled forward inner primers) was designed to specifically amplify a region of the VP2 gene of CPV. The optimum time and temperature for LAMP were 60 min and 65°C, respectively. The specific capture oligonucleotide probes, biotin-labeled CPV probe for LAMP–ELISA and fluorescein isothiocyanate-labeled CPV probe for LAMP–LFD were also designed for hybridization with LAMP amplicons on streptavidin-coated wells and LFD strips, respectively. For the comparison of detection sensitivity, conventional PCR and LAMP for CPV detection were also performed. The CPV detection limits by PCR, PCR–ELISA, LAMP, LAMP–ELISA and LAMP–LFD were 102, 102, 10−1, 10−1 and 10−1 TCID50/ml, respectively. In tests using artificially contaminated dog fecal samples, the samples with CPV inoculation levels of ≥1 TCID50/ml gave positive results by both LAMP–ELISA and LAMP–LFD. Our data indicated that both LAMP–ELISA and LAMP–LFD are promising as rapid, sensitive and specific methods for an efficient diagnosis of CPV infection. PMID:24334855

Investigation of spectral distribution and variation of irradiance with the passage time of CSI lamps which constitute a solar simulator; Solar simulator ni shiyosuru CSI lamp no supekutoru bunpu, hosha shodo no keiji henka ni kansuru chosa

Energy Technology Data Exchange (ETDEWEB)

Sugiyama, T; Yamada, T; Noguchi, T [Japan Quality Assurance Organization, Tokyo (Japan)

1997-11-25

Study was made on time-variation of the performance of CSI lamps for solar simulators. In order to accurately evaluate the standard heat collection performance of solar systems in a room, MITI installed an artificial solar light source in the Solar Techno-Center of Japan Quality Assurance Organization for trial use and evaluation. CSI lamp is superior in durability, and can simulate the solar light in the daytime. The light source is composed of 72 metal halide lamps of 1kW arranged in a plane of 3.5times3.5m. The study result on time-variation of a spectral distribution and irradiance by intermittent switching of lamps showed a sufficient durability of 2000h. To ensure the accuracy of a solar heat collector measurement system enough, periodic calibration is being carried out using reference goods. To ensure the reliability and stability for a switching system, periodic maintenance of a power source, stabilizer and electric system is also being carried out in addition to CSI lamps. The stable irradiance and accuracy are being kept by such maintenance and periodic exchange of lamps. 6 figs., 4 tabs.

Rapid and sensitive detection of Plesiomonas shigelloides by loop-mediated isothermal amplification of the hugA gene.

Directory of Open Access Journals (Sweden)

Shuang Meng

Full Text Available Plesiomonas shigelloides is one of the causative agents of human gastroenteritis, with increasing number of reports describing such infections in recent years. In this study, the hugA gene was chosen as the target to design loop-mediated isothermal amplification (LAMP assays for the rapid, specific, and sensitive detection of P. shigelloides. The performance of the assay with reference plasmids and spiked human stools as samples was evaluated and compared with those of quantitative PCR (qPCR. No false-positive results were observed for the 32 non-P. shigelloides strains used to evaluate assay specificity. The limit of detection for P. shigelloides was approximately 20 copies per reaction in reference plasmids and 5×10(3 CFU per gram in spiked human stool, which were more sensitive than the results of qPCR. When applied in human stool samples spiked with 2 low levels of P. shigelloides, the LAMP assays achieved accurate detection after 6-h enrichment. In conclusion, the LAMP assay developed in this study is a valuable method for rapid, cost-effective, and simple detection of P. shigelloides in basic clinical and field laboratories in the rural areas of China.

A Novel Loop-Mediated Isothermal Amplification (LAMP) Method for Detection of the JP2 Clone of Aggregatibacter actinomycetemcomitans in Subgingival Plaque

DEFF Research Database (Denmark)

Seki, M; Poulsen, Knud; Haubek, Dorte

2008-01-01

We developed a LAMP method that detects the JP2 clone of Aggregatibacter actinomycetemcomitans, which induces aggressive periodontitis in adolescents of North- and West-African descents. Being independent of special equipment this specific and sensitive method offers significant advantages...

Potential mercury emissions from fluorescent lamps production and obsolescence in mainland China.

Science.gov (United States)

Tan, Quanyin; Li, Jinhui

2016-01-01

The use of fluorescent lamps has expanded rapidly all over the world in recent years, because of their energy-saving capability. Consequently, however, mercury emissions from production, breakage, and discard of the lamps are drawing increasing concern from the public. This article focuses on evaluating the amount of mercury used for fluorescent lamp production, as well as the potential mercury emissions during production and breakage, in mainland China. It is expected to provide a comprehensive understanding about the risks present in the mercury from fluorescent lamps, and to know about the impacts of the policies on fluorescent lamps after their implementation. It is estimated that, in 2020, mercury consumption will be about 11.30-15.69 tonnes, a significant reduction of 34.9%-37.4% from that used in 2013, owing to improvement in mercury dosing dosage technology and tighter limitations on mercury content in fluorescent lamps. With these improvements, the amount of mercury remaining in fluorescent lamps and released during production is estimated to be 10.71-14.86 and 0.59-0.83 tonnes, respectively; the mercury released from waste fluorescent lamps is estimated to be about 5.37-7.59 tonnes. Also, a significant reduction to the mercury emission can be expected when a collection and treatment system is well established and conducted in the future. © The Author(s) 2015.

30 CFR 75.1703 - Portable electric lamps.

Science.gov (United States)

2010-07-01

... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Portable electric lamps. 75.1703 Section 75... HEALTH MANDATORY SAFETY STANDARDS-UNDERGROUND COAL MINES Miscellaneous § 75.1703 Portable electric lamps. [Statutory Provisions] Persons underground shall use only permissible electric lamps approved by the...

Loop-Mediated Isothermal Amplification-Lateral-Flow Dipstick (LAMP-LFD) to detect Toxoplasma gondii oocyst in ready-to-eat salad.

Science.gov (United States)

Lalle, Marco; Possenti, Alessia; Dubey, Jitender P; Pozio, Edoardo

2018-04-01

The apicomplexan parasite Toxoplasma gondii is the causative agent of toxoplasmosis, a foodborne zoonosis with a global distribution and estimated to cause up to 20% of the total foodborne disease burden in Europe. Association between T. gondii infection and the consumption of unwashed raw fruits and vegetables contaminated with oocysts has been reported and the increasing habit to eat pre-washed ready-to-eat salads poses a new potential risk for consumers. It is therefore important to trace the occurrence of potential contamination with this parasite to guarantee the safety of ready-to-eat vegetables. Detection of T. gondii in vegetables by molecular techniques has been achieved but low sensitivity (PCR) or expensive equipments (qPCR) limit routine applicability. Here, we describe the development and validation of a sensitive and robust method relying on a LAMP assay, targeting the 529 bp locus, to detect T. gondii oocysts down to 25 oocysts/50 g in ready-to-eat baby lettuce. The LAMP has been also adapted for a faster visualization of the result by a lateral flow dipstick chromatographic detection method. Copyright © 2017 Elsevier Ltd. All rights reserved.

Comparison of LAMP and PCR for molecular mass screening of sand flies for Leishmania martiniquensis infection.

