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WO2005019413A2 - A low maintenance detector for biological agents - Google Patents

A low maintenance detector for biological agents

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Publication number
WO2005019413A2
WO2005019413A2 PCT/US2004/018374 US2004018374W WO2005019413A2 WO 2005019413 A2 WO2005019413 A2 WO 2005019413A2 US 2004018374 W US2004018374 W US 2004018374W WO 2005019413 A2 WO2005019413 A2 WO 2005019413A2
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WO
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Patent type
Prior art keywords
detection
sample
target
probes
molecule
Prior art date
Application number
PCT/US2004/018374
Other languages
French (fr)
Other versions
WO2005019413A3 (en )
Inventor
Dennis M. Connolly
David R. Chafin
Original Assignee
Integrated Nano-Technologies, Llc
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    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12QMEASURING OR TESTING PROCESSES INVOLVING ENZYMES OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/68Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
    • C12Q1/6813Hybridisation assays
    • C12Q1/6816Hybridisation assays characterised by the means of detection
    • C12Q1/6825Nucleic acid detection involving sensors

Abstract

The present invention relates to a device for the detection of a target molecule in an environment. The device includes a detection system having a detection cartridge (303) containing a detection chip where capture probes are attached to two or more electrically separated conductors such that a gap exists between the capture probes on the electrically separated conductors. A sample can be analyzed for the presence of the target molecule by determining whether any target molecules present in the sample bind to the capture probes and electrically connect the conductors. The device includes a cartridge manipulator (302) having a plurality of detection cartridges (303) where an unused individual detection cartridge can be inserted into a support unit (300) for detection of the target molecule in the sample. The present invention also relates to a method for the detection of a biological agent using the device disclosed.

Description

A LOW MAINTENANCE DETECTOR FOR BIOLOGICAL AGENTS

[0001] This application claims the benefit of U.S. Provisional Patent

Application Serial No. 60/478,134, filed June 12, 2003, which is hereby incorporated by reference in its entirety.

FIELD OF THE INVENTION

[0002] The present invention relates to a device and a method for the detection and identification of a biological agent with a sensitive, highly accurate detection system that has the capability to run and store several tests per day without human intervention.

BACKGROUND OF THE INVENTION

[0003] Nucleic acids, such as DNA or RNA, have become of increasing interest as analytes for identifying unknown biological agents in an experimental or field sample. Powerful new biotechnologies enable one to detect infectious organisms in a clinical setting. These same technologies can be used to detect bio warfare agents in a variety of situations.

[0004] Some devices have been developed that monitor the environment for the presence of particles, the so-called particle counters. These devices typically use a laser to scan the air in front of the device and record the number and size of particles present. Most particle counters can be programmed to monitor for a specific sized particle, such as that relating to bacteria or bacterial spores. However, even the most sophisticated particle counters have a high rate of positive signals resulting from normal contamination in the environment. Since these devices merely "look" for particles, they cannot make an absolute identification of the type of particle present. Therefore, these devices need the assistance of a dedicated detector for biological materials to monitor the environment and examine the particles to make an exact genetic identification of the contaminant.

[0005] For the analysis and testing of nucleic acid molecules, amplification of a small amount of nucleic acid molecules, isolation of the amplified nucleic acid fragments, and other procedures are typically necessary. The science of amplifying small amounts of DNA has progressed rapidly and several methods now exist. These include linked linear amplification, ligation- based amplification, transcription-based amplification, and linear isothermal amplification. Linked linear amplification is described in detail in U.S. Patent No. 6,027,923 to Wallace et al. Ligation-based amplification includes the ligation amplification reaction (LAR) described in detail in Wu et al. Genomics 4:560 (1989) and the ligase chain reaction described in EP application No. 0320308B1. Transcription-based amplification methods are described in detail in U.S. Patent No. 5,766,849 to McDonough et al., U.S. Patent No. 5,654,142 to Kievits et al., Kwoh et al., "Transcription-Based Amplification System and Detection of Amplified Human Immunodeficiency Virus Type 1 with a Bead-Based Sandwich Hybridization Format," Proc. Natl. Acad. Sci. U.S.A. 86:1173(1989), and PCT Patent Application No. WO 88/10315 to Ginergeras et al. The more recent method of linear isothermal amplification is described in U.S. Patent No. 6,251,639 to Rum.

