Rapid pathogen detection by lateral-flow immunochromatographic assay with gold nanoparticle-assisted enzyme signal amplification.

To date most LF-ICA format for pathogen detection is based on generating color signals from gold nanoparticle (AuNP) tracers that are perceivable by naked eye but often these methods exhibit sensitivity lower than those associated with the conventional enzyme-based immunological methods or mandated by the regulatory guidelines. By developing AuNP avidin-biotin constructs in which a number of enzymes can be labeled we report on an enhanced LF-ICA system to detect pathogens at very low levels. With this approach we show that as low as 100 CFU/mL of Escherichia coli O157:H7 can be detected, indicating that the limit of detection can be increased by about 1000-fold due to our signal amplification approach. In addition, extensive cross-reactivity experiments were conducted (19 different organisms were used) to test and successfully validate the specificity of the assay. Semi-quantitative analysis can be performed using signal intensities which were correlated with the target pathogen concentrations for calibration by image processing.

[1]  H. Anany,et al.  Bacteriophages for detection and control of bacterial pathogens in food and food-processing environment. , 2012, Advances in food and nutrition research.

[2]  J. Paul Robinson,et al.  Discovering the unknown: Detection of emerging pathogens using a label‐free light‐scattering system , 2010, Cytometry. Part A : the journal of the International Society for Analytical Cytology.

[3]  Parin Chaivisuthangkura,et al.  Development and evaluation of a highly sensitive immunochromatographic strip test using gold nanoparticle for direct detection of Vibrio cholerae O139 in seafood samples. , 2013, Biosensors & bioelectronics.

[4]  Joseph Irudayaraj,et al.  Biofunctionalized magnetic nanoparticle integrated mid-infrared pathogen sensor for food matrixes. , 2009, Analytical chemistry.

[5]  Steven Ripp,et al.  Pathogen detection using engineered bacteriophages , 2012, Analytical and Bioanalytical Chemistry.

[6]  B. Swaminathan,et al.  Rapid detection of food-borne pathogenic bacteria. , 1994, Annual review of microbiology.

[7]  Joseph Maria Kumar Irudayaraj,et al.  Silver Nanosphere SERS Probes for Sensitive Identification of Pathogens , 2010 .

[8]  Il-Hoon Cho,et al.  Lateral-flow enzyme immunoconcentration for rapid detection of Listeria monocytogenes , 2013, Analytical and Bioanalytical Chemistry.

[9]  J. Irudayaraj,et al.  Multifunctional magnetic-optical nanoparticle probes for simultaneous detection, separation, and thermal ablation of multiple pathogens. , 2010, Small.

[10]  Khalil Arshak,et al.  An overview of foodborne pathogen detection: in the perspective of biosensors. , 2010, Biotechnology advances.

[11]  J. Fontecha,et al.  A spectrophotometric assay for biotinbinding sites of immobilized avidin , 1996 .

[12]  K. A. Razak,et al.  The effects of size and synthesis methods of gold nanoparticle-conjugated MαHIgG4 for use in an immunochromatographic strip test to detect brugian filariasis , 2012, Nanotechnology.

[13]  P. Fratamico,et al.  PCR detection and microbiological isolation of Salmonella spp. from fresh beef and cantaloupes. , 2009, Journal of food science.

[14]  R. Reissbrodt.  New chromogenic plating media for detection and enumeration of pathogenic Listeria spp.--an overview. , 2004, International journal of food microbiology.

[15]  Il-Hoon Cho,et al.  In-situ fluorescent immunomagnetic multiplex detection of foodborne pathogens in very low numbers. , 2014, Biosensors & bioelectronics.

[16]  Peter Rådström,et al.  Standardization of diagnostic PCR for the detection of foodborne pathogens. , 2003, International journal of food microbiology.

[17]  S. Baidoo,et al.  Use of chicken egg-yolk antibodies against K88+ fimbrial antigen for quantitative analysis of enterotoxigenic Escherichia coli (ETEC) K88+ by a sandwich ELISA , 1999 .

[18]  C. Suri,et al.  Immuno-fluorescence based Vi capsular polysaccharide detection for specific recognition of Salmonella enterica serovar Typhi in clinical samples. , 2014, Analytica chimica acta.

[19]  Tennant,et al.  Rapid and definitive detection of Salmonella in foods by PCR , 1998, Letters in applied microbiology.

[20]  J. Paul Robinson,et al.  Label-free detection of multiple bacterial pathogens using light-scattering sensor. , 2009, Biosensors & bioelectronics.

[21]  Arun K Bhunia,et al.  Biosensors and bio-based methods for the separation and detection of foodborne pathogens. , 2008, Advances in food and nutrition research.

[22]  Joseph Irudayaraj,et al.  Separation and detection of multiple pathogens in a food matrix by magnetic SERS nanoprobes , 2011, Analytical and bioanalytical chemistry.

[23]  Il-Hoon Cho,et al.  An enzyme immunoanalytical system based on sequential cross-flow chromatography. , 2005, Analytical chemistry.

[24]  Il-Hoon Cho,et al.  Plastic ELISA-on-a-chip based on sequential cross-flow chromatography. , 2006, Analytical chemistry.

[25]  Il-Hoon Cho,et al.  Membrane filter-assisted surface enhanced Raman spectroscopy for the rapid detection of E. coli O157:H7 in ground beef. , 2015, Biosensors & bioelectronics.

[26]  Joseph Irudayaraj,et al.  A mixed self-assembled monolayer-based surface plasmon immunosensor for detection of E. coli O157:H7. , 2006, Biosensors & bioelectronics.

[27]  G. Węgrzyn,et al.  Specific detection of Salmonella enterica and Escherichia coli strains by using ELISA with bacteriophages as recognition agents , 2011, European Journal of Clinical Microbiology & Infectious Diseases.

[28]  M. Griffiths,et al.  Development and Characterization of a Fluorescent-Bacteriophage Assay for Detection of Escherichia coli O157:H7 , 1999, Applied and Environmental Microbiology.