Science.gov (United States)

Tiwananthagorn, Saruda; Kato, Hirotomo; Yeewa, Ranchana; Muengpan, Amontip; Polseela, Raxsina; Leelayoova, Saovanee

2017-02-01

Leishmaniasis caused by Leishmania martiniquensis infection has been reported in human and domestic animals of Martinique Island, Germany, Switzerland, USA, Myanmar and Thailand. The peculiar clinical features of disseminated cutaneous and visceral forms co-existence render the urgent need of specific diagnostic tool to identify the natural sand fly vectors for effective prevention and control strategies. Loop-mediated isothermal amplification (LAMP) of 18S rRNA gene as well as polymerase chain reaction (PCR) of minicircle kinetoplast DNA gene (PCR-mkDNA) have never been applied to detect L. martiniquensis and L. siamensis in sand fly vectors. The present study was aimed to validate malachite green-LAMP (MG-LAMP) and PCR-mkDNA techniques to detect L. martiniquensis in sand fly vectors, compared with the conventional PCR of internal transcribed spacer 1 (PCR-ITS1). We compared the validity of LAMP of 18S rRNA gene and PCR-mkDNA, to PCR-ITS1 in simulation model of L. martiniquensis infection in Sergentomyia gemmea sand flies. Attributable to the sensitivity and specificity, PCR-mkDNA was consecutively applied to detect L. martiniquensis in 380 female sand fly individuals captured in the newly identified affected region of Lamphun Province, Thailand. Results showed that PCR-mkDNA could detect at least one promastigote per sand fly, which was 10-time superior to LAMP and PCR-ITS1. In addition, PCR-mkDNA was more specific, able to differentiate L. martiniquensis from other viscerotropic Leishmania species, such as L. siamensis, L. (L.) donovani, and L. (L.) infantum. Consecutively, mass screening of L. martiniquensis in 380 female sand fly individuals by PCR-mkDNA was implemented in a new affected area of Thailand where a patient with leishmaniasis/HIV co-infection resides; however Leishmania DNA was undetected. In conclusion, PCR-mkDNA is a promising tool for molecular mass screening of L. martiniquensis infection in outbreak areas where several species of Leishmania

Comparison of LAMP and PCR for molecular mass screening of sand flies for Leishmania martiniquensis infection

Science.gov (United States)

Tiwananthagorn, Saruda; Kato, Hirotomo; Yeewa, Ranchana; Muengpan, Amontip; Polseela, Raxsina; Leelayoova, Saovanee

2017-01-01

BACKGROUND Leishmaniasis caused by Leishmania martiniquensis infection has been reported in human and domestic animals of Martinique Island, Germany, Switzerland, USA, Myanmar and Thailand. The peculiar clinical features of disseminated cutaneous and visceral forms co-existence render the urgent need of specific diagnostic tool to identify the natural sand fly vectors for effective prevention and control strategies. Loop-mediated isothermal amplification (LAMP) of 18S rRNA gene as well as polymerase chain reaction (PCR) of minicircle kinetoplast DNA gene (PCR-mkDNA) have never been applied to detect L. martiniquensis and L. siamensis in sand fly vectors. OBJECTIVE The present study was aimed to validate malachite green-LAMP (MG-LAMP) and PCR-mkDNA techniques to detect L. martiniquensis in sand fly vectors, compared with the conventional PCR of internal transcribed spacer 1 (PCR-ITS1). METHODS We compared the validity of LAMP of 18S rRNA gene and PCR-mkDNA, to PCR-ITS1 in simulation model of L. martiniquensis infection in Sergentomyia gemmea sand flies. Attributable to the sensitivity and specificity, PCR-mkDNA was consecutively applied to detect L. martiniquensis in 380 female sand fly individuals captured in the newly identified affected region of Lamphun Province, Thailand. FINDINGS AND MAIN CONCLUSIONS Results showed that PCR-mkDNA could detect at least one promastigote per sand fly, which was 10-time superior to LAMP and PCR-ITS1. In addition, PCR-mkDNA was more specific, able to differentiate L. martiniquensis from other viscerotropic Leishmania species, such as L. siamensis, L. (L.) donovani, and L. (L.) infantum. Consecutively, mass screening of L. martiniquensis in 380 female sand fly individuals by PCR-mkDNA was implemented in a new affected area of Thailand where a patient with leishmaniasis/HIV co-infection resides; however Leishmania DNA was undetected. In conclusion, PCR-mkDNA is a promising tool for molecular mass screening of L. martiniquensis

Fluorescent discharge lamp

Science.gov (United States)

Mukai, E.; Otsuka, H.; Nomi, K.; Honmo, I.

1982-01-01

A rapidly illuminating fluorescent lamp 1,200 mm long and 32.5 mm in diameter with an interior conducting strip which is compatible with conventional fixtures and ballasts is described. The fluorescent lamp is composed of a linear glass tube, electrodes sealed at both ends, mercury and raregas sealed in the glass tube, a fluorescent substance clad on the inner walls of the glass tube, and a clad conducting strip extending the entire length of the glass tube in the axial direction on the inner surface of the tube.

BAD-LAMP controls TLR9 trafficking and signalling in human plasmacytoid dendritic cells.

Science.gov (United States)

Combes, Alexis; Camosseto, Voahirana; N'Guessan, Prudence; Argüello, Rafael J; Mussard, Julie; Caux, Christophe; Bendriss-Vermare, Nathalie; Pierre, Philippe; Gatti, Evelina

2017-10-13

Toll-like receptors (TLR) are essential components of the innate immune system. Several accessory proteins, such as UNC93B1, are required for transport and activation of nucleic acid sensing Toll-like receptors in endosomes. Here, we show that BAD-LAMP (LAMP5) controls TLR9 trafficking to LAMP1 + late endosomes in human plasmacytoid dendritic cells (pDC), leading to NF-κB activation and TNF production upon DNA detection. An inducible VAMP3 +/ LAMP2 +/ LAMP1 - endolysosome compartment exists in pDCs from which TLR9 activation triggers type I interferon expression. BAD-LAMP-silencing enhances TLR9 retention in this compartment and consequent downstream signalling events. Conversely, sustained BAD-LAMP expression in pDCs contributes to their lack of type I interferon production after exposure to a TGF-β-positive microenvironment or isolation from human breast tumours. Hence, BAD-LAMP limits interferon expression in pDCs indirectly, by promoting TLR9 sorting to late endosome compartments at steady state and in response to immunomodulatory cues.TLR9 is highly expressed by plasmacytoid dendritic cells and detects nucleic acids, but to discriminate between host and microbial nucleic acids TLR9 is sorted into different endosomal compartments. Here the authors show that BAD-LAMP limits type 1 interferon responses by sorting TLR9 to late endosomal compartments.

Collisional and radiative processes in fluorescent lamps

International Nuclear Information System (INIS)

Lister, Graeme G.

2003-01-01

Since electrode life is the major limiting factor in operating fluorescent lamps, many lighting companies have introduced 'electrodeless' fluorescent lamps, using inductively coupled discharges. These lamps often operate at much higher power loadings than standard lamps and numerical models have not been successful in reproducing experimental measurements in the parameter ranges of interest. A comprehensive research program was undertaken to study the fundamental physical processes of these discharges, co-funded by the Electric Power Research Institute (EPRI) and OSRAM SYLVANIA under the name of ALITE. The program included experiments and modeling of radiation transport, computations of electron-atom and atom-atom cross sections and the first comprehensive power balance studies of a highly loaded fluorescent lamp. Results from the program and their importance to the understanding of the physics of fluorescent lamps are discussed, with particular emphasis on the important collisional and radiative processes. Comparisons between results of experimental measurements and numerical models are presented

Phase out of incandescent lamps

Energy Technology Data Exchange (ETDEWEB)

NONE

2010-07-01

Since early 2007 almost all OECD and many non-OECD governments have announced policies aimed at phasing-out incandescent lighting within their jurisdictions. This study considers the implications of these policy developments in terms of demand for regulatory compliant lamps and the capacity and motivation of the lamp industry to produce efficient lighting products in sufficient volume to meet future demand. To assess these issues, it reviews the historic international screw-based lamp market, describes the status of international phase-out policies and presents projections of anticipated market responses to regulatory requirements to determine future demand for CFLs.

