A single-tube sample preparation method based on a dual-electrostatic interaction strategy for molecular diagnosis of gr
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ORIGINAL PAPER
A single-tube sample preparation method based on a dual-electrostatic interaction strategy for molecular diagnosis of gram-negative bacteria Feixiong Chen 1 & Soyeon Kim 2 & Jun-Hee Na 3 & Kyudong Han 4 & Tae Yoon Lee 1 Received: 28 March 2020 / Accepted: 27 August 2020 # Springer-Verlag GmbH Austria, part of Springer Nature 2020
Abstract A single-tube method based on a dual-electrostatic interaction (EI) strategy for bacteria capture and DNA extraction was designed to enable the highly sensitive detection of nucleic acids. Specially designed magnetic nanoparticles were developed to meet the opposing requirements of a single-tube method, which exist between the strong EI required for efficient bacteria capture and the weak EI required for DNA extraction with minimal DNA adsorption. A dual-EI strategy for the single-tube (DESIGN) method was thus developed to integrate bacteria enrichment, bacteria cell lysis, and DNA recovery in a single tube, thereby minimizing precious sample loss and reducing handling time. Subsequently, we evaluated the performance with a variety of concentrations from 5 to 100 colony-forming units (CFU)/10 mL human urine and milk samples. The DESIGN method achieved the simple and sensitive detection of Salmonella enterica serovar Typhimurium in 10 mL of human urine and milk samples up to 5 CFU by quantitative PCR. Furthermore, the DESIGN method detected Brucella ovis and Escherichia coli from 10 mL of human urine with a detection limit up to 5 CFU/10 mL.
Keywords Electrostatic interaction . Magnetic nanoparticle . Pathogenic bacteria enrichment . Pathogenic bacteria detection Abbreviations EI MNP DESIGN method S. Typhimurium B. ovis E. coli
Introduction Electrostatic interaction Magnetic nanoparticle Dual-electrostatic interaction strategy based on a single-tube method Salmonella enterica serotype Typhimurium Brucella ovis Escherichia coli
Bacterial culture is currently the gold standard for bacterial detection, but it has a long turnaround time [1, 2]. Therefore, nucleic acid amplification tests (NAATs) have emerged as alternatives which shorten the turnaround time required to identify causative microorganisms [3]. However, the success of these methods is hindered by inevitable challenges such as low abundances of bacteria or unknown inhibitors, which
Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00604-020-04536-9) contains supplementary material, which is available to authorized users. * Kyudong Han [email protected] * Tae Yoon Lee [email protected] 1
Department of Convergence System Engineering, Department of Biomedical Engineering, and Department of Technology Education, Chungnam National University, Daejeon 34134, Republic of Korea
2
Graduate School of Energy Science and Technology and Department of Chemical Engineering Education, Chungnam National University, Daejeon 34134, Republic of Korea
3
Department of Electrical, Electronics and Communication Engineering Education and Department of Convergence System En
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