[0006] The most common method of amplifying DNA is by PCR, described in detail by Mullis et al., "Specific Enzymatic Amplification of DNA in vitro: The Polymerase Chain Reaction," Cold Spring Harbor Quant. Biol. 51:263-273 (1986); EP 201,184 to Mullis; U.S. Patent Nos. 4,683,195 and

4,683,202 to Mullis et al.; EP 50,424, EP 84,796, EP 258017, EP 237362 to Erlich et al.; and U.S. Patent No. 4,683,194 to Saiki et al. The PCR reaction is based on multiple cycles of hybridization, nucleic acid synthesis, and denaturation in which an extremely small number of nucleic acid molecules or fragments can be multiplied by several orders of magnitude to provide detectable amounts of material. One of ordinary skill in the art knows that the effectiveness and reproducibility of PCR amplification is dependent, in part, on the purity and amount of the DNA template. Certain molecules present in biological sources of nucleic acids are known to stop or inhibit PCR amplification (Belec et al., "Myoglobin as a Polymerase Chain Reaction (PCR) Inhibitor: a Limitation for PCR from Skeletal Muscle Tissue Avoided by the Use of Thermus Thermophims Polymerase," Muscle and Nerve 21 (8): 1064 (1998); Wiedbrauk et al., "Inhibition of PCR by Aqueous and Vitreous Fluids," J. Clinical Microbiology 33(10):2643-6 (1995); Deneer et al., "Inhibition of the Polymerase Chain Reaction by Mucolytic Agents," Clinical Chemistry. (40(l):171-2 (1994)). In whole blood, for example, hemoglobin, lactoferrin, and immunoglobulin G are known to interfere with several DNA polymerases used to perform PCR (Al-Soud et al., J. Clinical Microbiology 39(2):485-493(2001); Al-Soud et al., "Purification and

Characterization of PCR-Inhibitory Components in Blood Cells," J. Clinical Microbiology 38(l):345-50 (2000)). These inhibitory effects can somewhat be overcome by the addition of certain protein agents, but these must be added in addition to the multiple components already used to perform the PCR. Thus, the removal or inactivation of such inhibitors is an important factor in amplifying DNA from select samples.

[0007] On the other hand, isolation and detection of particular nucleic acid molecules in a mixture requires a nucleic acid sequencer and fragment analyzer, in which gel electrophoresis and fluorescence detection are combined. Unfortunately, electrophoresis becomes very labor-intensive as the number of samples or test items increases.

[0008] For this reason, a simpler method of analysis using DNA oligonucleotide probes is becoming popular. New technology, called VLSIPS™, has enabled the production of chips smaller than a thumbnail where each chip contains hundreds of thousands or more different molecular probes. These techniques are described in U.S. Patent No. 5,143,854 to Pirrung et al., PCT Publication No. WO 92/10092, and PCT WO 90/15070. These biological chips have molecular probes arranged in arrays where each probe ensemble is assigned a specific location. These molecular array chips have been produced in which each probe location has a center-to-center distance measured on the micron scale. Use of these array type chips has the advantage that only a small amount of sample is required, and a diverse number of probe sequences can be used simultaneously. Array chips have been useful in a number of different types of scientific applications, including measuring gene expression levels, SNP (small nucleic acid polymorphism) identification, molecular diagnostics, and sequencing, as described in U.S. Patent No. 5,143,854 to Pirrung et al. [0009] Array chips where the probes are nucleic acid molecules have been increasingly useful for detecting the presence of specific DNA sequences. Most technologies related to array chips involve the coupling of a probe of known sequence to a substrate that can either be structural in nature or conductive. Structural types of array chips usually involve providing a platform where probe molecules can be constructed base by base or the completed molecule bound covalently. Typical array chips involve amplification of the target nucleic acid followed by detection with a fluorescent label to determine whether target nucleic acid molecules hybridize with any of the oligonucleotide probes on the chip. After exposing the array to a sample containing target nucleic acid molecules under selected test conditions, scanning devices can examine each location in the array and quantitate the amount of hybridized material at that location.

