Rapid Bacteria Detection at Low Concentrations Using Sequential Immunomagnetic Separation and Paper-Based Isotachophoresis
| Autor: | Charles S. Henry, Cody S. Carrell, Federico Schaumburg |
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| Rok vydání: | 2019 |
| Předmět: |
Paper
PAPER ELECTROPHORESIS INGENIERÍAS Y TECNOLOGÍAS 010402 general chemistry Immunomagnetic separation 01 natural sciences Analytical Chemistry chemistry.chemical_compound BACTERIA ASSAYS Escherichia coli Chlorophenol red Volume concentration Detection limit Chlorophenol Bacteriological Techniques Chromatography biology Isotachophoresis Immunomagnetic Separation 010401 analytical chemistry Paper based biology.organism_classification UPADS 0104 chemical sciences Ingeniería Química Fruit and Vegetable Juices chemistry Malus Food Microbiology Single-Cell Analysis Bacteria |
| Zdroj: | Analytical chemistry. 91(15) |
| ISSN: | 1520-6882 |
| Popis: | Detecting bacteria is important in the fields of human health, environmental monitoring, and food safety. Foodborne pathogens alone are estimated to cause 420 »000 deaths annually, with low-income regions affected most. Despite improvements in bacterial detection, fast, disposable, low-cost, sensitive, and user-friendly methods are still needed. Traditional methods for detecting bacteria rely primarily on cell culturing or polymerase chain reaction (PCR), which require highly trained personnel and a central laboratory and take several hours or even days to deliver results. Low-cost methods like lateral flow immunoassays exist but frequently suffer from poor sensitivity and/or lack quantitative results. Here, a rapid method for detecting bacteria at very low concentrations is presented using two sequential preconcentration steps. In the first preconcentration step, the sample is mixed with antibody-modified magnetic particles and free antibodies conjugated to β-galactosidase (β-gal). The target bacteria are isolated and concentrated using immunomagnetic separation. The isolated bacteria are then incubated with chlorophenol red-β-d-galactopyranoside (CPRG), which reacts with β-gal to produce chlorophenol red (CPR) in a bacteria concentration-dependent manner. In the second step, CPR and CPRG are separated and focused using an isotachophoretic microfluidic paper-based analytical device, significantly improving the final detection limit relative to paper-based devices lacking the focusing mechanism. Moreover, CPR and CPRG form two visible color bands that act as test and control bands, respectively, improving assay robustness. The method was tested with E. coli DH5-α and successfully detected concentrations as low as 9.2 CFU/mL in laboratory samples and 920 CFU/mL in apple juice samples in ∼90 min. Fil: Schaumburg, Federico. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Santa Fe. Instituto de Desarrollo Tecnológico para la Industria Química. Universidad Nacional del Litoral. Instituto de Desarrollo Tecnológico para la Industria Química; Argentina Fil: Carrell, Cody S.. State University of Colorado - Fort Collins; Estados Unidos Fil: Henry, Charles S.. State University of Colorado - Fort Collins; Estados Unidos |
| Databáze: | OpenAIRE |
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