A biosensor is an instrument that is sensitive to biological substances and converts their concentration into electrical signals for detection. It is made of immobilized biologically sensitive materials as identification elements (including enzymes, antibodies, antigens, microorganisms, cells, tissues, nucleic acids and other biologically active substances), appropriate physical and chemical transducers (such as oxygen electrodes, photosensitive tubes, field effect tubes, Piezoelectric crystals, etc.) and analysis tools or systems composed of signal amplification devices. Biosensors have the functions of receptors and converters.
composition structure
Biosensors consist of a molecular recognition part (sensitive element) and a conversion part (transducer):
*Identifying the target with the molecular recognition part is the main functional element that can cause a certain physical or chemical change. Molecular recognition moieties are the basis for selective assays in biosensors.
* Physical or chemical transducers (sensors) that convert biologically expressed signals into electrical signals
Various biosensors have the following common structure: including one or several related biologically active materials (biofilms) and physical or chemical transducers (sensors) that can convert the signals expressed by biological activity into electrical signals, the combination of the two Together, the reprocessing of biological signals is carried out with modern microelectronics and automated instrumentation technology to form various biosensor analysis devices, instruments and systems that can be used.
Biosensors fulfill the following three functions:
*Feeling: Extract the biological materials that animals and plants play a role in perception, including: biological tissues, microorganisms, organelles, enzymes, antibodies, antigens, nucleic acids, DNA, etc. Realize the mass production of biomaterials or biomaterials, reuse them repeatedly, and reduce the difficulty and cost of detection.
*Observation: Converting continuous, regular information felt by biological materials into information that people can understand.
*Reaction: Display information to people through optical, piezoelectric, electrochemical, temperature, electromagnetic, etc., to provide a basis for people’s decision-making.
The main function
Biosensors have the functions of receptors and converters. An instrument that is sensitive to biological substances and converts their concentration into electrical signals for detection.
Substances that can selectively distinguish specific substances in living organisms include enzymes, antibodies, tissues, cells, and the like. These molecular recognition functional substances can combine with the target to be detected into complexes through the recognition process, such as the combination of antibodies and antigens, and the combination of enzymes and substrates.
When designing a biosensor, it is an extremely important premise to select a substance with a recognition function suitable for the object to be measured. The properties of the resulting complex are taken into account. According to the chemical changes or physical changes caused by the sensitive elements prepared by molecular recognition functional substances, the selection of transducers is another important link in the development of high-quality biosensors. The generation or consumption of light, heat, and chemical substances in the sensitive element will produce corresponding changes. Based on these variations, an appropriate transducer can be selected.
The information generated by the biochemical reaction process is diverse, and the achievements of microelectronics and modern sensing technology have provided abundant means for detecting this information.
History
In 1967, S.J. Updick et al. produced the first biosensor glucose sensor. Glucose oxidase was contained in polyacrylamide colloid for solidification, and then the colloid film was fixed on the tip of the diaphragm oxygen electrode to make a glucose sensor. Other sensors that detect their counterparts can be fabricated when other immobilized films such as enzymes or microorganisms are used. The methods of immobilizing the sensing membrane include direct chemical bonding method; polymer carrier method; polymer membrane bonding method. The second generation of biosensors (microorganisms, immunity, enzyme immunity and organelle sensors) have been developed, and the third generation of biosensors, field-effect biosensors that combine systems biotechnology and electronic technology, were developed in the 1990s. Fluidics technology and microfluidic chip integration of biosensors provide new technological prospects for drug screening and genetic diagnosis. Biosensors have very high selectivity because enzyme membranes, mitochondrial electron transport system particle membranes, microbial membranes, antigen membranes, and antibody membranes have selective recognition functions for the molecular structure of biological substances, and only catalyze and activate specific reactions. The disadvantage is that the biocured film is not stable. Biosensors involve biological substances and are mainly used in clinical diagnostic examinations, monitoring during treatment, fermentation industry, food industry, environment and robotics.
