Biosensors or enzyme electrodes

INTRODUCTION OF BIOSENSORS:-

Biosensors or enzyme electrodes invariably refer to such devices that sence and analyze biological informations. A biosensor is a device that detects, records, and transmits information regarding a physiological change or the presence of various chemical or biological materials in the environment. More technically, a biosensor is a probe that integrates a biological component, such as a whole bacterium or a biological product (e.g., an enzyme or antibody) with an electronic component to yield a measurable signal. Biosensors, which come in a large variety of sizes and shapes, are used to monitor changes in environmental conditions. They can detect and measure concentrations of specific bacteria or hazardous chemicals; they can measure acidity levels (pH). In short, biosensors can use bacteria and detect them, too.

PRINCIPALS OF BIOSENSORS:-

A biosensor essentially comprise of the following two major parts

1- Biological component-

For sensing the presence as well as concentration of analyte. In the presence of a certain molecule the biological system changes the environment. The measuring device sensitive to this change sends a signal. This signal can be converted into the measurement parameter. Often the biological system is an actual cell. The key thing to remember is that it is an actual organism that detects the concentration change of the molecule in the media. This organism could be the same one as the one in the media or it could be different. In either case it must be kept separate from the media. This can be done with a membrane that is permeable to the molecule that is being measured but impermeable to the cells and most other macromolecules in the reactor media.

2- Physical component:-

Transducer:-

A device that converts energy from one form into another e.g., telephone companies use transducers to convert sound energy into electrical energy to be carried long- distance through telephone lines and then another transducer at the receiving end to convert the electrical energy back into sound A biosensor is a sensing device that consists of a biological component coupled to a transducer that converts biochemical activity into, most commonly, electrical energy.

Types of Biosensors:-

There are different types of biosensors, which have different applications. These are listed below.

1. Calorimetric biosensor
2. Potentiometric biosensor
3. Amperometric biosensors
4. Optical biosensor
5. Acoustic wave biosensors

Potentiometric Biosensors

At the transducer surface, an electrical potential is produced due to changed distribution of electrons and this type of biosensors are called potentiometric biosensors. They use ion sensitive electrodes, commonly pH meter glass electrodes for cations, glass pH electrodes coated with a gas-selective membrane for CO2, NH3 or H2S or solid-state electrodes. These electrodes convert the biological reaction into electric signal.

Optical Biosensor

A most promising optical biosensor utilizes luminescence due to firefly luciferase for detection of bacteria in food or clinical samples. The bacteria are specifically lysed to release ATP. This ATP is used by luciferase in the presence of O2 to produce light, which is measured by the biosensor.

Acoustic wave biosensors

Acoustic wave biosensors sense the change in mass of the biological components as a result of the reaction.

They are also called piezoelectric devices. The surface of the transducer is usually coated with antibodies which bind to the complementary antigen present in the sample solution. The resulting increase in mass reduces their frequency of vibration. This change in frequency is measured in terms of antigen present in the sample solution.

Applications of Biosensor:-

1. Health Care

Measurement of Metabolites

The initial impetus for advancing sensor technology came from health care area, where it is now generally recognized that measurements of blood gases, ions and metabolites are often essential and allow a better estimation of the metabolic state of a patient. In intensive care units for example, patients frequently show rapid variations in biochemical levels that require an urgent remedial action. Also, in less severe patient handling, more successful treatment can be achieved by obtaining instant assays. At present, the list of the most commonly required instant analyses is not extensive. In practice, these assays are performed by analytical laboratories, where discrete samples are analyzed, frequently using the more traditional analytical techniques.

Market Potential.

There is an increasing demand for inexpensive and reliable sensors to allow not only routine monitoring in the central or satellite laboratory, but also analysis with greater patient contact, such as in the hospital ward, emergency rooms, and operating rooms. Ultimately, patients themselves should be able to use biosensors in the monitoring and control of some treatable condition, such as diabetes. It is probably true to say that the major biosensor market may be found where an immediate assay is required. If the cost of laboratory maintenance are counted with the direct analytical costs, then low-cost biosensor devices can be desirable in the whole spectrum of analytical applications from hospital to home.

Diabetes.

The ‘classic’ and most widely explored example of closed-loop drugcontrol is probably to be found in the development of an artificial pancreas. Diabetic patients have a relative or absolute lack of insulin, a polypeptide hormone produced by the beta-cells of the pancreas, which is essential to the metabolism of a number of carbon sources. This deficiency causes various metabolic abnormalities, including higher than normal blood glucose levels. For such patients, insulin must be supplied externally. This has usually been achieved by subcutaneous injection, but fine control is difficult and hyperglycaemia cannot be totally avoided, or even hypoglycaemia is sometimes induced, causing impaired consciousness and the serious long-term complications to tissue associated with this intermittent low glucose condition.

