Details
| Original language | English |
|---|---|
| Pages (from-to) | 956-963 |
| Number of pages | 8 |
| Journal | Environmental Science: Water Research and Technology |
| Volume | 4 |
| Issue number | 7 |
| Publication status | Published - 1 Jul 2018 |
Abstract
Waterborne microbial pathogen detection via nucleic acid analysis on portable microfluidic devices is a growing area of research, development, and application. Traditional polymerase chain reaction (PCR)-based nucleic acid analysis detects total extracted DNA, but cannot differentiate live and dead cells. A propidium monoazide (PMA) pretreatment step before PCR can effectively exclude DNA from nonviable cells, as PMA can selectively diffuse through compromised cell membranes and intercalate with DNA to form DNA-PMA complex upon light exposure. The complex strongly inhibits the amplification of the bound DNA in PCR, and thus, only cells with intact cell membranes are detected. Herein, this study reports the development of a microfluidic device to carry out PMA pretreatment 'on-chip'. Chip design was guided by computer simulations, and prototypes were fabricated using a high-resolution 3D printer. The optimized design utilizes split and recombine mixers for initial PMA-sample mixing and a serpentine flow channel containing herringbone structures for dark and light incubation. On-chip PMA pretreatment to differentiate live and dead bacterial cells in buffer and natural pond water samples was successfully demonstrated.
ASJC Scopus subject areas
- Environmental Science(all)
- Environmental Engineering
- Environmental Science(all)
- Water Science and Technology
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In: Environmental Science: Water Research and Technology, Vol. 4, No. 7, 01.07.2018, p. 956-963.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Propidium monoazide pretreatment on a 3D-printed microfluidic device for efficient PCR determination of 'live versus dead' microbial cells
AU - Zhu, Yanzhe
AU - Huang, Xiao
AU - Xie, Xing
AU - Bahnemann, Janina
AU - Lin, Xingyu
AU - Wu, Xunyi
AU - Wang, Siwen
AU - Hoffmann, Michael R.
N1 - Funding information: The authors acknowledge the financial support of the Bill and Melinda Gates Foundation (BMGF OPP1111246).
PY - 2018/7/1
Y1 - 2018/7/1
N2 - Waterborne microbial pathogen detection via nucleic acid analysis on portable microfluidic devices is a growing area of research, development, and application. Traditional polymerase chain reaction (PCR)-based nucleic acid analysis detects total extracted DNA, but cannot differentiate live and dead cells. A propidium monoazide (PMA) pretreatment step before PCR can effectively exclude DNA from nonviable cells, as PMA can selectively diffuse through compromised cell membranes and intercalate with DNA to form DNA-PMA complex upon light exposure. The complex strongly inhibits the amplification of the bound DNA in PCR, and thus, only cells with intact cell membranes are detected. Herein, this study reports the development of a microfluidic device to carry out PMA pretreatment 'on-chip'. Chip design was guided by computer simulations, and prototypes were fabricated using a high-resolution 3D printer. The optimized design utilizes split and recombine mixers for initial PMA-sample mixing and a serpentine flow channel containing herringbone structures for dark and light incubation. On-chip PMA pretreatment to differentiate live and dead bacterial cells in buffer and natural pond water samples was successfully demonstrated.
AB - Waterborne microbial pathogen detection via nucleic acid analysis on portable microfluidic devices is a growing area of research, development, and application. Traditional polymerase chain reaction (PCR)-based nucleic acid analysis detects total extracted DNA, but cannot differentiate live and dead cells. A propidium monoazide (PMA) pretreatment step before PCR can effectively exclude DNA from nonviable cells, as PMA can selectively diffuse through compromised cell membranes and intercalate with DNA to form DNA-PMA complex upon light exposure. The complex strongly inhibits the amplification of the bound DNA in PCR, and thus, only cells with intact cell membranes are detected. Herein, this study reports the development of a microfluidic device to carry out PMA pretreatment 'on-chip'. Chip design was guided by computer simulations, and prototypes were fabricated using a high-resolution 3D printer. The optimized design utilizes split and recombine mixers for initial PMA-sample mixing and a serpentine flow channel containing herringbone structures for dark and light incubation. On-chip PMA pretreatment to differentiate live and dead bacterial cells in buffer and natural pond water samples was successfully demonstrated.
UR - http://www.scopus.com/inward/record.url?scp=85049244459&partnerID=8YFLogxK
U2 - 10.1039/c8ew00058a
DO - 10.1039/c8ew00058a
M3 - Article
AN - SCOPUS:85049244459
VL - 4
SP - 956
EP - 963
JO - Environmental Science: Water Research and Technology
JF - Environmental Science: Water Research and Technology
SN - 2053-1400
IS - 7
ER -