Combination of immunosensor detection with viability testing and confirmation using the polymerase chain reaction and culture.
Rapid and accurate differential determination of viable versus nonviable microbes is critical for formulation of an appropriate response after pathogen detection. Sensors for rapid bacterial identification can be used for applications ranging from environmental monitoring and homeland defense to food process monitoring, but few provide viability information. This study combines the rapid screening capability of the array biosensor using an immunoassay format with methods for determination of viability. Additionally, cells captured by the immobilized antibodies can be cultured following fluorescence imaging to further confirm viability and for cell population expansion for further characterization, e.g., strain identification or antibiotic susceptibility testing. Finally, we demonstrate analysis of captured bacteria using the polymerase chain reaction (PCR). PCR results for waveguide-captured cells were 3 orders of magnitude more sensitive than the fluorescence immunoassay and can also provide additional genetic information on the captured microbes. These approaches can be used to rapidly detect and distinguish viable versus nonviable and pathogenic versus nonpathogenic captured organisms, provide culture materials for further analysis on a shorter time scale, and assess the efficacy of decontamination or sterilization procedures.
Johnson-White
Brandy
B
Center for Bio/Molecular Science & Engineering, Naval Research Laboratory, Washington, D.C. 20375-0001, USA.
Lin
Baochuan
B
Ligler
Frances S
FS
D001412:Bacillus subtilis / Q000254:growth & development / Q000302:isolation & purification / Q000472:pathogenicity; D001419:Bacteria / Q000145:classification / Q000254:growth & development / Q000302:isolation & purification* / Q000472:pathogenicity; D015374:Biosensing Techniques; D018929:Cell Culture Techniques / Q000379:methods*; D015169:Colony Count, Microbial; D004586:Electrophoresis / Q000379:methods; D004926:Escherichia coli / Q000254:growth & development / Q000302:isolation & purification / Q000472:pathogenicity; D005516:Food Microbiology; D007118:Immunoassay; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity