Establishment of Sample-to-Answer Loop-Mediated Isothermal Amplification-Based Nucleic Acid Testing Using the Sampling, Processing, Incubation, Detection and Lateral Flow Immunoassay Platforms

被引：0
作者：
Pommies, Lilas [1 ]
Boutal, Herve [1 ]
Fras, David [2 ]
Volland, Herve [1 ]
机构：
[1] Univ Paris Saclay, Dept Med & Technol Sante DMTS, CEA, INRAE,SPI, F-91191 Gif Sur Yvette, France
[2] CEA DRT LIST DIN SIMRI, F-91191 Gif Sur Yvette, France
来源：
BIOSENSORS-BASEL | 2024年 / 14卷 / 12期
关键词：
sample-to-answer test; POCT; genetic detection; RECOMBINASE POLYMERASE AMPLIFICATION; MULTIPLE DISPLACEMENT AMPLIFICATION; RAPID DETECTION; DNA; SIGNAL; PAPER; ASSAY; LAMP; DIAGNOSIS;
D O I：
10.3390/bios14120609
中图分类号：
O65 [分析化学];
学科分类号：
070302 ; 081704 ;
摘要：
Diagnostics often require specialized equipment and trained personnel in laboratory settings, creating a growing need for point-of-care tests (POCTs). Among the genetic testing methods available, Loop-mediated Isothermal Amplification (LAMP) offers a viable solution for developing genetic POCT due to its compatibility with simplified devices. This study aimed to create a genetic test that integrates all steps from sample processing to analyzing results while minimizing the complexity, handling, equipment, and time required. Several challenges were addressed to achieve this goal: (1) the development of a buffer for bacterial DNA extraction that is compatible with both LAMP and immunochromatographic tests; (2) the adaption of the LAMP protocol for use with the SPID device; and (3) the optimization of the detection protocol for specific test conditions, with a lateral flow immunoassay format selected for its POCT compatibility. Following these developments, the test was validated using Escherichia coli (E. coli) and non-E. coli strains. A portable heating station was also developed to enable amplification without costly equipment. The resulting genetic POCT achieved 100% sensitivity and 85% specificity, with results available in 60 to 75 min. This study demonstrated that our POCT efficiently performs DNA extraction, amplification, and detection for bacterial identification. The test's simplicity and cost-effectiveness will support its implementation in various settings.
引用
收藏
页数：19
相关论文
共 55 条
  • [1] Fu S., Jiang Y., Jiang X., Zhao Y., Chen S., Yang X., Man C., Probe-Free Label System for Rapid Detection of Cronobacter Genus in Powdered Infant Formula, AMB Express, 8, (2018)
  • [2] Yager P., Domingo G.J., Gerdes J., Point-of-Care Diagnostics for Global Health, Annu. Rev. Biomed. Eng, 10, pp. 107-144, (2008)
  • [3] Land K.J., Boeras D.I., Chen X.-S., Ramsay A.R., Peeling R.W., REASSURED Diagnostics to Inform Disease Control Strategies, Strengthen Health Systems and Improve Patient Outcomes, Nat. Microbiol, 4, pp. 46-54, (2019)
  • [4] Wang S., Xu F., Demirci U., Advances in Developing HIV-1 Viral Load Assays for Resource-Limited Settings, Biotechnol. Adv, 28, pp. 770-781, (2010)
  • [5] Zhao Y., Chen F., Li Q., Wang L., Fan C., Isothermal Amplification of Nucleic Acids, Chem. Rev, 115, pp. 12491-12545, (2015)
  • [6] Karami A., Gill P., Motamedi M.H.K., Saghafinia M., A Review of the Current Isothermal Amplification Techniques: Applications, Advantages and Disadvantages, J. Glob. Infect. Dis, 3, (2011)
  • [7] Walker G.T., Fraiser M.S., Schram J.L., Little M.C., Nadeau J.G., Malinowski D.P., Strand Displacement Amplification—An Isothermal, in Vitro DNA Amplification Technique, Nucl. Acids Res, 20, pp. 1691-1696, (1992)
  • [8] Ali M.M., Li F., Zhang Z., Zhang K., Kang D.-K., Ankrum J.A., Le X.C., Zhao W., Rolling Circle Amplification: A Versatile Tool for Chemical Biology, Materials Science and Medicine, Chem. Soc. Rev, 43, pp. 3324-3341, (2014)
  • [9] Lobato I.M., O'Sullivan C.K., Recombinase Polymerase Amplification: Basics, Applications and Recent Advances, Trends Anal. Chem, 98, pp. 19-35, (2018)
  • [10] Mok E., Wee E., Wang Y., Trau M., Comprehensive Evaluation of Molecular Enhancers of the Isothermal Exponential Amplification Reaction, Sci. Rep, 6, (2016)
123456