Parametric amplification and bidirectional invisibility in PT -symmetric time-Floquet systems

Science.gov (United States)

Koutserimpas, Theodoros T.; Alù, Andrea; Fleury, Romain

2018-01-01

Parity-time (PT )-symmetric wave devices, which exploit balanced interactions between material gain and loss, exhibit extraordinary properties, including lasing and flux-conserving scattering processes. In a seemingly different research field, periodically driven systems, also known as time-Floquet systems, have been widely studied as a relevant platform for reconfigurable active wave control and manipulation. In this article, we explore the connection between PT -symmetry and parametric time-Floquet systems. Instead of relying on material gain, we use parametric amplification by considering a time-periodic modulation of the refractive index at a frequency equal to twice the incident signal frequency. We show that the scattering from a simple parametric slab, whose dynamics follows the Mathieu equation, can be described by a PT -symmetric scattering matrix, whose PT -breaking threshold corresponds to the Mathieu instability threshold. By combining different parametric slabs modulated out of phase, we create PT -symmetric time-Floquet systems that feature exceptional scattering properties, such as coherent perfect absorption (CPA)-laser operation and bidirectional invisibility. These bidirectional properties, rare for regular PT -symmetric systems, are related to a compensation of parametric amplification due to multiple scattering between two parametric systems modulated with a phase difference.

30 CFR 57.12035 - Weatherproof lamp sockets.

Science.gov (United States)

2010-07-01

... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Weatherproof lamp sockets. 57.12035 Section 57.12035 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND NONMETAL... Surface and Underground § 57.12035 Weatherproof lamp sockets. Lamp sockets shall be of a weatherproof type...

30 CFR 56.12035 - Weatherproof lamp sockets.

Science.gov (United States)

2010-07-01

... 30 Mineral Resources 1 2010-07-01 2010-07-01 false Weatherproof lamp sockets. 56.12035 Section 56.12035 Mineral Resources MINE SAFETY AND HEALTH ADMINISTRATION, DEPARTMENT OF LABOR METAL AND NONMETAL....12035 Weatherproof lamp sockets. Lamp sockets shall be of a weatherproof type where they are exposed to...

MINIX4RT: a real-time operating system based on MINIX

OpenAIRE

Pessolani, Pablo Andrés

2006-01-01

Tanenbaum's MINIX Operating System was extended with a Real-Time microkernel and services to conform MINIX4RT, a Real-Time Operating System for academic uses that includes more flexible Interprocess Communications facilities supporting basic priority inheritance protocol, a fixed priority scheduler, timer and event driven interrupt management, statistics and Real-Time metrics gathering keeping backward compatibility with standard MINIX versions. Facultad de Informática

Lamp with a truncated reflector cup

Science.gov (United States)

Li, Ming; Allen, Steven C.; Bazydola, Sarah; Ghiu, Camil-Daniel

2013-10-15

A lamp assembly, and method for making same. The lamp assembly includes first and second truncated reflector cups. The lamp assembly also includes at least one base plate disposed between the first and second truncated reflector cups, and a light engine disposed on a top surface of the at least one base plate. The light engine is configured to emit light to be reflected by one of the first and second truncated reflector cups.

47 CFR 17.54 - Rated lamp voltage.

Science.gov (United States)

2010-10-01

... 47 Telecommunication 1 2010-10-01 2010-10-01 false Rated lamp voltage. 17.54 Section 17.54... voltage. To insure the necessary lumen output by obstruction lights, the rated voltage of incandescent lamps used shall correspond to be within 3 percent higher than the voltage across the lamp socket during...

Implementation of antimicrobial peptides for sample preparation prior to nucleic acid amplification in point-of-care settings.

Science.gov (United States)

Krõlov, Katrin; Uusna, Julia; Grellier, Tiia; Andresen, Liis; Jevtuševskaja, Jekaterina; Tulp, Indrek; Langel, Ülo

2017-12-01

A variety of sample preparation techniques are used prior to nucleic acid amplification. However, their efficiency is not always sufficient and nucleic acid purification remains the preferred method for template preparation. Purification is difficult and costly to apply in point-of-care (POC) settings and there is a strong need for more robust, rapid, and efficient biological sample preparation techniques in molecular diagnostics. Here, the authors applied antimicrobial peptides (AMPs) for urine sample preparation prior to isothermal loop-mediated amplification (LAMP). AMPs bind to many microorganisms such as bacteria, fungi, protozoa and viruses causing disruption of their membrane integrity and facilitate nucleic acid release. The authors show that incubation of E. coli with antimicrobial peptide cecropin P1 for 5 min had a significant effect on the availability of template DNA compared with untreated or even heat treated samples resulting in up to six times increase of the amplification efficiency. These results show that AMPs treatment is a very efficient sample preparation technique that is suitable for application prior to nucleic acid amplification directly within biological samples. Furthermore, the entire process of AMPs treatment was performed at room temperature for 5 min thereby making it a good candidate for use in POC applications.

CALiPER Report 21.3. Cost Effectiveness of Linear (T8) LED Lamps

Energy Technology Data Exchange (ETDEWEB)

None

2014-05-01

Meeting performance expectations is important for driving adoption of linear LED lamps, but cost-effectiveness may be an overriding factor in many cases. Linear LED lamps cost more initially than fluorescent lamps, but energy and maintenance savings may mean that the life-cycle cost is lower. This report details a series of life-cycle cost simulations that compared a two-lamp troffer using LED lamps (38 W total power draw) or fluorescent lamps (51 W total power draw) over a 10-year study period. Variables included LED system cost ($40, $80, or $120), annual operating hours (2,000 hours or 4,000 hours), LED installation time (15 minutes or 30 minutes), and melded electricity rate ($0.06/kWh, $0.12/kWh, $0.18/kWh, or $0.24/kWh). A full factorial of simulations allows users to interpolate between these values to aid in making rough estimates of economic feasibility for their own projects. In general, while their initial cost premium remains high, linear LED lamps are more likely to be cost-effective when electric utility rates are higher than average and hours of operation are long, and if their installation time is shorter.

CALiPER Report 21.3: Cost-Effectiveness of Linear (T8) LED Lamps

Energy Technology Data Exchange (ETDEWEB)

Miller, Naomi J.; Perrin, Tess E.; Royer, Michael P.

2014-05-27

Meeting performance expectations is important for driving adoption of linear LED lamps, but cost-effectiveness may be an overriding factor in many cases. Linear LED lamps cost more initially than fluorescent lamps, but energy and maintenance savings may mean that the life-cycle cost is lower. This report details a series of life-cycle cost simulations that compared a two-lamp troffer using LED lamps (38 W total power draw) or fluorescent lamps (51 W total power draw) over a 10-year study period. Variables included LED system cost ($40, $80, or $120), annual operating hours (2,000 hours or 4,000 hours), LED installation time (15 minutes or 30 minutes), and melded electricity rate ($0.06/kWh, $0.12/kWh, $0.18/kWh, or $0.24/kWh). A full factorial of simulations allows users to interpolate between these values to aid in making rough estimates of economic feasibility for their own projects. In general, while their initial cost premium remains high, linear LED lamps are more likely to be cost-effective when electric utility rates are higher than average and hours of operation are long, and if their installation time is shorter.

