A dry-reagent assay to rapidly detect Mycobacterium tuberculosis using loop-mediated isothermal amplification.
10.3760/cma.j.cn112150-20230623-00488
- VernacularTitle:基于环介导等温扩增和干式试剂技术的结核分枝杆菌快速检测方法的建立
- Author:
Bao Hua LI
1
;
Ying ZHANG
2
;
Shun TAO
2
;
Yan Ni GUO
1
;
Qing LIU
1
;
Qi Qi SUN
2
Author Information
1. Clinical Laboratory, University of Chinese Academy of Sciences Shenzhen Hospital, Shenzhen 518107, China.
2. EDAN Instruments Inc., Shenzhen 518122, China.
- Publication Type:Journal Article
- MeSH:
Humans;
Mycobacterium tuberculosis/genetics*;
Indicators and Reagents;
Sensitivity and Specificity;
Nucleic Acid Amplification Techniques/methods*;
DNA
- From:
Chinese Journal of Preventive Medicine
2023;57(10):1625-1629
- CountryChina
- Language:Chinese
-
Abstract:
A molecular diagnostic assay which could be stored at room temperature was developed to rapidly detect Mycobacterium tuberculosis (MTB) based on loop-mediated isothermal amplification (LAMP) technology and dry-reagent process. LAMP uses 4 or 6 primers and Bst DNA polymerase to amplify DNA at a constant temperature. The results showed that the LAMP assay could detect the amplification of IS6110 target gene within 20 min using real-time fluorescence signal detection. The sensitive of LAMP assay was similar to the PCR technology while the precision of PCR was better than LAMP (coefficient of variation, LAMP 18.9%, PCR 3.4%), meaning LAMP was more suitable for qualitative detection. The LAMP assay did not amplify DNA of other 10 types of pathogens, including Neisseria meningitidis, Haemophilus influenzae, Staphylococcus aureus, Streptococcus pneumoniae, Rubivirus, mumps virus, adenovirus (type 3), adenovirus (type 7), respiratory syncytial virus B and parainfluenza virus type 2, indicating a good specificity. Furthermore, a dry-reagent assay was developed using air-drying and freeze-drying process. The performance of dried reagents did not change after 10 days storage at 50 ℃, meaning the dried reagents could be stored at room temperature (25 ℃) for more than six months. The dry-reagent LAMP assay also successfully amplified MTB DNA from several clinical samples within 20 min. In conclusion, the developed LAMP assay together with isothermal amplifier could rapidly detection MTB.
