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Rapid detection systems
- 1. Rapid detection of food borne pathogens & Biosensors. Dawrul Islam M.Tech (FSQM) 313004
- 2. Introduction • Biological defense and security applications demand rapid, sensitive detection of bacterial pathogens. • Standard culture methods for identifying pathogenic bacteria cannot provide fast results in an emergency situation such as a biological attack. • Advanced techniques are designed for detection of pathogens through gene identifications and biomarker signature.
- 3. The Issue of Food Safety • 48 million people get sick annually due to food borne diseases. • 128,000 hospitalizations • 5,000 deaths • According to the CDC- • Bacterial contamination accounts for 91% of total foodborne diseases. Annually
- 4. Needs for detection • Food borne diseases cost billions of dollars annually • The food industry is the main party concerned with the presence of pathogenic microorganisms, where failure to detect a pathogen may lead to a dreadful effect. • Pathogens may enter the food through many different ways • To minimize time and human error.
- 5. Today we will discuss…. • Conventional detection techniques. • What are biosensors , there types and applications ? • Current Research in Biosensors.
- 6. ConventionalTechniques • Culture & Colony Counting Methods. • Immunological Based Methods. • Nucleic acid-based Assays
- 7. Rapid Methods – Foodborne Pathogens • 1. Biochemical Identification pure cultures ; incubations • 2. Modification - Conventional methods (ATP & Chromogenic, fluorogenic substrates, etc.) substrates – incubations & presumptive data, cost • 3. Automated systems (eg. Biochemical , Fatty Acid identifiers) identification – Biochemical compositions -Optimized techniques
- 8. Rapid Methods – Food borne Pathogens • 4.Antibody-based Assays Highly specific interactions of antigen-antibody used for detection of pathogens. ELISA Immunoprecipitation • 5. Nucleic acid-basedAssays These include the methods based on the use of nucleic acids for detection of pathogens DNA hybridization DNA Microarray PCR: sensitive and reliable -Real time PCR
- 9. Food Analysis - timeline
- 10. Biosensors What Is a Biosensor? The bio-receptor is a biomolecule that recognizes the target analyte whereas the transducer converts the recognition event into a measurable signal.
- 11. Block Diagram of a Biosensor a) Target+Bioreceptor b) Transducer c) Amplifier d) Processor e) Monitor
- 13. Types of Biosensor in pathogen Detection • Whole-cell-based biosensors Genetically engineered yeast or bacteria cells to bear the luc or lux gene operon, expressing luminescence proteins as the green fluorescence protein (GFP). -Bacterial Bioluminescence. Detects 100 cells/hr. • Amperometric Biosensors • Amperometric biosensor are the most common electrochemical biosensor which has been used for pathogen detection. • Detection of Ecoli 0157 & Salmonella .
- 14. Types of Biosensor in pathogen Detection • Enzyme-based biosensors • An enzyme biosensor is an analytical device that combines an enzyme with a transducer to produce signal proportional to target analyte concentration. • Enzyme sensor array by model recognition using an artificial neural network . • This type of test is generally applied to analysis of dairy products, fish etc. • Optical biosensors • These types of sensors are based on measuring responses to light emission. • Optical biosensors are based on well founded methods including fluorescence, light absorbance, photothermal techniques, etc. These technical usages have been demonstrated to detect the presence of allergens, particularly peanuts, during food production.
- 15. Research in Biosensors • CARBON NANOTUBES The significant sensitivity of CNT conductivity to the surface adsorbates allows the use of CNT as highly sensitive Nano-scale sensors - Amperometric Nanobiosensors • Immunomagnetic Separation of BacterialTarget The electrically active NPs were conjugated with antibodies by direct physical adsorption of magnetic NP and confirmed in Polyclonal anti-Bacillus antibodies
- 17. Conclusions • As a rapid method is used more frequently, its benefits and at the same time, its limitations also become more apparent. • Lastly, technology continues to advance at a great pace and next generation assays, such as biosensors and DNA chips already are being developed that has the potential capability for near real-time and on-line monitoring of multiple pathogens in foods.