Alternatively, conductive types of array chips contain probe sequences linked to conductive materials such as metals. Hybridization of a target nucleic acid typically elicits an electrical signal that is carried to the conductive electrode and the signal analyzed. [0010] However, in order to detect target DNA molecules, the target must first be amplified by PCR to get a reliable signal. The ability to detect fluorescent or radioactive materials is generally needed. Such a system is expensive to use and is not amenable to a portable system for biological sample detection and identification. Furthermore, the hybridization reactions take up to two hours. For many potential uses, such as detecting biological warfare agents, this is simply not effective. Therefore, there is a need for a device and method which can rapidly detect small quantities of a target nucleic acid molecule without relying on PCR amplification. It would also be useful if such a device would be capable of running and storing multiple tests in a day, with limited human intervention. [0011] The present invention is directed to overcoming these and other deficiencies in the art.

SUMMARY OF THE INVENTION

[0012] The present invention relates to a method for detecting a target molecule. This method involves providing a detection system having a plurality of detection cartridges. Each detection cartridge includes a housing defining a first chamber and a detection chip within the first chamber defined by the housing. The detection chip includes two or more electrically separated conductors fabricated on a substrate and capture probes attached to the conductors such that a gap exists between the capture probes on the electrically separated conductors. A sample, potentially containing a target molecule, can be analyzed for the presence of the target molecule by determining whether the conductors are electrically connected. An electrical connector extends through the housing and is coupled to the electrically separated conductors so that the presence of a target molecule connecting the capture probes on the electrically separated conductors can be detected. The detection system includes a support unit into which the detection cartridge can be positioned to carry out a procedure for detecting the target molecule in the sample. The support unit has an electrical coupler suitable for electrical communication with the electrical connector of the detection cartridge, whereby the presence of the target molecule in the sample can be detected and communicated to the support unit. Also included in the detection system is a detection cartridge manipulator suitable for holding the plurality of detection cartridges and positioned to insert unused detection cartridges individually into the support unit for detection of a target molecule in a sample and for removing detection cartridges individually from the support unit after use. The method also involves inserting an unused detection cartridge into the support unit and processing a sample within the first chamber of the detection cartridge under conditions effective to permit any target molecules present in the sample to bind to the capture probes and thereby connect the capture probes. It is then determined whether the electrical conductors are electrically connected, thereby detecting the presence of the target molecule in the sample. [0013] The present invention also relates to a device for detecting a target molecule. This device includes a detection system having a plurality of detection cartridges. Each detection cartridge includes a housing defining a first chamber and a detection chip within the first chamber defined by the housing. The detection chip includes two or more electrically separated conductors fabricated on a substrate and capture probes attached to the conductors such that a gap exists between the capture probes on the electrically separated conductors. A sample, potentially containing a target molecule, can be analyzed for the presence of the target molecule by determining whether the conductors are electrically connected. An electrical connector extends through the housing and is coupled to the electrically separated conductors so that the presence of a target molecule connecting the capture probes on the electrically separated conductors can be detected. The detection system includes a support unit into which the detection cartridge can be positioned to carry out a procedure for detecting the target molecule in the sample. The support unit has an electrical coupler suitable for electrical communication with the electrical connector of the detection cartridge, whereby the presence of the target molecule in the sample can be detected and communicated to the support unit. Also included in the detection system is a detection cartridge manipulator suitable for holding the plurality of detection cartridges and positioned to insert unused detection cartridges individually into the support unit for detection of a target molecule in a sample and for removing detection cartridges individually from the support unit after use. [0014] The device and method of the present invention provide an efficient, low maintenance way to monitor and accurately identify biological agents in an environment. In particular, the present invention is useful for testing multiple biological samples in a day, storing and transmitting the results to a remote location, and operating for extended periods of time without human intervention. Thus, it is highly effective for many uses, including the detection of biowarfare agents.

BRIEF DESCRIPTION OF THE DRAWINGS

[0015] Figure 1 is a diagram of the bio-detection sensor device of the present invention coupled to a continuous particle counter. [0016] Figure 2 is a diagram of a bio-detection device in which the air is continuously monitored by a particle counter.

[0017] Figure 3 is a diagram of the detection device of the present invention having a detection cartridge manipulator that is a carousel.

[0018] Figure 4 is a diagram of the detection device of the present invention having a detection cartridge manipulator that is magazine.

[0019] Figure 5 is a diagram of detection device of the present invention having a detection cartridge manipulator that is a tape reel structure. [0020] Figures 6A-B are diagrams of the structure and use of a detection system in accordance with the present invention. Figure 6A depicts a single test structure on a detection chip suitable to be positioned in the detection cartridge in Figures 3-5, where oligonucleotide probes are attached to electrical conductors in the form of spaced apart conductive fingers. Figure 6B shows how a target nucleic acid molecule present in a sample is detected by the detection chip.