Dispenser printed electroluminescent lamps on textiles for smart fabric applications

Science.gov (United States)

de Vos, Marc; Torah, Russel; Tudor, John

2016-04-01

Flexible electroluminescent (EL) lamps are fabricated onto woven textiles using a novel dispenser printing process. Dispenser printing utilizes pressurized air to deposit ink onto a substrate through a syringe and nozzle. This work demonstrates the first use of this technology to fabricate EL lamps. The luminance of the dispenser printed EL lamps is compared to screen-printed EL lamps, both printed on textile, and also commercial EL lamps on polyurethane film. The dispenser printed lamps are shown to have a 1.5 times higher luminance than the best performing commercially available lamp, and have a comparable performance to the screen-printed lamps.

Dispenser printed electroluminescent lamps on textiles for smart fabric applications

International Nuclear Information System (INIS)

De Vos, Marc; Torah, Russel; Tudor, John

2016-01-01

Flexible electroluminescent (EL) lamps are fabricated onto woven textiles using a novel dispenser printing process. Dispenser printing utilizes pressurized air to deposit ink onto a substrate through a syringe and nozzle. This work demonstrates the first use of this technology to fabricate EL lamps. The luminance of the dispenser printed EL lamps is compared to screen-printed EL lamps, both printed on textile, and also commercial EL lamps on polyurethane film. The dispenser printed lamps are shown to have a 1.5 times higher luminance than the best performing commercially available lamp, and have a comparable performance to the screen-printed lamps. (paper)

Lamps and lighting

CERN Document Server

Cayless, MA; Marsden, A M

2012-01-01

This book is a comprehensive guide to the theory and practice of lighting. Covering the physics of light production, light sources, circuits and a wide variety of lighting applications, it is both suitable as a detailed textbook and as thoroughly practical guide for practising lighting engineers. This fourth edition of Lamps and Lighting has been completely updated with new chapters on the latest lamp technology and applications. The editors ahve called upon a wide range of expertise and as a result many sections have been broadened to include both European and US practice.The book begins with

Commercial Slit-Lamp Anterior Segment Photography versus Digital Compact Camera Mounted on a Standard Slit-Lamp with an Adapter.

Science.gov (United States)

Oliphant, Huw; Kennedy, Alasdair; Comyn, Oliver; Spalton, David J; Nanavaty, Mayank A

2018-06-16

To compare slit lamp mounted cameras (SLC) versus digital compact camera (DCC) with slit-lamp adaptor when used by an inexperienced technician. In this cross sectional study, where posterior capsule opacification (PCO) was used as a comparator, patients were consented for one photograph with SLC and two with DCC (DCC1 and DCC2), with a slit-lamp adaptor. An inexperienced clinic technician, who took all the photographs and masked the images, recruited one eye of each patient. Images were graded for PCO using ECPO2000 software by two independent masked graders. Repeatability between DCC1 & DCC2 and limits-of-agreement between SLC and DCC1 mounted on slit-lamp with an adaptor were assessed. Coefficient-of-repeatability and Bland-Altmann plots were analyzed. Seventy-two patients (eyes) were recruited in the study. First 9 patients (eyes) were excluded due to unsatisfactory image quality from both the systems. Mean EPCO score for SLC was 2.28 (95% CI: 2.09 -2.45), for DCC1 was 2.28 (95% CI: 2.11-2.45), and for the DCC2 was 2.11 (95% CI: 2.11-2.45). There was no significant difference in EPCO scores between SLC Vs. DCC1 (p = 0.98) and between DCC1 and DCC 2 (p = 0.97). Coefficient of repeatability between DCC images was 0.42, and the coefficient of repeatability between DCC and SLC was 0.58. DCC on slit-lamp with an adaptor is comparable to a SLC. There is an initial learning curve, which is similar for both for an inexperienced person. This opens up the possibility for low cost anterior segment imaging in the clinical, research and teaching settings.

MET amplification, expression, and exon 14 mutations in colorectal adenocarcinoma.

Science.gov (United States)

Zhang, Meng; Li, Guichao; Sun, Xiangjie; Ni, Shujuan; Tan, Cong; Xu, Midie; Huang, Dan; Ren, Fei; Li, Dawei; Wei, Ping; Du, Xiang

2018-04-08

MET amplification, expression, and splice mutations at exon 14 result in dysregulation of the MET signaling pathway. The aim of this study was to identify the relationship between MET amplification, protein or mRNA expression, and mutations in colorectal cancer (CRC). MET immunohistochemistry (IHC) was used for MET protein expression analysis and fluorescence in situ hybridization (FISH) was used for MET amplification detection. Both analyses were performed in tissue microarrays (TMA) containing 294 of colorectal adenocarcinoma tissue samples and 131 samples of adjacent normal epithelial tissue. MET mRNA expression was examined by real-time quantitative polymerase chain reaction (qRT-PCR) in 72 fresh colorectal adenocarcinoma tissue samples and adjacent normal colon tissue. PCR sequencing was performed to screen for MET exon 14 splice mutations in 59 fresh CRC tissue samples. Our results showed that MET protein expression was higher in colorectal tumor tissue than in adjacent normal intestinal epithelium. Positive MET protein expression was associated with significantly poorer overall survival (OS) and disease-free survival (DFS). Multivariate analysis revealed that positive MET protein expression was an independent risk factor for DFS, but not for OS. MET mRNA expression was upregulated in tumor tissues compared with the adjacent normal tissues. The incidence of MET amplification was 4.4%. None of the patients was positive for MET mutation. Collectively, MET was overexpressed in colorectal adenocarcinoma, and its positive protein expression predicted a poorer outcome in CRC patients. Furthermore, according to our results, MET amplification and 14 exon mutation are extremely rare events in colorectal adenocarcinoma. Copyright © 2018. Published by Elsevier Inc.

Penerapan Metode Diagnosis Cepat Virus Avian Influenza H5N1 dengan Metode Single Step Multiplex RT-PCR

Directory of Open Access Journals (Sweden)

Aris Haryanto

2010-12-01

Full Text Available Avian influenza (AI virus is a segmented single stranded (ss RNA virus with negative polarity andbelong to the Orthomyxoviridae family. Diagnose of AI virus can be performed using conventional methodsbut it has low sensitivity and specificity. The objective of the research was to apply rapid, precise, andaccurate diagnostic method for AI virus and also to determine its type and subtype based on the SingleStep Multiplex Reverse Transcriptase-Polymerase Chain Reaction targeting M, H5, and N1 genes. In thismethod M, H5 and NI genes were simultaneously amplified in one PCR tube. The steps of this researchconsist of collecting viral RNAs from 10 different AI samples originated from Maros Disease InvestigationCenter during 2007. DNA Amplification was conducted by Simplex RT-PCR using M primer set. Then, bysingle step multiplex RT-PCR were conducted simultaneously using M, H5 and N1 primers set. The RTPCRproducts were then separated on 1.5% agarose gel, stained by ethidum bromide and visualized underUV transilluminator. Results showed that 8 of 10 RNA virus samples could be amplified by Simplex RTPCRfor M gene which generating a DNA fragment of 276 bp. Amplification using multiplex RT-PCRmethod showed two of 10 samples were AI positive using multiplex RT-PCR, three DNA fragments weregenerated consisting of 276 bp for M gene, 189 bp for H5 gene, and 131 bp for N1. In this study, rapid andeffective diagnosis method for AI virus can be conducted by using simultaneous Single Step Multiplex RTPCR.By this technique type and subtype of AI virus, can also be determined, especially H5N1.

