Sensitive and simple detection of Escherichia coli strain based on time-resolved fluorescence DNA hybridization assay.
A two-probe tandem DNA hybridization assay based on time-resolved fluorescence was employed to detect Escherichia coli strain. The amino modified capture probe was covalently immobilized on the common glass slide surface. The Eu(TTA)(3)(5-NH(2)-phen) with the characteristics of long lifetime and intense luminescence was labeled with reporter probe. The original extracted DNA samples without the purification and amplification process were directly used in the hybridization assay. The concentration of capture probe, hybridization temperature, hybridization and washing time were optimized. The detection limit is about 1.49x10(3) CFU mL(-1) E. coli cells, which is comparable to the value of most microbiology methods. The proposed method has the advantages of easy operation, satisfactory sensitivity and specificity, which can provide a promising technique for monitoring the microorganisms.
College of Environmental Science and Engineering, Key Laboratory of Environmental Biology and Pollution Control, Ministry of Education, Hunan University, Changsha 410082, China.
Min
M
Ruan
Cheng-Gang
CG
Niu
Pin-Zhu
PZ
Qin
Guang-Ming
GM
Zeng
Zhao-Hui
ZH
Yang
Hui
H
He
Jing
J
Huang
D056831:Coordination Complexes / Q000737:chemistry; D015342:DNA Probes / Q000737:chemistry*; D004269:DNA, Bacterial / Q000737:chemistry*; D004926:Escherichia coli / Q000235:genetics / Q000302:isolation & purification*; D005063:Europium / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry*; D009693:Nucleic Acid Hybridization / Q000379:methods*
D056831:Coordination Complexes; D015342:DNA Probes; D004269:DNA, Bacterial; D005456:Fluorescent Dyes; D005063:Europium