Biosensor is an interdisciplinary subject that organically combines bioactive materials (enzymes, proteins, DNA, antibodies, antigens, biofilms, etc.) with physical and chemical transducers, and is an advanced detection method indispensable for the development of biotechnology. It is also a rapid and trace analysis method at the molecular level of substances. In the future development of knowledge economy in the 21st century, biosensor technology will definitely be a new growth point between information and biotechnology, in clinical diagnosis, industrial control, food and drug analysis (including biopharmaceutical research and development) in the national economy. It has a wide range of application prospects in researches such as environmental protection, biotechnology and biochips.
Technical Features
A sensor is a special device that can acquire and process information. For example, the human body’s sensory organs are a set of perfect sensing systems to perceive physical information such as light, sound, temperature, and pressure from the outside world through eyes, ears, and skin. The tongue senses chemical stimuli such as smell and taste. Biosensors are a special kind of sensors, which use biologically active units (such as enzymes, antibodies, nucleic acids, cells, etc.) as biologically sensitive units, and are highly selective detectors for target substances.
(1) Immobilized biologically active substances are used as catalysts, and expensive reagents can be used repeatedly, which overcomes the disadvantages of high cost of enzymatic analysis reagents and cumbersome and complicated chemical analysis in the past.
(2) It has strong specificity, only reacts to specific substrates, and is not affected by color and turbidity.
(3) The analysis speed is fast, and the result can be obtained in one minute.
⑷ High accuracy, generally the relative error can reach 1%
⑸ The operating system is relatively simple, and it is easy to realize automatic analysis
⑹ Low cost, in continuous use, only a few cents of RMB are needed for each measurement.
⑺Some biosensors can reliably indicate the oxygen supply status and the production of by-products in the microbial culture system. In production control, information that can only be obtained by the combined action of many complex physical and chemical sensors can be obtained. At the same time they also indicated the direction of increasing the product yield.
Equipment classification
Sensors that use immobilized biological components or organisms as sensitive elements are called biosensors. Biosensors do not specifically refer to sensors used in the field of biotechnology. Its application areas also include environmental monitoring, medical and health care, and food inspection. Biosensors mainly have the following three classification and naming methods:
1. According to the molecular recognition elements in biosensors, that is, sensitive elements can be divided into five categories: enzyme sensors (enzymesensor), microbial sensors (microbialsensor), cell sensors (organallsensor), tissue sensors (tis-suesensor) and immune sensors (immunolsensor). Obviously, the applied sensitive materials are enzymes, microorganisms, organelles, animal and plant tissues, antigens and antibodies in order.
2. According to the transducers of biosensors, namely signal converters, they are classified as: bioelectrode sensors, semiconductor biosensors, optical biosensors, thermal biosensors (calorimetric biosensor), piezoelectric crystal biosensors (piezoelectric biosensor) The transducers are electrochemical electrodes, semiconductors, photoelectric converters, thermistors, piezoelectric crystals, etc.
3. There are bioaffinity biosensors (affinity biosensors), metabolic biosensors or catalytic biosensors based on the interaction between the target and the molecular recognition element.
The three classification methods are actually used interchangeably.
Application field
Overview
Biosensor is a high-tech developed by the mutual penetration of various disciplines such as biology, chemistry, physics, medicine, and electronic technology. Because of its good selectivity, high sensitivity, fast analysis speed, low cost, and online continuous monitoring in complex systems, especially its features of high automation, miniaturization and integration, it has been widely used in recent decades. flourishing and rapid development.
It has broad application prospects in various sectors of the national economy such as food, pharmaceuticals, chemical industry, clinical testing, biomedicine, and environmental monitoring. In particular, the combination of molecular biology and new disciplines and technologies such as microelectronics, optoelectronics, microfabrication technology and nanotechnology is changing the face of traditional medicine, environmental science, and zoology and botany. The research and development of biosensors has become a new hot spot in the world’s scientific and technological development, forming an important part of the emerging high-tech industry in the 21st century, and has important strategic significance.
food industry
The application of biosensors in food analysis includes the determination and analysis of food components, food additives, harmful poisons and food freshness.
