reportBiological Detection
9.3.3 Short description

Detection of biological threats, which could be used against citizens and agricultural produce, involves the recognition of bacteria, viruses and toxins. The identification methods based on nucleic acid and immuno-based methods are used in detecting has been mentioned in the literature. These have also been used in food testing, clinical and environmental applications [50].

Table BW.2 - A selection of detection of commonly known bacterial pathogens as cited in the journal paper [50]

	Biorecognition molecules	Target	Type of assay	Estimated time of assay (h)	Sensitivity (no. of targets)	Specificity
Nucleic acid	pst gene probes and primers	Yersinia pestis	5% nuclease fluorogenic assay	2.5 h	3-30 cells	Very high
	Heat stable toxin gene (ST-gene)	E. coli	PCR-ELISA; color amplified PCR (CAPS)	3.5 h	270 cfu	High
	Genomic DNA; Escherichia coli	E. coli and lambda phage ,	Lambda phage Molecular combing, scanning force microscopy	2 h	103 cells	High
	Universal and specific 16s rRNA primers	B. subtilis, Y. pestis and E. coli	Multiplex PCR followed by gel electrophoresis	3 h	ND (5 ml of soil)	High

Immuno-assays	Fluorescently labeled MAb	Salmonella typhimurium	Flow cytometry	40 min	103 cells:ml of milk & egg	High
	Fluorescently labeled PAb to 0157 antigen	E. coli 0157:H7	Antibody-direct Epifluorescent filter technique (Ab-DEFT)	3 h	104-105 cfu:g feces	High

Table BW.3 - A selection of recognition techniques for detection of pathogenic viruses as cited in a journal paper [50]

	Biorecognition molecules	Target	Type of assay	Estimated time of assay (h)	Sensitivity (no. of targets)	Specificity
Nucleic acid	Biotinylated primers for gag gene and internal standard control	HIV-1 RNA:DNA	QC-PCR followed by amplicon capture, probe hybridization and Luminometry	4h	100	Very high quantitative
	Biotinylated primers for pol gene	HIV-1 RNA	PCR followed by amplicon capture and colorimetry	3 h	20 virions	Very high
	Primers for immediate early genes	Active human cytomegalovirus (hCMV)	In situ PCR followed by in situ colorimetry	4 h	100	High
	Primers for immediate early genes	hCMV	Nested PCR followed by gel electrophoresis	4 h	5 virions	High
	Primers and biotinylated probes	Hepatitis C virus (HCV)	PCR based, digoxigenin labled amplicons are captured by biotinylated probes	3 h	10-100 virions	Very high
	Digioxigenin labeled riboprobe targeting non-coding sequence	Enteroviruses	Dot blot hybridization and chemiluminescent detection	2.5 h	10⁴-10⁵ TCID	High
	Digioxigenin labeled Vp1 probe	Polioviruses serotyopes	Dot blot hybridization and chemiluminescent detection	2.5 h	10³-10⁴ TCID	High

Immuno-assays	Fluorescein labeled human recombinant Fab (rFab)	Cells infected with herpes simplex virus (HSV)	Indirect immunofluorescence	2.5 h	ND	High
	Fluorescein labeled murine monoclonal Ab to HSV	Cells infected with HSV	Direct immunofluorescence	2 h indirect	ND (less than	High
	Latex bound antibody to	Cytomegalovirus (CMV)	Latex agglutination, visual assay	10 min	10¹²	High
	IgM antibody to	CMV	Enzyme immuno assay	10 min	10⁶	High
	Antibody	HAV	Immuno electron microscopy	8 h	10⁴-10⁵ virions/ml	Moderate
	Radio labeled antibody	HAV	Radioimmunoassay	2 h	10⁵-10⁶ virions/ml	Moderate

Self assembled bilayers of Cu2+/L-cysteine have been coated on gold surfaces are used to detect biological agents in microcantilever sensors. Dimethyl methyl phosphonate was used as a sarin nerve gas stimulant in these tests [51].