Ultrafast Capillary Electrophoresis Isolation of DNA Aptamer for the PCR Amplification-Based Small Analyte Sensing

Directory of Open Access Journals (Sweden)

Emmanuelle eFiore

2015-08-01

Full Text Available Here, we report a new homogeneous DNA amplification-based aptamer assay for small analyte sensing. The aptamer of adenosine chosen as the model analyte was split into two fragments able to assemble in the presence of target. Primers were introduced at extremities of one fragment in order to generate the amplifiable DNA component. The amount of amplifiable fragment was quantifiable by Real-Time Polymerase Chain Reaction (RT-PCR amplification and directly reliable on adenosine concentration. This approach combines the very high separation efficiency and the homogeneous format (without immobilization of capillary electrophoresis and the sensitivity of real time PCR amplification. An ultrafast isolation of target-bound split aptamer (60 s was developed by designing a capillary electrophoresis input/ouput scheme. Such method was successfully applied to the determination of adenosine with a LOD of 1 µM.

Evidence of high-elevation amplification versus Arctic amplification.

Science.gov (United States)

Wang, Qixiang; Fan, Xiaohui; Wang, Mengben

2016-01-12

Elevation-dependent warming in high-elevation regions and Arctic amplification are of tremendous interest to many scientists who are engaged in studies in climate change. Here, using annual mean temperatures from 2781 global stations for the 1961-2010 period, we find that the warming for the world's high-elevation stations (>500 m above sea level) is clearly stronger than their low-elevation counterparts; and the high-elevation amplification consists of not only an altitudinal amplification but also a latitudinal amplification. The warming for the high-elevation stations is linearly proportional to the temperature lapse rates along altitudinal and latitudinal gradients, as a result of the functional shape of Stefan-Boltzmann law in both vertical and latitudinal directions. In contrast, neither altitudinal amplification nor latitudinal amplification is found within the Arctic region despite its greater warming than lower latitudes. Further analysis shows that the Arctic amplification is an integrated part of the latitudinal amplification trend for the low-elevation stations (≤500 m above sea level) across the entire low- to high-latitude Northern Hemisphere, also a result of the mathematical shape of Stefan-Boltzmann law but only in latitudinal direction.

Loop-mediated isothermal amplification assay for rapid detection of Streptococcus agalactiae (group B streptococcus) in vaginal swabs - a proof of concept study.

Science.gov (United States)

McKenna, James Patrick; Cox, Ciara; Fairley, Derek John; Burke, Rachael; Shields, Michael D; Watt, Alison; Coyle, Peter Valentine

2017-03-01

Neonatal sepsis caused by Streptococcus agalactiae [group B streptococcus (GBS)] is a life-threatening condition, which is preventable if colonized mothers are identified and given antibiotic prophylaxis during labour. Conventional culture is time consuming and unreliable, and many available non-culture diagnostics are too complex to implement routinely at point of care. Loop-mediated isothermal amplification (LAMP) is a method that, enables the rapid and specific detection of target nucleic acid sequences in clinical materials without the requirement for extensive sample preparation. A prototype LAMP assay targeting GBS sip gene is described. The assay was 100 % specific for GBS, with a limit of detection of 14 genome copies per reaction. The clinical utility of the LAMP assay for rapid direct molecular detection of GBS was determined by testing a total of 157 vaginal swabs with minimal sample processing using a rapid lysis solution. Compared to a reference quantitative real-time PCR assay, the direct LAMP protocol had a sensitivity and specificity of 95.4 and 100 %, respectively, with positive and negative predictive values of 100 and 98.3 %, respectively. Positive and negative likelihood ratios were infinity and 0.05, respectively. The direct LAMP method required a mean time of 45 min from the receipt of a swab to generation of a confirmed result, compared to 2 h 30 min for the reference quantitative real-time PCR test. The direct LAMP protocol described is easy to perform, facilitating rapid and accurate detection of GBS in vaginal swabs. This test has a potential for use at point of care.

Rapid colorimetric assay for detection of Listeria monocytogenes in food samples using LAMP formation of DNA concatemers and gold nanoparticle-DNA probe complex

Science.gov (United States)

Wachiralurpan, Sirirat; Sriyapai, Thayat; Areekit, Supatra; Sriyapai, Pichapak; Augkarawaritsawong, Suphitcha; Santiwatanakul, Somchai; Chansiri, Kosum

2018-04-01

ABSTRACT Listeria monocytogenes is a major foodborne pathogen of global health concern. Herein, the rapid diagnosis of L. monocytogenes has been achieved using loop-mediated isothermal amplification (LAMP) based on the phosphatidylcholine-phospholipase C gene (plcB). Colorimetric detection was then performed through the formation of DNA concatemers and a gold nanoparticle/DNA probe complex (GNP/DNA probe). The overall detection process was accomplished within approximately 1 h with no need for complicated equipment. The limits of detection for L. monocytogenes in the forms of purified genomic DNA and pure culture were 800 fg and 2.82 CFU mL-1, respectively. No cross reactions were observed from closely related bacteria species. The LAMP-GNP/DNA probe assay was applied to the detection of 200 raw chicken meat samples and compared to routine standard methods. The data revealed that the specificity, sensitivity and accuracy were 100%, 90.20% and 97.50%, respectively. The present assay was 100% in conformity with LAMP-agarose gel electrophoresis assay. Five samples that were negative by both assays appeared to have the pathogen at below the level of detection. The assay can be applied as a rapid direct screening method for L. monocytogenes.

Evaluation of colorimetric loop-mediated isothermal amplification assay for visual detection of Streptococcus agalactiae and Streptococcus iniae in tilapia.

Science.gov (United States)

Suebsing, R; Kampeera, J; Tookdee, B; Withyachumnarnkul, B; Turner, W; Kiatpathomchai, W

2013-10-01

Streptococcus agalactiae and Strep. iniae are bacterial pathogens that cause streptococcosis in many fish species. An accelerated colorimetric loop-mediated isothermal amplification (LAMP) assay with pre-addition of calcein was established, and the transmission and detection of Strep. agalactiae and Strep. iniae in tilapia under natural aquatic environment were investigated. A positive reaction was observed by a colour change from orange to green through the naked eyes after completion at 63°C for 30 min with 10 times higher sensitivity than that of nested PCR assays and without cross-amplification with other fish bacterial pathogens. All sample types of Nile and red tilapia (broodstock, fertilized egg, fry) were Strep. agalactiae- and Strep. iniae positive by this new method, implying that they could be vertically transmitted. With its application for screening broodstock and fry before stocking and for monitoring fish health in grow-out ponds, the method would become very useful in fish farming industry. The application of colorimetric LAMP with pre-addition of calcein offers simple, rapid and sensitive technique with applicability for small field laboratories. This technique explored the possible vertical transmission mode of Strep. agalactiae and Strep. iniae under natural aquatic environment. It could be such preliminary data provided for the screening broodstock before breeding and/or the specific-pathogen-free production. © 2013 The Society for Applied Microbiology.

Lamp reliability studies for improved satellite rubidium frequency standard

Science.gov (United States)

Frueholz, R. P.; Wun-Fogle, M.; Eckert, H. U.; Volk, C. H.; Jones, P. F.

1982-01-01

In response to the premature failure of Rb lamps used in Rb atomic clocks onboard NAVSTAR GPS satellites experimental and theoretical investigations into their failure mechanism were initiated. The primary goal of these studies is the development of an accelerated life test for future GPS lamps. The primary failure mechanism was identified as consumption of the lamp's Rb charge via direct interaction between Rb and the lamp's glass surface. The most effective parameters to accelerate the interaction between the Rb and the glass are felt to be RF excitation power and lamp temperature. Differential scanning calorimetry is used to monitor the consumption of Rb within a lamp as a function of operation time. This technique yielded base line Rb consumption data for GPS lamps operating under normal conditions.

