Rapid Bacteria Detection at Low Concentrations Using
Sequential Immunomagnetic Separation and Paper-Based Isotachophoresis
Posted on 2019-07-18 - 18:47
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.
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Schaumburg, Federico; Carrell, Cody S.; Henry, Charles S. (2019). Rapid Bacteria Detection at Low Concentrations Using
Sequential Immunomagnetic Separation and Paper-Based Isotachophoresis. ACS Publications. Collection. https://doi.org/10.1021/acs.analchem.9b01002