Label free biosensors also known as optical biosensors are based on direct measurements of a change taking place during a biochemical reaction on the surface of a transducer [52]. Ricin has been detected using optical biosensors within 30 minutes to a detection limit of 10ng/ml. The advantages optical biosensors offer is rapid screening and multi-analyte detection. The main disadvantage for these sensors is the reduced sensitivity for the rapid screening assays [53].

Portable fibre optic biosensors have been demonstrated in the detection of Staphylococcal Enterotoxin B. These biosensors have been demonstrated in the compact, light weight portable
identification system. The multi-channel identification system is based on simultaneous
fluorescence immunoassays on the surface of polystyrene fibre optic probes [54].

NASA has demonstrated the nanotechnology based biosensors that can be used to detect pathogens such as anthrax. These sensors are based on carbon nanofibres and the licensed technology is being commercialised by Early Warning Inc. Electrical signals are measured in
these sensors to identify the presence of a pathogen. These sensors are equipped with
microfludics as well that provides the advantage of field testing, allowing detection within 30 minutes [55].

Aptamers are functional RNA oligonucleotides that are used for sensing biologicial agents. These are also DNA based. Aptamers based biosensors have been reported to be good for security applications. Aptamers based biosensors are relatively immature, in relation to immunoassays, due to the limited availability of aptamers and knowledge of surface immobilization [56]. Single
walled carbon nanotubes based field effect transistors, has been demonstrated to monitor
aptamers - protein affinity binding processes. These offer an advantage over immunological
assays due to their small size in monitoring protein- protein interactions [57].

Array Biosensors have been used to detect targets such as staphylococcal enterotoxin B (SEB), ricin toxin, cholera toxin, mouse IgG and Bacillus globigii (anthrax spore simulant). Prototype for monitoring postal sorting machines was demonstrated to be successful. The array biosensors provide the advantage of being specific and non-destructive [58].

Inhibition of enzyme acetylcholinesterase has been the basis of detecting organophosphate
compounds due to high specificity and selectivity. The development of single thiocholine enzyme based biosensor has been reported in the literature using screen printed carbon electrodes doped with cobalt phthalocyanine. The sensors were observed to be fabricated using electropolymerisation and ablation with ultrasound. Detection limits in the order of 1 * 10-17M were experimentally reported for dichlorvos, parathion and azinphos. In a separate experiment the
same detection limit was reported for paraoxon. The biosensors were reported to have application in environmental monitoring [59,60].

Biosensors based on acetylcholinesterase functionalised carbon nanotubes have been demonstrated in detecting organophosphorus compounds. These sensors have shown excellent limits of detection (0.145 ppb), good precision, electrode to electrode stability, and reproducibility
[61].

Metallic nanowires striped with gold, silver and nickel nanoparticles in a suspended format have been used to identify biological warfare agents such as anthrax, smallpox, ricin and botulinum. Each nanowire relating to a particular antibody detects a pathogen. The made advantage offered by this method of detection is the ability to have up to 100 different striped nanowires which
reduce analysis time significant for multiple analytes [62].

Kane et al. have demonstrated the use of peptide bound carbon nanotubes in detecting toxins such as anthrax, and deactivating the anthrax toxin by using invisible and near infrared light. Carbon nanotubes coatings maybe applied as a thin coating on a range on surfaces [63]. SERS enhancements have been used to detect microorganisms using colloidal metal suspension [64].

Terahertz imaging has been demonstrated for anthrax stimulant bacillus cereus in postal
envelopes [65]. The need for research in detection methods of biological threats was driven by the anthrax spores distribution through the postal services in the United States. Laser induced breakdown spectroscopy (LIBS) is a technique that uses light induced from a laser induced microplasma to determine the composition of the sample, based on elemental and molecular
emission intensity. The main advantages of this technique are high sensitivity, selectivity and
minimal sample preparation. LIBS have been demonstrated to have been effectively used in the detection of bacillus subtilis and ovalbumin. Experimental research has established the
effectiveness of this method with few false positive or false negatives [66].

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Tags: biosensors, terahertz, Carbon Nanotubes, nanowires, optical biosensors, nucleic acid based sensing, immunoassay based sensing

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reportBiological Detection 9.3.3 Short description

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reportBiological Detection
9.3.3 Short description