DETAILED DESCRIPTION OF THE INVENTION

[0021] The present invention relates to a method for detecting a target molecule. This method involves providing a detection system having a plurality of detection cartridges. Each detection cartridge includes a housing defining a first chamber and a detection chip within the first chamber defined by the housing. The detection chip includes two or more electrically separated conductors fabricated on a substrate and capture probes attached to the conductors such that a gap exists between the capture probes on the electrically separated conductors. A sample, potentially containing a target molecule, can be analyzed for the presence of the target molecule by determining whether the conductors are electrically connected. An electrical connector extends through the housing and is coupled to the electrically separated conductors so that the presence of a target molecule connecting the capture probes on the electrically separated conductors can be detected. The detection system includes a support unit into which the detection cartridge can be positioned to carry out a procedure for detecting the target molecule in the sample. The support unit has an electrical coupler suitable for electrical communication with the electrical connector of the detection cartridge, whereby the presence of the target molecule in the sample can be detected and communicated to the support unit. Also included in the detection system is a detection cartridge manipulator suitable for holding the plurality of detection cartridges and positioned to insert unused detection cartridges individually into the support unit for detection of a target molecule in a sample and for removing detection cartridges individually from the support unit after use. The method also involves inserting an unused detection cartridge into the support unit and processing a sample within the first chamber of the detection cartridge under conditions effective to permit any target molecules present in the sample to bind to the capture probes and thereby connect the capture probes. It is then determined whether the electrical conductors are electrically connected, thereby detecting the presence of the target molecule in the sample. [0022] The present invention also relates to a device for detecting a target molecule. This device includes a detection system having a plurality of detection cartridges. Each detection cartridge includes a housing defining a first chamber and a detection chip within the first chamber defined by the housing. The detection chip includes two or more electrically separated conductors fabricated on a substrate and capture probes attached to the conductors such that a gap exists between the capture probes on the electrically separated conductors. A sample, potentially containing a target molecule, can be analyzed for the presence of the target molecule by determining whether the conductors are electrically connected. An electrical connector extends through the housing and is coupled to the electrically separated conductors so that the presence of a target molecule connecting the capture probes on the electrically separated conductors can be detected. The detection system includes a support unit into which the detection cartridge can be positioned to carry out a procedure for detecting the target molecule in the sample. The support unit has an electrical coupler suitable for electrical communication with the electrical connector of the detection cartridge, whereby the presence of the target molecule in the sample can be detected and communicated to the support unit. Also included in the detection system is a detection cartridge manipulator suitable for holding the plurality of detection cartridges and positioned to insert unused detection cartridges individually into the support unit for detection of a target molecule in a sample and for removing detection cartridges individually from the support unit after use. [0023] In one aspect, the present invention relates to a device and a method for coupling the detection system with a continuous particle-monitoring device. The device would provide a particle-monitoring device that is physically or electrically coupled to the detection system containing a plurality of detection cartridges. Detection cartridges would be provided in a magazine, carousel, or tape reel set-up, protected from the environment. Also provided is an air-filtering device. Such devices are commercially available from manufacturers such as Meso Systems Technology, Inc.

[0024] In order to conserve the detection cartridges, the particle counter would initially scan an environmental region based on parameters input from an operator. If enough particles are present in the environmental region, or, alternatively, particles of a given size are detected, a signal would be sent to run a test for identification of the target molecule of interest. Once a signal is received that activates the detection system, a detection test cartridge is displaced from a magazine, carousel or sensor tape spot and loaded into the support unit. At the same time, a separate signal is sent to an air-sampling device and a known quantity of air is filtered. The air goes through a specific filter that will trap bacteria and bacterial spores. Next, the filter is washed with a small amount of fluid to displace the bacteria. [0025] The sample is injected into the detection test cartridge where the bacteria are lysed and the genetic material released. The genetic material is filtered to remove any cell debris, sheared to a specific size, and pushed onto the chip sensor. The chip sensor can contain multiple test sites and contain capture probes specific for a set of bacteria. After a short incubation time under conditions effective to permit any target nucleic acid molecule present in the sample to hybridize to both of the spaced apart probes to bridge the gap and electrically couple the pair of probes with the hybridized target nucleic acid molecule, the sample is pushed into a waste collection container on the detection cartridge. [0026] Next, the DNA bridges formed are made conductive such that an electrical signal can be generated from individual test sites. After hybridization of the target nucleic acid molecule to the probes, the hybridized target nucleic acid molecule is coated with a conductive material. Examples of conductive materials include, without limitation, silver and gold. Determination of electrical current between the probes indicates the presence of the target nucleic acid molecule in the sample which has sequences complementary to the probes.