Hinweis der Redaktion
- Coming to the introduction section
- Mostly the FB epidemic is associated with bacteria stain eg. Shigella , Salmonella etcBioterrorism , Epidemic and Health impacts , Industrial and economical losses , detailed analysis shows biosensor are much better for accuracy , lets see what are the basic diffrences are there in conventional and advanced techniques and how development of designer biosensor with multiply testing ability can be produced.Gene id : (fim A) salmonella , (afa- ecoli 0157)
- Bioterrisom, advancement of genetically modified strains, Most of them are Salmonella sp. ,Escherichia coli, Listeria monocytogenes, Staphylococcus aureus, were found to be main source of bacterial contaminations in our food supply.
- 2. Thousands of product samples would be recalled and dumped for minimization of food borne epidemics & would lower the brand value .3. It can enter though pre harvesting , cultivation , post harvesting or at place its being served.4. In colony counting there is high risk of human error.
- How rapid detection methods are developed and , were these are applicable 3. How they are faster then conv. Methods
- Culturing the sample , then observed for pathogen though plate counting methods , and are subseptable to human error.Are based on ab-ag interactions – eg ELISA , Imunoprecipitation. Are Hybridization techniques , faster and reliable then above
- BROAD CLASSIFICATION OF IDENTIFICATIONBiochemical identifiers like specific enzyme , Fatty acid compostionIn a study that was done in year 2007 ,they selected a specific monoclonal antibody (mouse IgG1) against L. monocytogenes, and also attempted to detect it directly.
- DNA hy- similar to the sandwich ‘elisa’ but instead enzyme conjugates with the ab which forms color.Probes are labelled with radioactive labels for detection.Autoradiography is then used to detect the probe-target complexesDNA microarray- advanced technique which are extensively used for analysis of genetic constraint , also analyses level of expression of toxic gene or protiens
- Complex carbohydrates , hormones etc
- These are the following types of recognition elements which binds to biosensor and produces quantifiable results.Calorimetric biosensors – food quality analysis , Heat based detection .
- Those sensors based on eitherthe fusion of the lux operon to result in detectable amount of bioluminanceBact.Biolum.Capable of detecting 100 cells/hrAmperometric detection of micro-organisms involvesthe measurement of the current generated through electrooxidation/reduction catalysed by their enzymesAmp-E. coli O157:H7 |81cfu| ( 6 min ) Salmonella 1–5 6hL. monocytogenes, and C. jejuni 10 cells/mL.Another study on detection of the freshness; based on L-lactate detection in tomato paste and infant food or by using a continuous measurement format of enzyme flow reactors [Baeumner et al. 2003]Phytotoxins,especially those produced by algae and found in seafoodwere of interest in Baeumner study 2003
- Enzyme based--simultaneous determination of the three biogenic amines (putrescine, histamine and Tyramine).A monoamine oxidase, a diamine oxidase (with specific activities adequate for rapid detection)Mycotoxins, such as aflatoxin B1, have been currently used as targets for analysis performed with screen-printed immunosensor devices.Biosensor kit: Cobas IDA BC : EnterobacteriaMicroID : BC: Listeria Riboprinter – Nucliec acid – Salmonella , stplycocus, EcoliBactometer- salmonella
- CNT-oxi-The accelerated electron transfer reaction of hydrogen peroxide at the CNT-based paste electrode offered a rapid low-potential (0.10 V) detection of the substrate
- The electrically active NPs were conjugated with antibodies by direct physical adsorption as described and confirmed in Polyclonal anti-Bacillus antibodies The bio-modified particles (immuno-c/sNPs) were magnetically separated to remove any unbound antibody in the supernatant, washed twice with blocking buffer, re-suspended in phosphate buffer.GE healthcare Genequant 100
- Because of the complex designs and formats of these tests, coupled with the difficulties of testing foods, users must exercise caution when selecting rapid methods and to also evaluate these tests thoroughly, as some may be more suitable than others for distinct testing situations or for assaying certain types of food