Monitoring biosensor capture efficiencies: development of a model using GFP-expressing Escherichia coli O157:H7.
One of the known limitations for biosensor assays is the high limit of detection for target cells within complex samples (e.g., Escherichia coli at 10(4) to 10(5) CFU/mL) due to poor capture efficiencies. Currently, researchers can only estimate the cell capture efficiency necessary to produce a positive signal for any type of biosensor using either cumbersome techniques or regression modeling. To solve this problem, green fluorescent protein (GFP) transformed E. coli O157:H7 was used to develop a novel method for directly and easily measuring the cell capture efficiency of any given biosensor platform. For demonstration purposes, E. coli-GFP was assayed on both fiber optic and planar waveguide biosensor platforms. Cells were enumerated using an epifluorescent microscope and digital camera to determine the number of cells captured on the surfaces. Conversion algorithms were used with these digital images to determine the cell density of entire waveguide surface areas. For E. coli-GFP, the range of cell capture efficiency was between 0.4 and 1.2%. This indicates that although the developed model works for calculating cell capture, there is still need for significant improvements in capture methods themselves, to increase the capture efficiency and thereby lower detection limits. The use of GFP-transformed target cells and cell capture efficiency calculations can facilitate the development and optimization processes by allowing direct enumeration of new biosensor design configurations and sample processing strategies.
Simpson-Stroot
Joyce M
JM
Division of Cell Biology, Microbiology, and Molecular Biology, Department of Biology, University of South Florida, Tampa, FL 33620-5200, USA. jstroot@cas.usf.edu
Kearns
Elizabeth A
EA
Stroot
Peter G
PG
Magaña
Sonia
S
Lim
Daniel V
DV
D000465:Algorithms; D000907:Antibodies, Bacterial / Q000276:immunology; D015374:Biosensing Techniques; D019453:Escherichia coli O157 / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification* / Q000378:metabolism; D005336:Fiber Optic Technology; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism*; D007118:Immunoassay; D008856:Microscopy, Fluorescence; D055100:Optical Fibers; D014169:Transformation, Bacterial
D000907:Antibodies, Bacterial; D049452:Green Fluorescent Proteins