[0027] Signals from the particle counter and test results from the detection system can be sent to a central monitoring station or to other remote locations. [0028] The basic configurations of the overall system are shown in Figure

1 and Figure 2.

[0029] According to Figure 1, air is continuously monitored by a particle counter 100. The air enters the air intake 101, passes through the air bridge 102, passes through the bio-aerosol collector/concentrator 103, and exits the air exhaust 104. If the particle counter 100 registers a positive event, it signals the bio- detection sensor 106 and the bio-aerosol collector/concentrator 103, via communication lines 107, to start the bio-detection process. The bio-aerosol collector/concentrator 103 concentrates the air sample into a liquid form. While this is happening, the bio-detection sensor 106 initiates the bio-detection process. The liquefied air sample is transferred to the bio-detection sensor 106 via the sample bridge 105. The bio-detection sensor 106 performs the detection process and transmits the results to a monitoring system. [0030] According to Figure 2, air is continuously monitored by a particle counter 200. The air 201 is scanned by the particle counter 200. If the particle counter 200 registers a positive event, it signals the bio-detection sensor 206 and the bio-aerosol collector/concentrator 203, via communication lines 207, to start the bio-detection process. The bio-aerosol collector/concentrator 203 concentrates the air sample into a liquid form. While this is happening, the bio-detection sensor 206 initiates the bio-detection process. The liquefied air sample is transferred to the bio-detection sensor 206 via the sample bridge 205. The bio- detection sensor 206 performs the detection process and transmits the results to a central monitoring station or to other remote locations [0031] The bio-detection sensor 100, 200, 300, 400, and 500 takes the liquefied air sample and performs a sample prep step. The bacteria are lysed and the genetic material released. The genetic material is filtered to remove any cell debris, and sheared to a specific size. The sheared DNA sample is then transferred to the interface manifold 307, 407, and 507 via the sample injection line 306, 406, and 506 using the pump module 310, 411, and 510. The sheared DNA sample is flowed over the microchip, which is embedded in the detection cartridges 303, 403, and 503. The chip sensor preferably contains multiple test sites and multiple capture probes specific for a set of bacteria. After a short incubation time, the sample is pushed into a waste collection container on the detection cartridge 303, 403, and 503. Next, the DNA bridges formed are made conductive such that an electrical signal can be generated from individual test sites. If any bacteria are present in the sample, test sites containing those probes sequences will result in positive electrical tests, thereby identifying the organisms present in the original environmental sample. Signals from the particle counter 100 and 200 and test results from the bio-detection sensor 300, 400, and 500 can be sent to a central monitoring station or to other remote locations. [0032] In one aspect of this invention, as shown in Figure 3, detection cartridges 303 are housed in a carousel 302. Similar to slide projector methods, the detection test cartridges 303 would reside in numbered slots. The carousel is rotated on a platform, moving the test cartridges successively to a spot just above the support unit of the detection sensor. When rotated into place, a detection cartridge 303 would be lowered into the support unit of the bio-detection sensor 300 and the interface manifold 307 would be attached. The liquefied air sample would enter the bio-detection sensor 300 via the liquid sample intake 305. The sample would be processed in the sample prep module 304. The liquefied air sample would then be injected into the detection cartridges 303 via the sample bridge 306, interface manifold 307, and the pump module 310. After testing is complete, the detection cartridge 303 would be disconnected from the interface manifold 307 and returned to the carousel 302 in the same position. The test results would be delivered and the carousel 302 rotated one position to receive the next detection cartridge 303. The number of tests run would be logged and a maintenance technician notified when all of the detection cartridges 303 are used. The carousel 302 would then be removed and replaced with a new carousel 302 containing fresh detection test cartridges 303. Carousels 302 could be designed to hold a variable number of test cartridges depending on the end user and the application. Analysis, control, and inter-device communications functions would be performed by the electrical module 301 via the communication lines 309. Communications to the central monitoring station or to other remote locations would be performed by the communications module 308.