⑴ food composition analysis In the food industry, the content of glucose is an important indicator to measure the maturity and storage life of fruits. The developed enzyme-electrode biosensor can be used to analyze glucose in liquor, apple juice, jam and honey. Other sugars, such as fructose, maltose in beer and wort, also have mature measurement sensors.
Niculescu et al. developed an amperometric biosensor that can be used to detect ethanol content in beverages. In this biosensor, a ligand protein alcohol dehydrogenase is embedded in polyethylene. Different ratios of enzyme and polymer can affect the performance of the biosensor. In the experiments conducted, the measurement limit of this biosensor for ethanol was 1 nmol/L.
(2) Analysis of food additives
Sulfite is usually used as a bleaching agent and preservative in the food industry. The current-type sulfur dioxide enzyme electrode made of sulfite oxidase as a sensitive material can be used to determine the content of sulfite in food. The linear range of the determination is 0～ 6 to the negative fourth power mol/L. Another example is the sweetener in beverages, pudding, vinegar and other foods. Guibault et al. used aspartase combined with ammonia electrode to determine the linear range of 2 × 10 negative fifth power to 1 × 10 negative third power mol/L. In addition, there are also reports of using biosensors to measure pigments and emulsifiers.
(3) Analysis of pesticide residues
People are paying more and more attention to the problem of pesticide residues in food, and governments around the world are constantly strengthening the detection of pesticide residues in food.
Yamazaki et al. invented an amperometric biosensor for the determination of organophosphorus pesticides using artificial enzymes, using organophosphorus pesticide hydrolase, p-nitrophenol and diethylphenol with a detection limit of minus 10 to the seventh power mol, it only takes 4 minutes to measure at 40 °C. Albareda et al. used glutaraldehyde cross-linking method to immobilize acetylcholineesterase on the surface of copper wire carbon paste electrode to make a detectable concentration of 10 minus tenth mol/L of paraoxon and 10 minus tenth mol/L. The biosensor of 11 mol/L Carbosvir can be used to directly detect the residues of two pesticides in tap water and fruit juice samples.
⑷ Inspection of microorganisms and toxins
The existence of pathogenic microorganisms in food will bring great harm to the health of consumers. There are not only many types of toxins in food, but also high toxicity. Most of them have carcinogenic, teratogenic and mutagenic effects. The detection of microorganisms and toxins is crucial.
Edible beef is easily infected by Escherichia coli 0157.H7. Therefore, rapid and sensitive methods are needed to detect and defend against bacteria such as Escherichia coli 0157.H7. The fiber-optic biosensors studied by Kramerr et al. can detect pathogens (such as E. coli 0157.H7.) in food in minutes, compared with days for traditional methods. The biosensor took just 1 day from detecting the pathogen to recovering the pathogen from the sample and allowing it to grow independently on the medium, compared to 4 days for the traditional method.
There is also a fast and sensitive immunobiosensor that can be used to measure dihydropyridoxine residues in milk, which is based on cytoplasmic genome responses that transmit signals through an optical system. The detection limit achieved was 16.2 ng/mL. 20 milk samples can be tested in a day.
⑸ Detection of food freshness
In the food industry, the freshness detection of food, especially fish and meat, is a major indicator for evaluating food quality. Volpe et al. used Huangxiong oxidase as a biologically sensitive material, combined with a hydrogen peroxide electrode, and measured the levels of inosine monophosphate (IMP), inosine (HXR) and hypoxanthine (HX) produced during fish degradation. Concentration, so as to evaluate the freshness of fish, the linear range is 5×10 minus 10th power to 2×10 minus 4th power mol/L.
Environmental monitoring
The problem of environmental pollution is becoming more and more serious, and people are eager to have an instrument that can monitor pollutants continuously, rapidly and online. Biosensors meet people’s requirements. A considerable number of biosensors have been used in environmental monitoring.
