Tags

Type your tag names separated by a space and hit enter

Real-time detection of Escherichia coli O157:H7 sequences using a circulating-flow system of quartz crystal microbalance.
Biosens Bioelectron. 2007 Jun 15; 22(12):2967-75.BB

Abstract

A DNA piezoelectric biosensing method for real-time detection of Escherichia coli O157:H7 in a circulating-flow system was developed in this study. Specific probes [a 30-mer oligonucleotide with or without additional 12 deoxythymidine 5'-monophosphate (12-dT)] for the detection of E. coli O157:H7 gene eaeA, synthetic oligonucleotide targets (30 and 104 mer) and PCR-amplified DNA fragments from the E. coli O157:H7 eaeA gene (104 bp), were used to evaluate the efficiency of the probe immobilization and hybridization with target DNA in the circulating-flow quartz crystal microbalance (QCM) device. It was found that thiol modification on the 5'-end of the probes was essential for probe immobilization on the gold surface of the QCM device. The addition of 12-dT to the probes as a spacer, significantly enhanced (P<0.05) the hybridization efficiency (H%). The results indicate that the spacer enhanced the H% by 1.4- and 2-fold when the probes were hybridized with 30- and 104-mer targets, respectively. The spacer reduced steric interference of the support on the hybridization behavior of immobilized oligonucleotides, especially when the probes hybridized with relatively long oligonucleotide targets. The QCM system was also applied in the detection of PCR-amplified DNA from real samples of E. coli O157:H7. The resultant H% of the PCR-amplified double-strand DNA was comparable to that of the synthetic target T-104AS, a single-strand DNA. The piezoelectric biosensing system has potential for further applications. This approach lays the groundwork for incorporating the method into an integrated system for rapid PCR-based DNA analysis.

Authors+Show Affiliations

Department of Food Science and Human Nutrition, University of Maine, Orono, ME 04469-5735, USA.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

17223335

Citation

Wu, Vivian C H., et al. "Real-time Detection of Escherichia Coli O157:H7 Sequences Using a Circulating-flow System of Quartz Crystal Microbalance." Biosensors & Bioelectronics, vol. 22, no. 12, 2007, pp. 2967-75.
Wu VC, Chen SH, Lin CS. Real-time detection of Escherichia coli O157:H7 sequences using a circulating-flow system of quartz crystal microbalance. Biosens Bioelectron. 2007;22(12):2967-75.
Wu, V. C., Chen, S. H., & Lin, C. S. (2007). Real-time detection of Escherichia coli O157:H7 sequences using a circulating-flow system of quartz crystal microbalance. Biosensors & Bioelectronics, 22(12), 2967-75.
Wu VC, Chen SH, Lin CS. Real-time Detection of Escherichia Coli O157:H7 Sequences Using a Circulating-flow System of Quartz Crystal Microbalance. Biosens Bioelectron. 2007 Jun 15;22(12):2967-75. PubMed PMID: 17223335.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Real-time detection of Escherichia coli O157:H7 sequences using a circulating-flow system of quartz crystal microbalance. AU - Wu,Vivian C H, AU - Chen,Sz-Hau, AU - Lin,Chih-Sheng, Y1 - 2007/01/16/ PY - 2006/08/07/received PY - 2006/11/28/revised PY - 2006/12/07/accepted PY - 2007/1/16/pubmed PY - 2007/7/27/medline PY - 2007/1/16/entrez SP - 2967 EP - 75 JF - Biosensors & bioelectronics JO - Biosens Bioelectron VL - 22 IS - 12 N2 - A DNA piezoelectric biosensing method for real-time detection of Escherichia coli O157:H7 in a circulating-flow system was developed in this study. Specific probes [a 30-mer oligonucleotide with or without additional 12 deoxythymidine 5'-monophosphate (12-dT)] for the detection of E. coli O157:H7 gene eaeA, synthetic oligonucleotide targets (30 and 104 mer) and PCR-amplified DNA fragments from the E. coli O157:H7 eaeA gene (104 bp), were used to evaluate the efficiency of the probe immobilization and hybridization with target DNA in the circulating-flow quartz crystal microbalance (QCM) device. It was found that thiol modification on the 5'-end of the probes was essential for probe immobilization on the gold surface of the QCM device. The addition of 12-dT to the probes as a spacer, significantly enhanced (P<0.05) the hybridization efficiency (H%). The results indicate that the spacer enhanced the H% by 1.4- and 2-fold when the probes were hybridized with 30- and 104-mer targets, respectively. The spacer reduced steric interference of the support on the hybridization behavior of immobilized oligonucleotides, especially when the probes hybridized with relatively long oligonucleotide targets. The QCM system was also applied in the detection of PCR-amplified DNA from real samples of E. coli O157:H7. The resultant H% of the PCR-amplified double-strand DNA was comparable to that of the synthetic target T-104AS, a single-strand DNA. The piezoelectric biosensing system has potential for further applications. This approach lays the groundwork for incorporating the method into an integrated system for rapid PCR-based DNA analysis. SN - 0956-5663 UR - https://www.unboundmedicine.com/medline/citation/17223335/Real_time_detection_of_Escherichia_coli_O157:H7_sequences_using_a_circulating_flow_system_of_quartz_crystal_microbalance_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0956-5663(06)00606-3 DB - PRIME DP - Unbound Medicine ER -