Insulin Therapy.

Better methods for the treatment of insulin-dependent diabetes havebeen sought and infusion systems for continuous insulin delivery have been developed. However, regardless of the method of insulin therapy, its induction must be made in response to information on the current blood glucose levels in the patient. Three schemes are possible (Fig. 1.6), the first two dependent on discrete manual glucose measurement and the third a ‘closed-loop’ system, where insulin delivery is controlled by the output of a glucose sensor which is integrated with the insulin infuser. In the former case, glucose has been estimated on ‘finger-prick’ blood samples with a colorimetric test strip or more recently with an amperometric ‘pen’-size biosensor device by the patient themselves. Obviously these diagnostic kits must be easily portable, very simple to use and require the minimum of expert interpretation. However, even with the ability to monitor current glucose levels, intensive conventional insulin therapy requires multiple daily injections and is unable to anticipate future states between each application, where diet and exercise may require modification of the insulin dose. For example, it was shown that administration of glucose by subcutaneous injection, 60 min before a meal provides the best glucose/insulin management.

Artificial Pancreas.

The introduction of a closed-loop system, where integrated glucose measurements provide feedback control on a pre-programmed insulin administration based on habitual requirement, would therefore relieve the patient of frequent assay requirements and perhaps more desirably frequent injections. Ultimately, the closed-loop system becomes an artificial pancreas, where the glycaemic control is achieved through an implantable glucose sensor. Obviously, the requirements for this sensor are very different to those for the discrete measurement kits. As summarized in Table 1.4, the prolonged life-time and biocompatibility represent the major requirements.

2. Industrial Process Control

Bioreactor Control. Real-time monitoring of carbon sources, dissolved gases,. in fermentation processes (Fig. 1.7a) could lead to optimization of the procedure giving increased yields at decreased materials cost. While real-time monitoring with feedback control involving automated systems does exist, currently only a few common variables are measured on-line (e.g. pH, temperature, CO2, O2)) which are often only indirectly related with the process under control.

Seven requirements for an implantable glucose sensor.

• Linear in 0 – 20 mM range with 1 mM resolution
• Specific for glucose; not affected by changes in metabolite concentrations and ambient conditions
• Biocompatible
• Small—causes minimal tissue damage during insertion and there is better patient
• acceptability for a small device
• External calibration and < 10% drift in 24h
• Response time < 10 min
• Prolonged lifetime-at least several days, preferably weeks in use

Three different methods of controlling a bioreactor are:

1. Off-line distant: central laboratory coarse control with significant time lapse
2. Off-line local: fine control with short time lapse
3. On-line: real-time monitoring and control

On-Line Control.

Method 3 is most desirable, which allows the process to follow an ideal pre-programmed fermentation profile to give maximum output. However, many problems exist with on-line measurements including in situ sterilization, sensor life-time, sensor fouling, etc. Some of the problems can be overcome if the sensor is situated so that the sample is run to waste, but this causes a volume loss, which can be particularly critical with small volume fermentations.

Off-Line Control.

Although Method 3 may be the ultimate aim, considerable advantage can be gained in moving from Method 1 to Method 2 giving a rapid analysis and thus enabling finer control of the fermentation. The demands of the sensor are perhaps not as stringent in Method 2 as in Method 3.

Benefits of Control.

The benefits which are achievable with process-control technology are considerable:

• Improved product quality; reduction in rejection rate following manufacture
• Increased product yield; process tuned in real time to maintain optimum conditions throughout and not just for limited periods
• Increased tolerance in quality variation of some raw materials. These variations can be compensated in the process-control management
• Reduced reliance on human ‘seventh sense’ to control process
• Improved plant performance-processing rate and line speed automated, so no unnecessary dead-time allocated to plant
• Optimized energy efficiency
• The use of biosensors in industrial process in general could facilitate plant automation, cut analysis costs and improve quality control of the product.

3. Military Applications

Dip Stick Test.

The requirement for rapid analysis can also be anticipated in military applications. The US army, for example, have looked at dipstick tests

Summary of potential applications for biosensors

• Clinical diagnosis and biomedicine
• Farm, garden and veterinary analysis
• Process control: fermentation control and analysis food and drink
• production and analysis
• Microbiology: bacterial and viral analysis
• Pharmaceutical and drug analysis
• Industrial effluent control
• Pollution control and monitoring o Mining, industrial and toxic gases
• Military applications

based on monoclonal antibodies. While these dipsticks are stable and highly specific (to Q-fever, nerve agents, yellow rain fungus, soman, etc.) they are frequently two-step analyses taking up to 20 min to run. Such a time lapse is not always suited to battlefield diagnostics; the resulting consequences are suggested in Fig. 1.7(c).