⑴Water environment monitoring
Biochemical oxygen demand (BOD) is a widely used comprehensive indicator to characterize the degree of organic pollution. Biochemical oxygen demand is also one of the most commonly used and important indicators in water body monitoring and operation control of sewage treatment plants. Conventional BOD measurement requires a 5-day incubation period, and the operation is complicated, with poor repeatability, time-consuming and labor-intensive, and large interference, so it is not suitable for on-site monitoring. SiyaWakin et al. used a species of Trichosporoncutaneum and Bacillus licheniformis to make a microbial BOD sensor. The BOD biosensor can accurately measure the concentration of glucose and glutamate simultaneously. The measurement range is 0.5～40mg/L, and the sensitivity is 5.84nA/mgL. The biosensor is stable, with a standard deviation of only 0.0362 in 58 experiments. The required reaction time is 5～10min.
Nitrate ions are one of the major water pollutants and are extremely harmful to human health if added to food. Zatsll et al. proposed a method for the detection of nitrate ions with an integrated enzymatically functional FET device. The detection limit of the device for nitrate ions is 7×10 minus 5 mol, the response time is less than 50s, and the system operation time is about 85s.
In addition, Han et al. invented a novel microbial sensor that can be used to measure trichloroethylene. The sensor immobilized Pseudomonas JI104 on a Teflon film (diameter: 25 mm, pore size: 0.45 μm). The film was then immobilized on the chloride ion electrode. The chloride ion electrode with AgCl/Ag2S thin film (7024L, DKK, Japan) and the Ag/AgCI reference electrode were connected to an ion meter (IOL-50, DKK, Japan), and the change in voltage was recorded and compared with the standard curve. the concentration of trichloroethylene. The linear concentration range of the sensor is 0.1-4 mg/L, which is suitable for detecting industrial wastewater. Under optimal conditions, the response time is less than 10min.
(2) Atmospheric environment monitoring
Sulfur dioxide (S02) is the main reason for the formation of acid rain and acid fog, and the traditional detection method is very complicated. Martyr et al. immobilized subcellular lipids (liver microsomes containing sulfite oxidase) on a cellulose acetate membrane, and made an amperometric biosensor with an oxygen electrode to detect the acid rain and acid mist sample solution formed by S02. lOmin can get stable test results.
NOx is not only one of the causes of acid rain and acid smog, but also the main culprit of photochemical smog. Charles et al. used a microbial sensor composed of a porous permeable membrane, immobilized nitrifying bacteria and an oxygen electrode to measure the nitrite content in the sample, thereby inferring the concentration of NOx in the air. Its detection limit is 0.01xl0 minus 6 mo1/L.
Fermentation industry
Among various biosensors, microbial sensors have the characteristics of low cost, simple equipment, not limited by the degree of turbidity of fermentation broth, and may eliminate the interference of interfering substances in the fermentation process. Therefore, microbial sensors are widely adopted as an effective measurement tool in the fermentation industry.
⑴ Determination of raw materials and metabolites
Microbial sensors can be used to measure raw materials (such as molasses, acetic acid, etc.) and metabolites (such as cephalosporins, glutamic acid, formic acid, alcohols, lactic acid, etc.) in the fermentation industry. The measurement devices are basically composed of suitable microbial electrodes and oxygen electrodes. The principle is to use the assimilation of microorganisms to consume oxygen, and to measure the reduction of oxygen by measuring the change in the current of the oxygen electrode, so as to achieve the purpose of measuring the concentration of the substrate. .
In 2002, Tkac et al. used a ferricyanide-mediated glucose oxidase cell biosensor to measure the ethanol content in the fermentation industry, and the measurement could be completed within 13s with a measurement sensitivity of 3.5nA/mM. The detection limit of the microbial sensor is 0.85nM, the measurement range is 2-270nM, and the stability is very good. In the continuous 8.5h detection, there was no decrease in sensitivity.
(2) Determination of the number of microbial cells
Determination of the number of cells in the fermentation broth is important. The number of cells (cell concentration) is the number of cells in a unit of fermentation broth. Under normal circumstances, a certain fermentation broth sample needs to be taken and measured by microscopic counting method, which is time-consuming and not suitable for continuous measurement. A simple and continuous method for direct determination of cell number is urgently needed in fermentation control. It was found that on the surface of the anode (Pt), the bacteria can be directly oxidized and generate electric current. This electrochemical system can be applied to the determination of cell number. The results were similar to those determined by conventional cytometry. Using this electrochemical microbial cell count sensor can realize continuous and online determination of bacterial concentration.
medicine
Biosensors in the medical field play a growing role. Biosensing technology not only provides a fast and simple new method for basic medical research and clinical diagnosis, but also has broad application prospects in military medicine because of its specificity, sensitivity, and fast response.
