Targeted Capture of Pathogenic Bacteria Using a Mammalian Cell Receptor Coupled with Dielectrophoresis on a Biochip

Abstract

Efficient capture of target analyte on biosensor platforms is a prerequisite for reliable and specific detection of pathogenic microorganisms in a microfluidic chip. Antibodies have been widely used as ligands; however, because of their occasional unsatisfactory performance, a search for alternative receptors is underway. Heat shock protein 60 (Hsp60), a eukaryotic mitochondrial chaperon protein is a receptor for Listeria adhesion protein (LAP) during Listeria monocytogenes infection. This paper reports application of biotinylated Hsp60 as a capture molecule for living (viable) L. monocytogenes in a microfluidic environment. Hsp60, immobilized on the surface of streptavidin-coated silicon dioxide exhibited specific capture of pathogenic Listeria against a background of other Listeria species, Salmonella, Escherichia, Bacillus, Pseudomonas, Serratia, Hafnia, Enterobacter, Citrobacter, and Lactobacillus. The capture efficiency of L. monocytogenes was 83 times greater than another Listeria receptor, the monoclonal antibody, mAb-C11E9. Additionally, the capture rate was further increased on a Hsp60-coated biochip by 60% when a dielectrophoresis force was applied for 5 min at the beginning of the final 1 h incubation step. Our data show that Hsp60 could be used for specific detection of L. monocytogenes on a biochip sensor platform.

LAP, an alcohol acetaldehyde dehydrogenase enzyme in Listeria, promotes bacterial adhesion to enterocyte-like Caco-2 cells only in pathogenic species

“…5a) and transmission immunoelectron microscopy ( Fig. 5b), and previously on a biochip sensor, where a Hsp60-coated biochip selectively detected only pathogenic Listeria species, with reduced or no interaction with non-pathogenic species (Koo et al, 2009). Although there is no apparent difference in lap transcript levels for L. monocytogenes and L. innocua strains, the observed differences in protein amounts could be due to either the mRNA not being translated into proteins or a post-transcriptional mechanism preventing protein translation (Johansson et al, 2002;Pelech, 2004).…”

Dielectrophoretic monitoring of microorganisms in environmental applications

“…An application of DEP to improve the efficient capture of target analytes on biosensor platforms was reported in 2009 by Koo et al [97]. In a microfluidic chip, the reliable and specific detection of pathogenic microorganisms is obligated.…”

A Novel Handheld Fluorimeter for Rapid Detection of <italic>Escherichia coli</italic> in Drinking Water

“…Main methods for detection of E. coli in drinking water are based on DNA hybridization, PCR (and qRT-PCR for quantification), immunoassays, immunomagnetic separation, lateral flow tests, incubation micro-chambers as VITEK, labelled nanoparticles, NIRS (near infrared spectroscopy), DEP-FFF (dielectrophoretic field-flow fractionation), or β-Dglucuronidase assays [14][15][16][17][18][19][20][21][22]. But these current technologies are not inexpensive in equipment (as NIRS) or reactives (as labelled antibodies) and usually require an intense water sample manipulation and laboratory equipment (as DNAhybridization), or may need several hours of incubation (β-Dglucuronidase assays) which impairs its portability as well as the economic feasibility of tests every few minutes.…”