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An Electrochemical Immunosensor for Detection of Staphylococcus aureus Bacteria Based on Immobilization of Antibodies on Self-Assembled Monolayers-Functionalized Gold Electrode.

Braiek M, Rokbani KB, Chrouda A, Mrabet B, Bakhrouf A, Maaref A, Jaffrezic-Renault N - Biosensors (Basel) (2012)

Bottom Line: The EIS technique was used for affinity assays: antibody-cell binding.The limit of detection (LOD) was observed at 10 CFU/mL, and the reproducibility was calculated to 8%.Finally, a good selectivity versus E. coli and S. epidermidis was obtained for our developed immunosensor demonstrating its specificity towards only S. aureus.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Physique et Chimie des Interfaces, Faculté des Sciences de Monastir, Tunisie, Avenue de l'Environnement, 5019 Monastir, Tunisia. mohamed_braiek@yahoo.fr.

ABSTRACT
The detection of pathogenic bacteria remains a challenge for the struggle against biological weapons, nosocomial diseases, and for food safety. In this research, our aim was to develop an easy-to-use electrochemical immunosensor for the detection of pathogenic Staphylococcus aureus ATCC25923. The biosensor was elaborated by the immobilization of anti-S. aureus antibodies using a self-assembled monolayer (SAMs) of 3-Mercaptopropionic acid (MPA). These molecular assemblies were spontaneously formed by the immersion of the substrate in an organic solvent containing the SAMs that can covalently bond to the gold surface. The functionalization of the immunosensor was characterized using two electrochemical techniques: cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Here, the analysis was performed in phosphate buffer with ferro/ferricyanide as the redox probe. The EIS technique was used for affinity assays: antibody-cell binding. A linear relationship between the increment in the electron transfer resistance (RCT) and the logarithmic value of S. aureus concentration was observed between 10 and 106 CFU/mL. The limit of detection (LOD) was observed at 10 CFU/mL, and the reproducibility was calculated to 8%. Finally, a good selectivity versus E. coli and S. epidermidis was obtained for our developed immunosensor demonstrating its specificity towards only S. aureus.

No MeSH data available.


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Different stages in the development of the biofilm.
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biosensors-02-00417-f001: Different stages in the development of the biofilm.

Mentions: The Au electrodes were ultrasonically cleaned in acetone for 10 min, followed by 5 min in a mixture of piranha solution (H2O2/H2SO4, 3/7, v/v). The electrodes were thoroughly rinsed with ultrapure water, followed by absolute ethanol, and then dried in a flow of nitrogen. Subsequently, the electrodes were incubated in a MPA (10 mM) solution made-up with ethanol for 12 h to formulate the SAMs. After the formation, the electrodes were rinsed in ethanol. Then, the terminal carboxylic acid (–COOH) groups were activated in a solution of NHS/EDC (0.1 M/0.1 M) for 1 h at room temperature. After rinsing with PBS, the electrodes were incubated for 1 h in a 0.3 mg/mL solution of anti-S. aureus antibodies. The terminal amine groups on the antibody enable covalent bonding to occur through the activated carboxylic functions from MPA functionalized NHS/EDC. Finally, after rinsing with PBS, the electrodes were incubated for 20 min in ethanolamine. This blocks the remaining acidic functionalities. The mechanism for this activation can be seen in Figure 1.


An Electrochemical Immunosensor for Detection of Staphylococcus aureus Bacteria Based on Immobilization of Antibodies on Self-Assembled Monolayers-Functionalized Gold Electrode.

Braiek M, Rokbani KB, Chrouda A, Mrabet B, Bakhrouf A, Maaref A, Jaffrezic-Renault N - Biosensors (Basel) (2012)

Different stages in the development of the biofilm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263564&req=5

biosensors-02-00417-f001: Different stages in the development of the biofilm.
Mentions: The Au electrodes were ultrasonically cleaned in acetone for 10 min, followed by 5 min in a mixture of piranha solution (H2O2/H2SO4, 3/7, v/v). The electrodes were thoroughly rinsed with ultrapure water, followed by absolute ethanol, and then dried in a flow of nitrogen. Subsequently, the electrodes were incubated in a MPA (10 mM) solution made-up with ethanol for 12 h to formulate the SAMs. After the formation, the electrodes were rinsed in ethanol. Then, the terminal carboxylic acid (–COOH) groups were activated in a solution of NHS/EDC (0.1 M/0.1 M) for 1 h at room temperature. After rinsing with PBS, the electrodes were incubated for 1 h in a 0.3 mg/mL solution of anti-S. aureus antibodies. The terminal amine groups on the antibody enable covalent bonding to occur through the activated carboxylic functions from MPA functionalized NHS/EDC. Finally, after rinsing with PBS, the electrodes were incubated for 20 min in ethanolamine. This blocks the remaining acidic functionalities. The mechanism for this activation can be seen in Figure 1.

Bottom Line: The EIS technique was used for affinity assays: antibody-cell binding.The limit of detection (LOD) was observed at 10 CFU/mL, and the reproducibility was calculated to 8%.Finally, a good selectivity versus E. coli and S. epidermidis was obtained for our developed immunosensor demonstrating its specificity towards only S. aureus.

View Article: PubMed Central - PubMed

Affiliation: Laboratoire de Physique et Chimie des Interfaces, Faculté des Sciences de Monastir, Tunisie, Avenue de l'Environnement, 5019 Monastir, Tunisia. mohamed_braiek@yahoo.fr.

ABSTRACT
The detection of pathogenic bacteria remains a challenge for the struggle against biological weapons, nosocomial diseases, and for food safety. In this research, our aim was to develop an easy-to-use electrochemical immunosensor for the detection of pathogenic Staphylococcus aureus ATCC25923. The biosensor was elaborated by the immobilization of anti-S. aureus antibodies using a self-assembled monolayer (SAMs) of 3-Mercaptopropionic acid (MPA). These molecular assemblies were spontaneously formed by the immersion of the substrate in an organic solvent containing the SAMs that can covalently bond to the gold surface. The functionalization of the immunosensor was characterized using two electrochemical techniques: cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS). Here, the analysis was performed in phosphate buffer with ferro/ferricyanide as the redox probe. The EIS technique was used for affinity assays: antibody-cell binding. A linear relationship between the increment in the electron transfer resistance (RCT) and the logarithmic value of S. aureus concentration was observed between 10 and 106 CFU/mL. The limit of detection (LOD) was observed at 10 CFU/mL, and the reproducibility was calculated to 8%. Finally, a good selectivity versus E. coli and S. epidermidis was obtained for our developed immunosensor demonstrating its specificity towards only S. aureus.

No MeSH data available.


Related in: MedlinePlus