NASA SPoRT Initialization Datasets for Local Model Runs in the Environmental Modeling System

Science.gov (United States)

Case, Jonathan L.; LaFontaine, Frank J.; Molthan, Andrew L.; Carcione, Brian; Wood, Lance; Maloney, Joseph; Estupinan, Jeral; Medlin, Jeffrey M.; Blottman, Peter; Rozumalski, Robert A.

2011-01-01

The NASA Short-term Prediction Research and Transition (SPoRT) Center has developed several products for its National Weather Service (NWS) partners that can be used to initialize local model runs within the Weather Research and Forecasting (WRF) Environmental Modeling System (EMS). These real-time datasets consist of surface-based information updated at least once per day, and produced in a composite or gridded product that is easily incorporated into the WRF EMS. The primary goal for making these NASA datasets available to the WRF EMS community is to provide timely and high-quality information at a spatial resolution comparable to that used in the local model configurations (i.e., convection-allowing scales). The current suite of SPoRT products supported in the WRF EMS include a Sea Surface Temperature (SST) composite, a Great Lakes sea-ice extent, a Greenness Vegetation Fraction (GVF) composite, and Land Information System (LIS) gridded output. The SPoRT SST composite is a blend of primarily the Moderate Resolution Imaging Spectroradiometer (MODIS) infrared and Advanced Microwave Scanning Radiometer for Earth Observing System data for non-precipitation coverage over the oceans at 2-km resolution. The composite includes a special lake surface temperature analysis over the Great Lakes using contributions from the Remote Sensing Systems temperature data. The Great Lakes Environmental Research Laboratory Ice Percentage product is used to create a sea-ice mask in the SPoRT SST composite. The sea-ice mask is produced daily (in-season) at 1.8-km resolution and identifies ice percentage from 0 100% in 10% increments, with values above 90% flagged as ice.

A one-step multiplex RT-PCR assay for simultaneous detection of four viruses that infect peach.

Science.gov (United States)

Yu, Y; Zhao, Z; Jiang, D; Wu, Z; Li, S

2013-10-01

A multiplex reverse transcription polymerase chain reaction (mRT-PCR) assay was developed to enable the simultaneous detection and differentiation of four viruses that infect peach, namely Apple chlorotic leaf spot virus (ACLSV), Cherry green ring mottle virus (CGRMV), Prunus necrotic ringspot virus (PNRSV) and Apricot pseudo-chlorotic leaf spot virus (APCLSV). In this study, four pairs of primers, one specific for each virus, were designed; the corresponding PCR products were 632, 439, 346 and 282 bp in length for ACLSV, CGRMV, PNRSV and APCLSV, respectively, and the fragments could be distinguished clearly by agarose gel electrophoresis. The sensitivity and specificity of the method were tested using individual RT-PCR and enzyme-linked immunosorbent assay (ELISA), and the identity of the RT-PCR amplification products was also confirmed by DNA sequencing. The results of RT-PCR and ELISA, along with batch detection using samples collected from peach orchards, revealed that this rapid and simple technique is an effective way to identify the four viruses simultaneously. The mRT-PCR assay described in this study was developed for the simultaneous detection of four peach viruses from infected peach samples is reliable and sensitive. In contrast to conventional uniplex RT-PCR, mRT-PCR is more efficient, reducing costs, time and handling when testing large numbers of samples. This rapid and simple method is useful for large-scale surveys of viruses that infect peach. © 2013 The Society for Applied Microbiology.

Determining putative vectors of the Bogia Coconut Syndrome phytoplasma using loop-mediated isothermal amplification of single-insect feeding media

Science.gov (United States)

Lu, Hengyu; Wilson, Bree A. L.; Ash, Gavin J.; Woruba, Sharon B.; Fletcher, Murray J.; You, Minsheng; Yang, Guang; Gurr, Geoff M.

2016-01-01

Phytoplasmas are insect vectored mollicutes responsible for disease in many economically important crops. Determining which insect species are vectors of a given phytoplasma is important for managing disease but is methodologically challenging because disease-free plants need to be exposed to large numbers of insects, often over many months. A relatively new method to detect likely transmission involves molecular testing for phytoplasma DNA in sucrose solution that insects have fed upon. In this study we combined this feeding medium method with a loop-mediated isothermal amplification (LAMP) assay to study 627 insect specimens of 11 Hemiptera taxa sampled from sites in Papua New Guinea affected by Bogia coconut syndrome (BCS). The LAMP assay detected phytoplasma DNA from the feeding solution and head tissue of insects from six taxa belonging to four families: Derbidae, Lophopidae, Flatidae and Ricaniidae. Two other taxa yielded positives only from the heads and the remainder tested negative. These results demonstrate the utility of combining single-insect feeding medium tests with LAMP assays to identify putative vectors that can be the subject of transmission tests and to better understand phytoplasma pathosystems. PMID:27786249

A linear accelerator power amplification system for high gradient structure research

International Nuclear Information System (INIS)

Haimson, J.; Mecklenburg, B.

1999-01-01

The ongoing development of linear collider high power RF sources and pulse compression systems has resulted in substantial progress towards a goal of providing a peak RF power level of approximately 250 MW at the input of the accelerator structure. While the immediate development and the high power testing of specialized waveguide components required for power transmission at these high levels have proceeded expeditiously due to the availability of resonant ring systems, the testing of high gradient accelerator structures at very high power levels, and the investigation of coupler cavity RF breakdown problems have, typically, been curtailed due to the unavailability of suitable 200 to 300 MW RF test facilities. We describe herein a compact, high peak power amplification system based on a dual hybrid bridge configuration that avoids the need for power splitters at the accelerator dual feed couplers, and also provides a convenient interface for installing high gradient accelerator test structures. Design parameters are presented for a proposed power amplification system that makes use of a 75 MW, 1/2 μs flat-top RF source to produce 280 MW, 1/4 μs flat-top power for testing dual feed TW experimental accelerator sections

Optimized positioning of autonomous surgical lamps

Science.gov (United States)

Teuber, Jörn; Weller, Rene; Kikinis, Ron; Oldhafer, Karl-Jürgen; Lipp, Michael J.; Zachmann, Gabriel

2017-03-01

We consider the problem of finding automatically optimal positions of surgical lamps throughout the whole surgical procedure, where we assume that future lamps could be robotized. We propose a two-tiered optimization technique for the real-time autonomous positioning of those robotized surgical lamps. Typically, finding optimal positions for surgical lamps is a multi-dimensional problem with several, in part conflicting, objectives, such as optimal lighting conditions at every point in time while minimizing the movement of the lamps in order to avoid distractions of the surgeon. Consequently, we use multi-objective optimization (MOO) to find optimal positions in real-time during the entire surgery. Due to the conflicting objectives, there is usually not a single optimal solution for such kinds of problems, but a set of solutions that realizes a Pareto-front. When our algorithm selects a solution from this set it additionally has to consider the individual preferences of the surgeon. This is a highly non-trivial task because the relationship between the solution and the parameters is not obvious. We have developed a novel meta-optimization that considers exactly this challenge. It delivers an easy to understand set of presets for the parameters and allows a balance between the lamp movement and lamp obstruction. This metaoptimization can be pre-computed for different kinds of operations and it then used by our online optimization for the selection of the appropriate Pareto solution. Both optimization approaches use data obtained by a depth camera that captures the surgical site but also the environment around the operating table. We have evaluated our algorithms with data recorded during a real open abdominal surgery. It is available for use for scientific purposes. The results show that our meta-optimization produces viable parameter sets for different parts of an intervention even when trained on a small portion of it.