⑴ clinical medicine
In clinical medicine, enzyme electrodes are the earliest developed and most widely used sensors, and have been successfully used in the detection of blood sugar, lactic acid, vitamin C, uric acid, urea, glutamic acid, transaminase and other substances. The principle is: the enzyme is mounted on the biosensitive membrane by immobilization technology, and if the detection sample contains the corresponding enzyme substrate, it can react to produce acceptable information substances, indicating that the electrode responds to changes that can be converted into electrical signals. According to this change, the presence and amount of a certain substance can be determined. Using microorganisms with different biological properties to replace enzymes, microbial sensors can be made. The microbial sensors used in clinical include sensors such as glucose, acetic acid, and cholesterol. If a suitable tissue containing a certain enzyme is selected to replace the corresponding enzyme, the sensor is called a bioelectrode sensor. For example, sensors made of pig kidney, rabbit liver, beef liver, beet, pumpkin and cucumber leaves can be used to detect glutamine, guanine, hydrogen peroxide, tyrosine, vitamin C and cystine, respectively.
DNA sensor is one of the most reported biosensors, and its use in clinical disease diagnosis is the biggest advantage of DNA sensors. It can help doctors understand the occurrence and development of diseases from the levels of DNA, RNA, proteins and their interactions, which is helpful for for timely diagnosis and treatment of diseases. In addition, drug detection is also a highlight of DNA sensors. Brabec et al. used DNA sensors to study the mechanism of action of commonly used platinum-based anticancer drugs and measured the concentration of such drugs in blood.
(2) Military medicine
In military medicine, the timely and rapid detection of biological toxins is an effective measure to defend against biological weapons. Biosensors have been applied to monitor a variety of bacteria, viruses and their toxins, such as Bacillus anthracis, Yersinia pestis, Ebola hemorrhagic fever virus, botulinum toxoid, etc.
In 2000, the U.S. military reported that it had developed an immunosensor that could detect four biological warfare agents, including staphylococcal enterotoxin B, ricin, tularemia and botulinum. The detection time is 3 ~ 10min, and the sensitivity is 10, 50mg/L, 5×10 to the 5th power, and 5×10 to the 4th power of cfu/ml. Song et al. made a biosensor to detect cholera virus. The biosensor can detect cholera toxin less than 1×10 minus 5 mol/L within 30 minutes, has high sensitivity and selectivity, and is easy to operate. This method can be used for the detection of protein toxins and pathogens with multiple signal recognition sites.
In addition, in forensic science, biosensors can be used for DNA identification and paternity verification, among others.
Operation example
There are many potential applications for various types of sensors. The demand for biosensors in the research and commercial fields is mainly driven by the identification of specific target molecules, the availability of biometric components, and single-use detection systems that are superior to laboratory techniques in some cases. Here are some examples:
Used to detect glucose concentration
Researchers at Purdue University and other institutions have created new biosensors that can non-invasively test for diabetes and detect extremely low concentrations of glucose in human saliva and tears. The technology eliminates the need for overly complex production steps, reducing the cost of manufacturing the sensor and potentially helping to eliminate or reduce the use of needle sticks for diabetes testing.
Prospects
Overview
With the development of biological science, information science and material science, biosensor technology is developing rapidly. However, the wide application of biosensors still faces some difficulties. In the future, the research work of biosensors will mainly focus on selecting biosensing elements with strong activity and high selectivity; improving the service life of signal detectors; improving signal conversion The service life of the biosensor; the stability of the biological response and the miniaturization and portability of the biosensor. It is foreseeable that future biosensors will have the following characteristics.
