Abstract
An aptamer-based electrochemical biosensor was successfully developed and applied in the rapid detection of pathogenic Escherichia coli (E. coli) in licorice extract. The thiolated capture probes were firstly immobilized on a gold electrode, and then the biotinylated aptamer probes for E. coli were introduced by hybridization with the capture probes. Due to the stronger interaction between the aptamer and the E. coli, a part of the biotinylated aptamers will dissociate from the capture probes in the presence of E. coli. The residual biotinylated aptamer probes can quantitatively bind with streptavidin-alkaline phosphatase. Subsequently, α-naphthyl phosphate substrate was catalytically hydrolyzed to generate electrochemical response, which could be recorded by a differential pulse voltammetry. The dependence of the peak current on the logarithm of E. coli concentration in the range from 5.0 × 102 colony forming units (CFU)/mL to 5.0 × 107 CFU/mL exhibited a linear trend with a detection limit of 80 CFU/mL. The relative standard deviation of 5 successive scans was 5.3%, 4.5% and 1.1% for 5.0 × 102, 5.0 × 105 and 5.0 × 107 CFU/mL E. coli, respectively. In the detection of the licorice extract samples, the results obtained from the proposed strategy and traditional culture counting method were close to each other, but the time consumption was only ~1/30 compared with the traditional method. These results demonstrate that the designed biosensor can be potentially utilized for rapid microbial examination in traditional Chinese medicine and relevant fields.
Highlights
Pathogenic Escherichia coli (E. coli) is one of the most harmful bacteria related to food-borne diseases and can cause inflammation, abdominal pain, diarrhea and even severe cases like hemorrhagic enteritis and hemolysis, especially in infants and youngsters [1]
The thiol-modified capture probes (SH-Capture) are immobilized on the surface of the polished gold electrode by Au-SH covalent bond, and the unreacted active sites on the electrode surface are blocked with MCH [38]
Streptavidin-alkaline phosphatase (ST-AP) can dissociate from the biotinylated aptamer probes and catalyze the hydrolysis of α-Naphthyl phosphate (α-NP) substrate generating the electrochemical signals [39]
Summary
Pathogenic Escherichia coli (E. coli) is one of the most harmful bacteria related to food-borne diseases and can cause inflammation, abdominal pain, diarrhea and even severe cases like hemorrhagic enteritis and hemolysis, especially in infants and youngsters [1]. A fast, sensitive, and easy-to-use method for the detection of E. coli is an urgent demand in clinical diagnosis, environment monitoring, pharmacy and food safety fields. E. coli detection methods mainly include the traditional culture counting method [3], molecular biological detection technology [4], immunological detection method [5], surface plasmon resonance (SPR) [6], Chemiluminescence immunoassay [7], etc. The traditional counting method for E. coli detection is based on culturing and isolation of bacteria in a specific medium, followed by observing.
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