Evaluation of commercial kit based on loop-mediated isothermal amplification for rapid detection of low levels of uninjured and injured Salmonella on duck meat, bean sprouts, and fishballs in Singapore.

Science.gov (United States)

Lim, Hazel Sin Yue; Zheng, Qianwang; Miks-Krajnik, Marta; Turner, Matthew; Yuk, Hyun-Gyun

2015-06-01

The objective of this study was to evaluate performance of the commercial kit based on loop-mediated isothermal amplification (LAMP) in comparison with the International Organization for Standardization method for detecting uninjured and sublethally injured Salmonella cells artificially inoculated at levels of 10(0) and 10(1) CFU/25 g on raw duck wing, raw mung bean sprouts, and processed fishballs. Injured cells were prepared by a heat treatment for duck wings and fishball samples and a chlorine treatment for bean sprout samples. Additionally, a validation study was performed on naturally contaminated food samples sold in Singapore. A total of 110 samples of each commodity were analyzed in this study. Regardless of inoculum levels, the detection by the commercial LAMP kit showed 100% sensitivity and specificity for both inoculated and uninoculated samples compared with the International Organization for Standardization method, with the exception of bean sprout samples. Only 20% of bean sprout samples inoculated with 10(0) CFU/25 g injured Salmonella cells were positive by using the commercial LAMP-based kit. However, all negative samples became positive following a secondary enrichment in Rappaport-Vassiliadis medium with soy broth or after concentration by centrifugation. These results suggest that secondary enrichment or centrifugation should be considered as an additional step to increase the sensitivity of the commercial LAMP-based kit with low numbers of injured target cells in samples with high background microflora (such as mung bean sprouts). The validation study also showed that the commercial LAMP-based kit provided 91% sensitivity and 95% specificity for naturally contaminated samples. Thus, this study demonstrates that the commercial LAMP-based kit might be a cost-effective method, as this system could provide rapid, accurate detection of both uninjured and injured Salmonella cells on raw duck wings, raw mung bean sprouts, and processed fishballs in

Tumor target amplification: Implications for nano drug delivery systems.

Science.gov (United States)

Seidi, Khaled; Neubauer, Heidi A; Moriggl, Richard; Jahanban-Esfahlan, Rana; Javaheri, Tahereh

2018-04-10

Tumor cells overexpress surface markers which are absent from normal cells. These tumor-restricted antigenic signatures are a fundamental basis for distinguishing on-target from off-target cells for ligand-directed targeting of cancer cells. Unfortunately, tumor heterogeneity impedes the establishment of a solid expression pattern for a given target marker, leading to drastic changes in quality (availability) and quantity (number) of the target. Consequently, a subset of cancer cells remains untargeted during the course of treatment, which subsequently promotes drug-resistance and cancer relapse. Since target inefficiency is only problematic for cancer treatment and not for treatment of other pathological conditions such as viral/bacterial infections, target amplification or the generation of novel targets is key to providing eligible antigenic markers for effective targeted therapy. This review summarizes the limitations of current ligand-directed targeting strategies and provides a comprehensive overview of tumor target amplification strategies, including self-amplifying systems, dual targeting, artificial markers and peptide modification. We also discuss the therapeutic and diagnostic potential of these approaches, the underlying mechanism(s) and established methodologies, mostly in the context of different nanodelivery systems, to facilitate more effective ligand-directed cancer cell monitoring and targeting. Copyright © 2018 Elsevier B.V. All rights reserved.

Selection of Housekeeping Genes for Transgene Expression Analysis in Eucommia ulmoides Oliver Using Real-Time RT-PCR

Directory of Open Access Journals (Sweden)

Ren Chen

2010-01-01

Full Text Available In order to select appropriate housekeeping genes for accurate calibration of experimental variations in real-time (RT- PCR results in transgene expression analysis, particularly with respect to the influence of transgene on stability of endogenous housekeeping gene expression in transgenic plants, we outline a reliable strategy to identify the optimal housekeeping genes from a set of candidates by combining statistical analyses of their (RT- PCR amplification efficiency, gene expression stability, and transgene influences. We used the strategy to select two genes, ACTα and EF1α, from 10 candidate housekeeping genes, as the optimal housekeeping genes to evaluate transgenic Eucommia ulmoides Oliver root lines overexpressing IPPI or FPPS1 genes, which are involved in isoprenoid biosynthesis.

A pulse generator for xenon lamps

CERN Document Server

Janata, E

2002-01-01

A pulse generator is described, which enhances the analyzing light emitted from a xenon lamp as used in kinetic photospectrometry experiments. The lamp current is increased to 600 A for a duration of 3 ms; the current is constant within +-0.2% during a time interval of 2 ms. Because of instabilities of the lamp arc during pulsing, the use of the enhanced light source is limited to measuring times up to 500 mu s. The enhancement in light intensity depends on the wavelength and amounts to more than 400-fold in the UV-region.

Biomaterials in light amplification

Science.gov (United States)

Mysliwiec, Jaroslaw; Cyprych, Konrad; Sznitko, Lech; Miniewicz, Andrzej

2017-03-01

Biologically produced or inspired materials can serve as optical gain media, i.e. they can exhibit the phenomenon of light amplification. Some of these materials, under suitable dye-doping and optical pumping conditions, show lasing phenomena. The emerging branch of research focused on obtaining lasing action in highly disordered and highly light scattering materials, i.e. research on random lasing, is perfectly suited for biological materials. The use of biomaterials in light amplification has been extensively reported in the literature. In this review we attempt to report on progress in the development of biologically derived systems able to show the phenomena of light amplification and random lasing together with the contribution of our group to this field. The rich world of biopolymers modified with molecular aggregates and nanocrystals, and self-organized at the nanoscale, offers a multitude of possibilities for tailoring luminescent and light scattering properties that are not easily replicated in conventional organic or inorganic materials. Of particular importance and interest are light amplification and lasing, or random lasing studies in biological cells and tissues. In this review we will describe nucleic acids and their complexes employed as gain media due to their favorable optical properties and ease of manipulation. We will report on research conducted on various biomaterials showing structural analogy to nucleic acids such as fluorescent proteins, gelatins in which the first distributed feedback laser was realized, and also amyloids or silks, which, due to their dye-doped fiber-like structure, allow for light amplification. Other materials that were investigated in that respect include polysaccharides, like starch exhibiting favorable photostability in comparison to other biomaterials, and chitosan, which forms photonic crystals or cellulose. Light amplification and random lasing was not only observed in processed biomaterials but also in living

The breakdown and glow phases during the initiation of discharges for lamps

International Nuclear Information System (INIS)

Pitchford, L.C.; Peres, I.; Liland, K.B.; Boeuf, J.P.; Gielen, H.