Diversified functions
Future biosensors will further involve various fields of healthcare, disease diagnosis, food detection, environmental monitoring, and fermentation industries. One of the important contents of biosensor research is to study biosensors that can replace biological sense organs such as vision, smell, taste, hearing and touch, which are biomimetic sensors, also known as biosensors based on biological systems.
miniaturization
With the advancement of micro-processing technology and nano-technology, biosensors will continue to be miniaturized. The emergence of various portable biosensors makes it possible for people to diagnose diseases at home and directly detect food in the market.
Intelligent integration
In the future, biosensors must be closely integrated with computers to automatically collect and process data, provide results more scientifically and accurately, and realize one-stop sampling, injection, and results, forming an automated system for detection. At the same time, chip technology will increasingly enter the sensor to realize the integration and integration of the detection system.
Low cost, high sensitivity, high stability and long life
The continuous progress of biosensor technology will inevitably require continuous reduction of product cost and improvement of sensitivity, stability and lifespan. The improvement of these characteristics will also accelerate the marketization and commercialization of biosensors. In the near future, biosensors will bring great changes to people’s lives. It has broad application prospects and will surely shine in the market.
Biosensor practicality
Biological components (enzymes, antigens, antibodies, hormones, DNA) or organisms themselves (cells, organelles, tissues) that can specifically recognize and react to various measured substances; the latter mainly include electrochemical electrodes, Ion-sensitive field effect transistors (ISFETs), thermistors, photocells, optical fibers, piezoelectric crystals (PZ), etc., whose function is to convert the biochemical signals sensed by the sensitive elements into measurable electrical signals.
Biosensors can be divided into enzyme sensors, microbial sensors, tissue sensors, organelle sensors, immunosensors, etc. according to the different molecular recognition elements used; Type biosensors, photometric biosensors, acoustic biosensors, etc.; according to different measurement methods of output electrical signals, they can be divided into potentiometric biosensors, current biosensors and voltammetric biosensors. Microbial sensors are an important branch of biosensors. Divies made the first microbial sensor in 1975, thus opening up another new field of biosensor development.
Under the condition of not damaging the function of microorganisms, microorganisms can be immobilized on the carrier to make a microbial sensor. Compared with enzyme sensors, microbial sensors have the following characteristics:
(1) The price of microbial strains is much lower than that of isolated and purified enzymes, so the sensors made are easy to popularize;
(2) The activity of the enzymes in the microbial cells is not easy to decrease under the appropriate environment, so the life of the microbial sensor is longer;
(3) Even if the catalytic activity of the enzyme in the microorganism has been lost, it can be regenerated by cell proliferation;
(4) For complex continuous reactions that require cofactors, it is easier to use microorganisms.
Derivatives
DNA biosensor
A DNA biosensor is a sensing device that converts the presence of target DNA into a detectable electrical signal. It consists of two parts, one is the recognition element, the DNA probe, and the other is the transducer. The identification element is mainly used to sense whether the sample contains the target DNA to be detected; the transducer converts the signal sensed by the identification element into a signal that can be observed and recorded. Usually, a single-stranded DNA is solidified on the transducer, and another DNA containing a complementary sequence is recognized by DNA molecular hybridization to form a stable double-stranded DNA. detection.
The principle of DNA biosensor is to hybridize a single-stranded DNA molecule with a known nucleotide sequence immobilized on the surface of the sensor or transducer probe and another complementary ss-DNA molecule, and the formed double-stranded DNA will show a certain The physical signal is finally reflected by the transducer.
Skin biosensor
Blood tests may be a common way to track certain indicators of human health, but a new project led by the U.S. military has the potential to change the way health is monitored. It turns out that many of the health indicators of human blood flow are also present in sweat.
The U.S. military’s project aims to develop skin “biosensors” that can track the flow of substances in soldiers’ sweat to monitor their health and improve their performance. The high-tech device, which looks and feels like an adhesive bandage, can be used to collect real-time measurements of heart rate, breathing rate, and more, the researchers said.
The sensor, a flat electronic chip embedded in a bandage, is designed to record health information that can be downloaded to smartphones and computers. The U.S. military hopes to use this technology to learn how to deploy troops most effectively and get them into battle at their best.
