EIS microfluidic chips for flow immunoassay and ultrasensitive cholera toxin detection

Chiriaco, Maria; Primiceri, Elisabetta; D’Amone, Eliana; Ionescu, Rodica Elena; Rinaldi, R.; Maruccio, Giuseppe

doi:10.1039/c0lc00409j

Cited by 61 publications

(57 citation statements)

References 39 publications

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“…Advances in biotechnology over the past decades have led to the development of genomic and proteomic methods for the detection and characterization of pathogenic toxins (31). In this study, we report for the first time the use of real-time, dynamic, CTmediated cytotoxicity monitoring of cell lines for detecting and identifying CT activities.…”

Section: Discussionmentioning

confidence: 99%

“…In addition, classic CT determinations require the use of either animal methods (5,6) or tissue culture methods (7,8), which are also time-consuming and are subjective in the interpretation of results. Efforts have been made recently to develop more-sensitive methods, including enzyme-linked immunosorbent assays (9), latex agglutination assays (10), coagglutination assays (5), liposome-based assays (9,11), radioimmunoassays (12), hydrogel-based immunoassays (13), monosaccharide-and antibody array-based assays (14)(15)(16)(17), PCR-based molecular assays (10,(18)(19)(20)(21), and biosensor-based assays (22)(23)(24)(25)(26)(27)(28)(29)(30)(31). A reliable laboratory tool is desirable for clinical services and epidemiological investigation, to characterize CT activities rapidly and quantitatively.…”

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confidence: 99%

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Quantitative Detection of Vibrio cholera Toxin by Real-Time and Dynamic Cytotoxicity Monitoring

et al. 2013

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eWe report here the quantitative detection of Vibrio cholerae toxin (CT) in isolates and stool specimens by dynamic monitoring of the full course of CT-mediated cytotoxicity in a real-time cell analysis (RTCA) system. Four cell lines, including Y-1 mouse adrenal tumor cells, Chinese hamster ovary (CHO) cells, small intestine epithelial (FHs74Int) cells, and mouse adrenal gland (PC12-Adh) cells, were evaluated for their suitability for CT-induced cytotoxicity testing. Among them, the Y-1 line was demonstrated to be the most sensitive for CT-mediated cytotoxicity, with limits of detection of 7.0 pg/ml for purified CT and 0.11 ng/ml for spiked CT in pooled negative stool specimens. No CT-mediated cytotoxicity was observed for nontoxigenic V. cholerae, non-V. cholerae species, or non-V. cholerae enterotoxins. The CT-RTCA assay was further validated with 100 stool specimens consecutively collected from patients with diarrhea and 200 V. cholerae isolates recovered from patients and the environment, in comparison to a reference using three detection methods. The CT-RTCA assay had sensitivities and specificities of 97.5% and 100.0%, respectively, for V. cholerae isolates and 90.0% and 97.2% for stool specimens. For stool specimens spiked with CT concentrations ranging from 3.5 pg/ml to 1.8 ng/ml, the inoculation-to-detection time was 1.12 ؎ 0.38 h, and the values were inversely correlated with CT concentrations ( ‫؍‬ ؊1; P ‫؍‬ 0.01). The results indicate that the CT-RTCA assay with the Y-1 cell line provides a rapid and sensitive tool for the quantitative detection of CT activities in clinical specimens. Vibrio cholerae is a Gram-negative, comma-shaped, bacterial pathogen causing cholera, an acute secretory diarrheal disease. Epidemic cholera is common in developing countries and affects about 100,000 people annually (1). Cholera toxin (CT) is a major virulence determinant of V. cholerae, leading to rapidly progressing dehydration, shock, metabolic acidosis, and even death within hours without adequate and appropriate therapy (2). CT is a key biomarker of V. cholerae that is used for forecasting and assessing epidemic disease, monitoring and controlling cholera outbreaks, preventing epidemics, and guiding medical personnel in providing timely treatment of patients. There has been urgent demand for the development of novel sensitive assays for the detection and identification of CT proteins.Conventional biochemical and immunological methods for CT detection and identification can be completed only after V. cholerae is isolated. The bacterial isolation and subsequent CT detection and identification are laborious and usually require several days, resulting in significant delays in patient care and disease control (3, 4). In addition, classic CT determinations require the use of either animal methods (5, 6) or tissue culture methods (7,8), which are also time-consuming and are subjective in the interpretation of results. Efforts have been made recently to develop more-sensitive methods, including enzyme-linked immunosorbent...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Quantitative Detection of Vibrio cholera Toxin by Real-Time and Dynamic Cytotoxicity Monitoring

