[Electrochemical detection of toxin gene in Listeria monocytogenes].
Listeria monocytogenes (LM) is a food-borne pathogen inducing listeriosis, an illness characterized by encephalitis, septicaemia, and meningitis. Listeriolysin O (LLO) is absolutely required for virulence by L. monocytogenes, and is found only in virulent strains of the species. One of the best ways to detect and confirm the pathogen is detection of one of the virulence factors, LLO, produced by the microorganism. This paper focused on the electrical method used to detect the LLO toxin gene in food products and organism without labeling the target DNA. The electrochemical sensor was obtained by immobilizing single-stranded oligonucleotides onto the gold electrode with the mercaptan activated by N-hydroxysulfosuccinimide (NHS) and N-(3-dimethylamion)propyl-N'-ethyl carbodiimidehydrochloride (EDC). The hy-bridization reaction that occurred on the electrode surface was evidenced by Cyclic Voltammetry (CV) analysis using [Co(phen)3](ClO4)3 as an indicator. The covalently immobilized single-stranded DNA could selectively hybridize to its complementary DNA in solution to form double-stranded DNA on the gold surface. A significant increase of the peak cur-rent of Cyclic Voltammetry (CV) upon hybridization of immobilized ssDNA with PCR amplification products in the solu-tion was observed. This peak current change was used to monitor the amount of PCR amplification products. Factors deter-mining the sensitivity of the electrochemical assay, such as DNA target concentration and hybridization conditions, were investigated. The coupling of DNA to the electrochemical sensors has the potential of the quantitative evaluation of gene.
Wu
Ling-Wei
LW
State Key Laboratory of Bioelectronics, Southeast University, Nanjing 210096, China. jxoai@hotmail.com
Liu
Quan-Jun
QJ
Wu
Zhong-Wei
ZW
Lu
Zu-Hong
ZH
D001427:Bacterial Toxins / Q000032:analysis* / Q000235:genetics; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods; D002234:Carbodiimides / Q000737:chemistry; D003035:Cobalt / Q000737:chemistry; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004277:DNA, Single-Stranded / Q000737:chemistry / Q000235:genetics; D055664:Electrochemical Techniques / Q000295:instrumentation / Q000379:methods*; D004566:Electrodes; D006046:Gold / Q000737:chemistry; D006360:Heat-Shock Proteins / Q000032:analysis* / Q000235:genetics; D006460:Hemolysin Proteins / Q000032:analysis* / Q000235:genetics; D006801:Humans; D008089:Listeria monocytogenes / Q000235:genetics / Q000378:metabolism; D008088:Listeriosis / Q000175:diagnosis / Q000382:microbiology; D009942:Organometallic Compounds / Q000737:chemistry; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013388:Succinimides / Q000737:chemistry; D013438:Sulfhydryl Compounds / Q000737:chemistry
D001427:Bacterial Toxins; D002234:Carbodiimides; D004269:DNA, Bacterial; D004277:DNA, Single-Stranded; D006360:Heat-Shock Proteins; D006460:Hemolysin Proteins; D009942:Organometallic Compounds; D013388:Succinimides; D013438:Sulfhydryl Compounds; D003035:Cobalt; D006046:Gold; C035761:N-hydroxysulfosuccinimide; C038705:hlyA protein, Listeria monocytogenes