Biomedical Engineering Reference
In-Depth Information
increased current of a redox indicator or by changes in conductivity or capacitance. In this
chapter, however, we would like to concentrate on the use of LAPS, which are proving to
be popular as a platform for detecting microbes. These are semiconductor-based systems
with an electrolyte-insulator-semiconductor (EIS) structure. When a current is applied
across the EIS region, a depletion layer is formed at the insulator-semiconductor interface
[98,99]. The capacitance of the depletion layer changes with the surface potential, which is
a function of the ion concentration in the electrolyte. To determine the capacitance, the
semiconductor is illuminated by modulated light and the current is measured. LAPS have
several advantages when compared with other sensors: the surface is flat, there is no need
for wires or passivation, and they can measure pH and concentration. Researchers at the
USDA have used a LAPS system in combination with an immunoligand assay (ILA) to
detect live
E. coli
O157:H7. They have reported that both live and dead bacteria can be
detected in 30-45 min. In this system, bacteria are captured onto a filter membrane by
using specific antibodies [100]. A silicone-based sensor is then placed adjacent to the mem-
brane and upon illumination small changes in acidity are detected. The signal is propor-
tional to the number of bacteria present, and it was possible to detect 710 dead or 25,000
live
E. coli
O157:H7 organisms/ml [101]. In a recent development, a LAPS approach was
used to detect
E. coli
in drinking water [94].
An immunoassay was developed such that there was specificity to a particular capsular
protein present in the bacterium. The transducer, based on the LAPS principle, was able to
detect the production of ammonia by urease-
E. coli
-antibody conjugate. It was claimed that
10 cells/ml were detected in 1.5 h. Generally, amperometric biosensors work by enzymati-
cally generating current between two electrodes. They have fast response time, dynamic
ranges, and sensitivities similar to potentiometric biosensors. Many amperometric biosen-
sors depend on dissolved oxygen concentration that can pose a major problem. To over-
come this situation, mediators are employed. These mediators transfer electrons directly to
the electrode thereby eliminating the need for the reduction of an oxygen cosubstrate [102].
The most commonly used mediators are the ferrocenes. Amperometric biosensors have
been used for the detection of
E. coli
in water (screen-printed electrodes), bacterial
vaginosis, studies of bacterial contamination, detection of agents of biological warfare
(e.g., anthrax) and
E. coli
heat-labile enterotoxin, and other neurotoxins. Amperometric
biosensors [103,104] have also been used to study bacterial luciferase reactions, nanoscale
bacterial surface proteins and growth, and viability of bacterial populations.
Many biosensors are based on the
UIDA
gene, or the
-D-glucuronidase (GLUase)
enzyme for which it encodes [105]. Although it is possible to target the
UIDA
gene directly
[106], usually GLUase activity is used as an enzymatic marker for the identification of
Salmonella and Shigella
. The GLUase activity is detected by its ability to cleave specific
chromogenic or fluorogenic artificial substrates added to the culture medium [107] or
directly to filtered cells [108]. This approach has led to considerably faster and specific
methods for the detection of
Salmonella and Shigella
contamination in water, although the
analysis time is still measured in hours. Also, it still requires either extensive manipulation
and incubation time [109-112] or sophisticated equipment [108].
According to the recent literatures cited, the method of immunoassay has become rapidly
popular due to factors like high sensitivity, decrease in the time of analysis, and reproducible
data that is reliable. The class of amperometric biosensors has become extremely successful
as they can be used under field conditions without the need for skilled personnel. With these
amperometric biosensors, the pathogens can be detected directly in the original samples
without any preenrichment of the sample. Recently developed techniques for the detection
of
Salmonella
and
Shigella
include methods based on the integration of several technologies
like the microelectromechanical systems (MEMS), self-assembled monolayer (SAMS), DNA
hybridization, and enzyme amplification [113]
.
The group of Jen-Jr Gau has tried to detect