1997-01-01

High intensity discharge (HID) lamps are often initiated by the application of one or more short, high-voltage, breakdown pulses superimposed on a 50 or 60 Hz generator voltage. A successful transition from the breakdown event to steady-state operating conditions in HID lamps requires that the lamp-circuit system be adequate to sustain the plasma created during breakdown until the electrodes are heated to thermionic temperatures. In this article, we use a one-dimensional (in the axial direction) transient discharge model to study the conditions needed to sustain the cold-cathode discharge after a breakdown event has occurred. While the application of our one-dimensional model to real lamps is approximate, we find that the model predictions are consistent with experimental results in HID lamps, a few of which are presented here. The main conclusion from this work is that, after breakdown, the voltage necessary to sustain a glow discharge is dependent on the source impedance, the gas composition, and on the plasma density created by the breakdown event. copyright 1997 American Institute of Physics

Study on residential appliances energy efficiency standards Refrigerators, air-conditioners, incandescent lamps, fluorescent lamps, color TVs

Energy Technology Data Exchange (ETDEWEB)

Lee, S.G.; Cho, S.K.; Choi, S.H.; Jung, B.M.; Han, S.B.; Kim, K.D. [Korea Inst. of Energy Research, Taejon (Korea, Republic of)

1995-12-01

The energy efficiency standards and rating act, as amended by the rational energy utilization act, provides energy efficiency standards and ratings for 6 types of consumer products(refrigerators, air-conditioners, fluorescent lamps, incandescent lamps, ballasts and cars) authorizes the Ministry of Trade, Industry and Energy(MOTIE) to prescribe amended or new energy efficiency standards and rating standards. This study was initiated by the KIER in 1992. KIER`s assessment of the standards is designed to evaluate their statistical and engineering analysis according to Korean(Industrial) Standards(KS). And to make distinction between the poor efficiency and good efficiency models, 5 grades are classified depending on their tested energy efficiency. This year, based on our analysis, MOTIE mandated updated standards for refrigerators, air-conditioners, incandescent lamps, and fluorescent lamps. Also the objective of this study is to set the energy efficiency standards and to grade for color TV sets. (author). 37 refs., 89 figs., 85 tabs.

Rapid, simple and sensitive detection of Q fever by loop-mediated isothermal amplification of the htpAB gene.

Directory of Open Access Journals (Sweden)

Lei Pan

Full Text Available BACKGROUND: Q fever is the most widespread zoonosis, and domestic animals are the most common sources of transmission. It is not only difficult to distinguish from other febrile diseases because of the lack of specific clinical manifestations in humans, but it is also difficult to identify the disease in C. burnetii-carrying animals because of the lack of identifiable features. Conventional serodiagnosis requires sera from the acute and convalescent stages of infection, which are unavailable at early diagnosis. Nested PCR and real-time PCR require equipment. In this study, we developed a Loop-Mediated Isothermal Amplification (LAMP assay to identify C. burnetii rapidly and sensitively. METHODS: A universal LAMP primer set was designed to detect the repeated sequence IS1111a of the htpAB gene of C. burnetii using PrimerExplorer V4 software. The sensitivity of the LAMP assay was evaluated using known quantities of recombined reference plasmids containing the targeted genes. The specificity of the developed LAMP assay was determined using 26 members of order Rickettsiae and 18 other common pathogens. The utility of the LAMP assay was further compared with real time PCR by the examination 24 blood samples including 6 confirmed and 18 probable Q fever cases, which diagnosed by IFA serological assessment and real time PCR. In addition, 126 animal samples from 4 provinces including 97 goats, 7 cattle, 18 horses, 3 marmots and 1 deer were compared by these two methods. RESULTS: The limits of detection of the LAMP assay for the htpAB gene were 1 copy per reaction. The specificity of the LAMP assay was 100%, and no cross-reaction was observed among the bacteria used in the study. The positive rate of unknown febrile patients was 33.3%(95%CI 30.2%-36.4% for the LAMP assay and 8.3%(95%CI 7.4%-9.2% for the real time PCR(P<0.05. Similarly, the total positive rate of animals was 7.9%(95%CI 7.1%-8.7% for the LAMP assay and 0.8%(95%CI 0.7%-0.9%for the real time

Nonlinear behavior in high-intensity discharge lamps

NARCIS (Netherlands)

Baumann, Bernd; Schwieger, Joerg; Wolff, Marcus; Manders, Freddy; Suijker, Jos

2016-01-01

The light flicker problem of high intensity discharge lamps is studied numerically and experimentally. It is shown that in some respects the systems behave very similar to the forced Duffing oscillator with a softening spring. In particular, the jump phenomenon and hysteresis are observed in the

RT/OS - realtime programming and application environment for the COSY control system

International Nuclear Information System (INIS)

Weinert, A.; Hacker, U.; Haberbosch, Ch.; Henn, K.; Simon, M.

1994-01-01

This paper presents the highlights and constraints of RT/OS and how it can be used at other places. The support of approximately 1000 VME and VXI CPUs in network-based multiprocessor systems needs a special set of programming tools and application environment which are bundled as the RT/OS Real-Time-Operating-System. Supporting the Motorola 680x0 processors and micro-controller with a development system on UNIX and MS-DOS platforms, the tool set gives the opportunity of bringing small embedded controllers as well as VME multiprocessor systems into operation. The tool set includes a cross compiler and a special version of a remote debugger, as well as application support such as downloading and configuration management. The main feature is a modular kernel with message-based task switching, using the rendezvous principle to allow easy extension. Special features of this kernel for a distributed system are networking with the Berkeley socket library supporting TCP/IP, XNS and Novell Netware, a multiprocessor system on VMEbus with the interprocessor Communication Module (IPC), a device driver library for almost every G64 I/O card using a FieldBus Interface called PDV and a X11R4 port. ((orig.))

RT/OS - realtime programming and application environment for the COSY control system

Energy Technology Data Exchange (ETDEWEB)

Weinert, A [Forschungszentrum, Juelich, Postfach 1913, 52425 Juelich (Germany); Hacker, U [Forschungszentrum, Juelich, Postfach 1913, 52425 Juelich (Germany); Haberbosch, Ch [Forschungszentrum, Juelich, Postfach 1913, 52425 Juelich (Germany); Henn, K [Forschungszentrum, Juelich, Postfach 1913, 52425 Juelich (Germany); Simon, M [Forschungszentrum, Juelich, Postfach 1913, 52425 Juelich (Germany)

1994-12-15

This paper presents the highlights and constraints of RT/OS and how it can be used at other places. The support of approximately 1000 VME and VXI CPUs in network-based multiprocessor systems needs a special set of programming tools and application environment which are bundled as the RT/OS Real-Time-Operating-System. Supporting the Motorola 680x0 processors and micro-controller with a development system on UNIX and MS-DOS platforms, the tool set gives the opportunity of bringing small embedded controllers as well as VME multiprocessor systems into operation. The tool set includes a cross compiler and a special version of a remote debugger, as well as application support such as downloading and configuration management. The main feature is a modular kernel with message-based task switching, using the rendezvous principle to allow easy extension. Special features of this kernel for a distributed system are networking with the Berkeley socket library supporting TCP/IP, XNS and Novell Netware, a multiprocessor system on VMEbus with the interprocessor Communication Module (IPC), a device driver library for almost every G64 I/O card using a FieldBus Interface called PDV and a X11R4 port. ((orig.))

The AMI speaker diarization system for NIST RT06s meeting data

NARCIS (Netherlands)

Leeuwen, D.A. van; Huijbregts, Marijn

2006-01-01

We describe the systems submitted to the NIST RT06s evaluation for the Speech Activity Detection (SAD) and Speaker Diarization (SPKR) tasks. For speech activity detection, a new analysis methodology is presented that generalizes the Detection Erorr Tradeoff analysis commonly used in speaker

The AMI speaker diarization system for NIST RT06s meeting data

NARCIS (Netherlands)

van Leeuwen, David A.; Huijbregts, M.A.H.

2007-01-01

We describe the systems submitted to the NIST RT06s evaluation for the Speech Activity Detection (SAD) and Speaker Diarization (SPKR) tasks. For speech activity detection, a new analysis methodology is presented that generalizes the Detection Erorr Tradeoff analysis commonly used in speaker detection