Rapid electrochemical detection and identification of catalase positive micro-organisms.
The rapid detection and identification of bacteria has application in a number of fields, e.g. the food industry, environmental monitoring and biomedicine. While in biomedicine the number of organisms present during infection is multiples of millions in the other fields it is the detection of low numbers of organisms that is important, e.g. an infective dose of Escherichia coli O157:H7 from contaminated food is less than 100 organisms. A rapid and sensitive technique has been developed to detect low numbers of the model organism E. coli O55, combining Lateral Flow Immunoassay (LFI) for capture and amperometry for sensitive detection. Nitrocellulose membranes were used as the solid phase for selective capture of the bacteria using antibodies to E. coli O55. Different concentrations of E. coli O55 in Ringers solution were applied to LFI strips and allowed to flow through the membrane to an absorbent pad. The capture region of the LFI strip was placed in close contact with the electrodes of a Clarke cell poised at +0.7 V for the detection of hydrogen peroxide. Earlier research identified that the consumption of hydrogen peroxide by bacterial catalase provided a sensitive indicator of aerobic and facultative anaerobic microorganisms numbers. Modification and application of this technique to the LFI strips demonstrated that the consumption of 8 mM hydrogen peroxide was correlated with the number of microorganisms presented to the LFI strips in the range of 2 x 10(1)-2 x 10(7) colony forming units (cfu). Capture efficiency was dependent on the number of organisms applied and varied from 71% at 2 x 10(2) cfu to 25% at 2 x 10(7) cfu. The procedure was completed in less than 10 min and could detect less than 10 cfu captured from a 200 microl sample applied to the LFI strip. The approached adopted provides proof of principle for the basis of a new technological approach to the rapid, quantitative and sensitive detection of bacteria that express catalase activity.
Sippy
Namrata
N
Faculty of Applied Sciences, Centre for Research in Biomedicine, University of the West of England, Coldharbour Lane, BS16 1QY, Bristol, UK.
Luxton
Richard
R
Lewis
Roger J
RJ
Cowell
David C
DC
D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D002374:Catalase / Q000032:analysis* / Q000378:metabolism*; D015169:Colony Count, Microbial / Q000295:instrumentation / Q000379:methods; D004563:Electrochemistry / Q000295:instrumentation* / Q000379:methods; D004867:Equipment Design; D019544:Equipment Failure Analysis; D004926:Escherichia coli / Q000145:classification / Q000201:enzymology* / Q000302:isolation & purification*; D005506:Food Contamination / Q000032:analysis; D005516:Food Microbiology; D006861:Hydrogen Peroxide / Q000032:analysis / Q000378:metabolism; D007118:Immunoassay / Q000295:instrumentation* / Q000379:methods; D011786:Quality Control; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
D006861:Hydrogen Peroxide; D002374:Catalase