A particularly promising approach to this unknown hazard detection seems to be via acetylcholine receptor systems. It has been calculated that with this biorecognition system, a matrix of 13-20 proteins are required to give 95% certainty of all toxin detection.

4. Environmental Monitoring

Air and Water Monitoring.

Another assay situation which may involve a considerable degree of the unknown is that of environmental monitoring. The primary measurement media here will be water or air, but the variety of target analytes is vast. At sites of potential pollution, such as in factory effluent, it would be desirable to install on-line real-time monitoring and alarm, targeted at specific analytes, but in many cases random or discrete monitoring of both target species or general hazardous compounds would be sufficient. The possible analytes include biological oxygen demand (BOD) which provides a good indication of pollution, atmospheric acidity, and river water pH, detergent, herbicides, and fertilizers (organophosphates, nitrates, etc.). The survey of market potential has identified the increasing significance of this area and this is now substantiated by a strong interest from industry. The potential applications of biosensors are summarized in Table 1.4.

Tuning to Application.

The potential for biosensor technology is enormous and is likely to revolutionize analysis and control of biological systems. It is possible therefore to identify very different analytical requirements and biosensor developments must be viewed under this constraint. It is often tempting to expect a single sensor targeted at a particular analyte, to be equally applicable to on-line closed-loop operation in a fermenter and pin-prick blood samples. In practice, however, the parallel development of several types of sensor, frequently employing very different measurement parameters is a more realistic.

Advantages of biosensors over other measurement schemes

1. They can measure nonpolar molecules that do not repond to most measurement devices.
2. They are as specific as the immobilized system used in them.
3. They allow rapid continuous control.

Disadvantages of biosensors

• Heat sterilization is not possible as this would denature the biological part of the biosensor.
• The membrane that separates the reactor media from the immobilized cells of the sensor can become fouled by deposits.
• The cells in the biosensor can become intoxicated by other molecules that are capable of diffusing through the membrane
• Changes in the reactor broth (i.e., pH) can put chemical and mechanical stress on the biosensor that might eventually impair it.

Future of Biosensors:-

Biosensors have the potential to affect many areas. Field application areas including medicine, physical therapy, music, and the video game industry, can all benefit from the introduction of biosensors. Although biosensors are not limited to any group of people, they are particulary useful for the handicapped. Even completely paralyzed individuals have electrical activity in their bodies that can be detected.

One biosensor application developed for the handicapped is an electronic instrument that produces music from bioelectric signals. Signal inputs such as eye movements, muscle tensions, and muscle relaxations are converted to MIDI (Musical Instrument Data Interface) and output to a synthesizer. Before being mapped to MIDI, the signals are analyzed for specific intensity and spectral characteristics for the particular individual. For dysfunctional or weak muscles the signals can be amplified according the the level of tension and relaxation. These signal inputs are then interpreted to control volume, pitch, tempo, and other aspects of musical composition.

Medical applications are presently seen in the diagnosis and correction of eye disorders . Strabismus is a condition in which an individual’s eyes are not aligned properly, and thus do not move in conjunction with one another. This can be corrected by surgery but the current use of prisms to determine the degree of correction necessary is not very accurate. Biosensors tracking the eye movements can determine with high accuracy the number of degrees in both the X and Y planes that the eyes need to be adjusted.

Just as biosensors can be used to determine amounts of eye correction, they can also be used to train the eye as they can be an input device to video game exercises to realign eye tracking. This same method of muscle training through a video game could be used for rehabilitation of potentially any muscle group, as biosensors can be individually customized to detect levels of muscle activity for most muscle groups. In the same way that patients undergoing rehabilitation could use biosensors as an input device for their video exercises, the video game industry could use biosensors as yet another powerful input device for entertainment.

Also contributing to physical therapy, biosensors can help to create custom exercise programs for injured patients and athletes, can be used by athletes to check muscle condition, and can be connected to a multitude of external monitoring devices.

Some Future Goals :-

There are future applications that make biosensors ideal input devices. Eye tracking devices that can focus and select objects in 3D virtual environments would couple sight and limb 3D selection creating powerful immersive environments. The laser abilities from the eyes of ‘Superman’ could be realized by users in a virtual environment.

Possible use of prosthetic limbs where just the bioelectric activity to the nerve endings of a missing limb could be used to control an artificial limb. In cases of paralysis, the nerves, prior to loss of transport ability, or brainwaves might be electrically monitored for instructions to control/move a mechanical device attached to the paralyzed limb.

When brainwaves can be reliably monitored, we can study relationships between EEG (brain activity) and specific cognitive activities such as sleep behaviors and sleep states. Simple brain wave detection has been successful in early research stages, but breaking through the use of subvocal commands would be perhaps the most powerful input controller we have yet seen. Just picture monitoring brain activity so that when you think “draw a circle”, a circle appears on your monitor or in your virtual environment.