et al. 2013

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“…Typically, a pair of electrodes as an electrical transducer is utilized to measure the impedance change caused by the existence of the biological substances. Literature has demonstrated the use of the similar principle for the detection of various biological substances such as enzymes [23], antibodies and antigens [10,[24][25][26], DNA [27,28], and cells [17,[29][30][31][32][33]. This technique provides a non-invasive and label-free measurement, and is found practically useful for the detection of substances in miniaturized analytical devices like microfluidic systems.…”

Section: Introductionmentioning

confidence: 99%

Review on Impedance Detection of Cellular Responses in Micro/Nano Environment

Lei

2014

Micromachines

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Abstract:In general, cell culture-based assays, investigations of cell number, viability, and metabolic activities during culture periods, are commonly performed to study the cellular responses under various culture conditions explored. Quantification of cell numbers can provide the information of cell proliferation. Cell viability study can understand the percentage of cell death under a specific tested substance. Monitoring of the metabolic activities is an important index for the study of cell physiology. Based on the development of microfluidic technology, microfluidic systems incorporated with impedance measurement technique, have been reported as a new analytical approach for cell culture-based assays. The aim of this article is to review recent developments on the impedance detection of cellular responses in micro/nano environment. These techniques provide an effective and efficient technique for cell culture-based assays.

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“…This electrode can be used to detect cholera toxin from 1-100 ng mL À1 . 44 Thus, there is an increased demand for the availability of a rapid, sensitive and reproducible analytical tool for the detection of Vibrio cholerae (Vc) concentration.…”

Section: Introductionmentioning

confidence: 99%

Reduced graphene oxide–titania based platform for label-free biosensor

et al. 2014

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A label-free biosensor has been fabricated using a reduced graphene oxide (RGO) and anatase titania (antTiO 2 ) nanocomposite, electrophoretically deposited onto an indium tin oxide coated glass substrate. The RGO-ant-TiO 2 nanocomposite has been functionalized with protein (horseradish peroxidase) conjugated antibodies for the specific recognition and detection of Vibrio cholerae. The presence of Ab-Vc on the RGO-ant-TiO 2 nanocomposite has been confirmed using electron microscopy, Fourier transform infrared spectroscopy and electrochemical techniques. Electrochemical studies relating to the fabricated Ab-Vc/RGO-ant-TiO 2 /ITO immunoelectrode have been conducted to investigate the binding kinetics.This immunosensor exhibits improved biosensing properties in the detection of Vibrio cholerae, with a sensitivity of 18.17 Â 10 6 F mol À1 L À1 m À2 in the detection range of 0.12-5.4 nmol L À1 , and a low detection limit of 0.12 nmol L À1 . The association (k a ), dissociation (k d ) and equilibrium rate constants have been estimated to be 0.07 nM, 0.002 nM and 0.41 nM, respectively. This Ab-Vc/RGO-ant-TiO 2 /ITO immunoelectrode could be a suitable platform for the development of compact diagnostic devices.

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EIS microfluidic chips for flow immunoassay and ultrasensitive cholera toxin detection

Cited by 61 publications

References 39 publications

Quantitative Detection of Vibrio cholera Toxin by Real-Time and Dynamic Cytotoxicity Monitoring

Quantitative Detection of Vibrio cholera Toxin by Real-Time and Dynamic Cytotoxicity Monitoring

Review on Impedance Detection of Cellular Responses in Micro/Nano Environment

Reduced graphene oxide–titania based platform for label-free biosensor

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