3.1.3 Cells Based Biosensors
Cell-based biosensors have been fabricated to study the level of water pollutants,
which are genotoxic, cytotoxic, membrane or protein-damaging or causing oxidative stress. These microorganisms are genetically modified to exhibit measurable
signals in the form of charge, light or colour as an indicator of water contaminants.
The advantages associated with cell-based toxicity evaluation include low cost,
high sensitivity, rapid response. A biosensor was developed for detection of
non-ionic surfactant nonylphenolethoxylate with a lower detection limit of
0.25 mg/L. The biosensor was analyzed for its ability to detect many organic
compounds, which include surfactants, carbohydrates, polyaromatic compounds
etc. Comamonas testosterone was the microorganism used for detection [30].
The performance of such biosensors is affected by the parameters such as rate of
gene transcription, metabolism, membrane permeability, and protein synthesis.
Whole cell based biosensors can be assayed based on increase and decrease of the
light intensity emitted due to metabolic changes in the microbes in the presence of
specific contaminants. The cells are frequently immobilized on the surface electrode. The immobilization of the cells on the surface of electrodes are governed by
conditions as (1) maintenance of the biological activity of the cells (2) the distance
between the transducers and the biological detection element (3) the constancy and
robustness of the microbial layer (4) the specificity of the detection element
(5) possibility of reuse [31]. The most commonly used methods for immobilization
include cross-linking, adsorption, covalent binding, self-aggregation and entrapment. The covalent binding involves the binding of the biological element to the
transducer with the help of a functional group such as carboxyl, amino, hydroxyl,
sulfhydryl, imidazole and indole.
Premkumar et al. developed a whole cell biosensor by immobilization of
luminescent Escherichia coli strain on aminosylilated glass surface. The immobilization was carried out by aminosylilation of glass surface which was immersed in
glutaraldehyde followed by antibody (Rabbit, Anti-E. coli) solution. The E. coli
suspension was then added and washed with LB medium. E. coli cells were
attached to the optical fibres similarly. A gold-coated glass slide was also prepared
with the same method. The recombinant E. coli strain was prepared by fusion of the
lux genes to a heat shock promoter. The biosensor was analyzed for the detection of
pesticides such as malathion, aldicarb and DDT. The presence of pesticide was
confirmed with the increase in luminescence by the transducer. The repeatability,
reproducibility and stability of the biosensor were studied [32]. A quick and simple
method for detection of lead was proposed, which employed a recombinant strain of
E. coli DH5a containing a regulatory protein and a lead resistance promoter which
also controlled the expression of green fluorescence protein (GFP) reporter gene
[33]. Different types of microbial cell-based biosensors have been developed, such
as optical biosensor having bioluminescence [34], fluorescence [35] or colorimetric
[36] detectors.
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R. Sinha et al.
Cell-based biosensors have been fabricated to study the level of water pollutants,
which are genotoxic, cytotoxic, membrane or protein-damaging or causing oxidative stress. These microorganisms are genetically modified to exhibit measurable
signals in the form of charge, light or colour as an indicator of water contaminants.
The advantages associated with cell-based toxicity evaluation include low cost,
high sensitivity, rapid response. A biosensor was developed for detection of
non-ionic surfactant nonylphenolethoxylate with a lower detection limit of
0.25 mg/L. The biosensor was analyzed for its ability to detect many organic
compounds, which include surfactants, carbohydrates, polyaromatic compounds
etc. Comamonas testosterone was the microorganism used for detection [30].
The performance of such biosensors is affected by the parameters such as rate of
gene transcription, metabolism, membrane permeability, and protein synthesis.
Whole cell based biosensors can be assayed based on increase and decrease of the
light intensity emitted due to metabolic changes in the microbes in the presence of
specific contaminants. The cells are frequently immobilized on the surface electrode. The immobilization of the cells on the surface of electrodes are governed by
conditions as (1) maintenance of the biological activity of the cells (2) the distance
between the transducers and the biological detection element (3) the constancy and
robustness of the microbial layer (4) the specificity of the detection element
(5) possibility of reuse [31]. The most commonly used methods for immobilization
include cross-linking, adsorption, covalent binding, self-aggregation and entrapment. The covalent binding involves the binding of the biological element to the
transducer with the help of a functional group such as carboxyl, amino, hydroxyl,
sulfhydryl, imidazole and indole.
Premkumar et al. developed a whole cell biosensor by immobilization of
luminescent Escherichia coli strain on aminosylilated glass surface. The immobilization was carried out by aminosylilation of glass surface which was immersed in
glutaraldehyde followed by antibody (Rabbit, Anti-E. coli) solution. The E. coli
suspension was then added and washed with LB medium. E. coli cells were
attached to the optical fibres similarly. A gold-coated glass slide was also prepared
with the same method. The recombinant E. coli strain was prepared by fusion of the
lux genes to a heat shock promoter. The biosensor was analyzed for the detection of
pesticides such as malathion, aldicarb and DDT. The presence of pesticide was
confirmed with the increase in luminescence by the transducer. The repeatability,
reproducibility and stability of the biosensor were studied [32]. A quick and simple
method for detection of lead was proposed, which employed a recombinant strain of
E. coli DH5a containing a regulatory protein and a lead resistance promoter which
also controlled the expression of green fluorescence protein (GFP) reporter gene
[33]. Different types of microbial cell-based biosensors have been developed, such
as optical biosensor having bioluminescence [34], fluorescence [35] or colorimetric
[36] detectors.
194
R. Sinha et al.