ABSTRACT
We compared the RT-LAMP,LAMP and L-J for detecting MTB to provide the rapid diagnosis method for tu-berculosis.In this assay,NTM and other common respiratory bacteria were used to detect specificity,Mycobacterium tubercu-losis specific products were identified by restriction enzyme digestion.To detect sensitivity,we used RT-LAMP,LAMP and L-J to detect 100 cases sputum specimens from patients with tuberculosis and 22 cases control sputum specimens,the 10 times dilution of 1 ng/μL H37Rv standard strains was used to detect RT-LAMP limit.The results showed that the positive rate of RT-LAMP,LAMP and L-J were 100%,92%,88%.RT-LAMP and L-J,RT-LAMP and LAMP were statistically signifi-cant,RT-LAMP had 10 times higher sensitivity than LAMP,RT-LAMP not only to identify viable but also capable of detec-ting a single copy of MTB.So RT-LAMP is superior to LAMP and L-J and is practical for use in primary medical care institu-tion or peripheral laboratory.
ABSTRACT
Purpose: The objective of this study was to establish a reverse-transcription loop-mediated isothermal amplification (RT-LAMP) method for rapid detection of human immunodeficiency virus type 1 (HIV-1). Materials and Methods: The HIV-1 integrase gene region was selected because it was a conserved part of the HIV-1 genome. Six primers specific to eight regions of the HIV-1 integrase gene were designed. A total of 171 samples (18 HIV-1 confirmed positive samples and 153 serum specimens were collected in this study) were tested by RT-LAMP and reverse-transcription polymerase chain reaction (RT-PCR). After amplification in an isothermal water bath for 45 min, samples containing HIV-1 generated the expected ladder-like products while other viruses generated no product. Results: The sensitivity and specificity of the RT-LAMP assay were evaluated by comparison with RT-PCR. The assay was significantly more sensitive than normal gel-based RT-PCR. Conclusion: Because it is specific and simple, the RT-LAMP assay can be widely applied in clinical laboratories for rapid detection of HIV-1.
Subject(s)
HIV-1/analysis , HIV-1/genetics , Humans , Nucleic Acid Amplification Techniques/methods , Reverse Transcriptase Polymerase Chain Reaction/methods , RNA, Viral/analysis , RNA, Viral/genetics , Sensitivity and SpecificityABSTRACT
A real-time monitoring reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay was developed for the sensitive and specific detection of prototypic,prevalent North American porcine reproductive and respiratory syndrome virus (PRRSV)strains.As a higher sensitivity and specificity method than reverse transcription polymerase chain reaction (RT-PCR),the RT-LAMP method only used a turbidimeter,exhibited a detection limit corresponding to a 10-4 dilution of template RNA extracted from 250 μL of 105 of the 50% tissue culture infective dose (TCID50) of PRRSV containing cells,and no cross-reactivity was observed with other related viruses including porcine circovirus type 2,swine influenza virus,porcine rotavirus and classical swine fever virus.From forty-two field samples,33 samples in the RT-LAMP assay was detected positive,whereas three of which were not detected by RT-PCR.Furthermore,in 33 strains of PRRSV,an identical detection rate was observed with the RT-LAMP assay to what were isolated using porcine alveolar macrophages.These findings demonstrated that the RT-LAMP assay has potential clinical applications for the detection of highly pathogenic PRRSV isolates,especially in developing countries.
ABSTRACT
A simple and rapid assay for the detection of Classical swine fever virus(CSFV)was established using reverse transcription loop-mediated isothermal amplification(RT-LAMP).This study describes the amplification of the genomic RNA of CSFV under isothermal conditions(63℃)within one hour,using a set of six primers(two outer primers,two inner primers and two loop primers).This RT-LAMP assay showed 100-fold higher sensitivity than the standard RT-PCR method and identified eighteen additional positive cases that were negative when tested by RT-PCR.This RT-LAMP was able to detect all the 13 strains of CSFV but not the BVDV.PRRSV.SIV.PRV-PCV,thus showed a good specificity.Products amplified by RT-LAMP can be visualized by agarose gel electrophoresis and in addition,either as a white precipitate at the bottom of the tube after a pulse spin or as a color change when dyed with SYBR Green I which are visible to the naked eye.Because RT-LAMP is low-cost and produces rapid results,it has the potential to be an excellent tool for CSFV surveillance in the field,especially in developing countries.