[0033] In another aspect of this invention, as shown in Figure 4, detection cartridges 403 are housed in a magazine 402. A detection cartridge 403 would be lowered into the support unit of the bio-detection sensor and the interface manifold 407 would be attached. The liquefied air sample would enter the detection sensor 400 via the liquid sample intake 404. The sample would be processed in the sample prep module 405. The liquefied air sample would then be injected into the detection cartridges 403 via the sample bridge 406, interface manifold 407, and the pump module 411. After testing is complete, the detection cartridge 403 would be disconnected from the interface manifold 407 and returned to the magazine 402 in the same position. The test results would be delivered and the interface manifold 407 would be moved along the interface manifold rail 410 to the next position in order to receive the next fresh detection cartridge 403. The number of tests run would be logged and a maintenance technician notified when all of the detection cartridges 403 are used. The magazine 402 would then be removed and replaced with a new magazine 402 containing fresh detection cartridges 403. Magazines 402 could be designed to hold a variable number of test cartridges depending on the end user and the application. Analysis, control, and inter-device communications functions would be performed by the electrical module 401 via the communication lines 409. Communications to the central monitoring station or to other remote locations would be performed by the communications module 408. [0034] In yet another aspect of the present invention, as shown in Figure 5, detection cartridges 503 are housed on a tape reel 502. A detection cartridge 503 would be moved to a designated position and the interface manifold 507 would be attached. The liquefied air sample would enter the bio-detection sensor 500 via the liquid sample intake 504. The sample would be processed in the sample prep module 505. The liquefied air sample would then be injected into the detection cartridges 503 via the sample bridge 506, interface manifold 507, and the pump module 510. After testing is complete, the interface manifold 507 would be removed from the detection cartridge 503. The test results would be delivered and the tape reel 502 would be advanced to the next position in order to receive the next fresh detection cartridge 503. The number of tests run would be logged and a maintenance technician notified when all of the detection cartridges 503 are used. The tape reel 502 would then be removed and replaced with a new tape reel 502 containing fresh detection cartridges 503. Tape reels 502 could be designed to hold a variable number of test cartridges depending on the end user and the application. Analysis, control, and inter-device communications functions would be performed by the electrical module 501 via the communication lines 509. Communications to the central monitoring station or to other remote locations would be performed by the communications module 508. [0035] Figure 6A depicts a single test structure on a detection chip suitable to be positioned in detection cartridge 303, 403, and 503 of the system shown in Figures 3-5. According to Figure 6 A, oligonucleotide probes 600 attached to spaced apart conductive fingers 601 are physically located at a distance sufficient that they cannot come into contact with one another. A sample, containing a mixture of nucleic acid molecules (e.g., M1-M6) to be tested is contacted with the fabricated device on which conductive fingers 601 are fixed, as shown in Figure 6B. If a target nucleic acid molecule (e.g., Ml) that is capable of binding to the two oligonucleotide probes is present in the sample, the target nucleic acid molecule will bind to the two probe molecules. If bound, the nucleic acid molecule can bridge the gap between the two electrodes and provide an electrical connection. Any unhybridized nucleic acid molecules (e.g., M2-M6) not captured by the probes are washed away. Here, the electrical conductivity of nucleic acid molecules is relied upon to transmit the electrical signal. Hans-Werner Fink and Christian Schoenenberger reported in Nature, 398:407-410 (1999), which is hereby incorporated by reference in its entirety, that DNA conducts electricity like a semiconductor. This flow of current can be sufficient to construct a simple switch, which will indicate whether or not a target nucleic acid molecule is present within a sample. The presence of a target molecule can be detected as an "on" switch, while a set of probes not connected by a target molecule would be an "off switch. The information can be processed by a digital computer which correlates the status of the switch with the presence of a particular target. The information can be quickly identified to the user as indicating the presence of the target molecule of interest. Optionally, after hybridization of the target molecules to sets of biological probes, the target molecule can be coated with a conductor, such as a metal. The coated target molecule can then conduct electricity across the gap between the pair of probes, thus producing a detectable signal indicative of the presence of a target molecule. [0036] The detection cartridges 303 and 403 would be constructed in one of two potential fashions. The first would be to laminate the microchip within a plastic package containing microfluidic features and flexible circuitry. The microfluidic features would include reservoirs, mixing chambers, channels, valves, and waste chambers. The flexible circuitry would create the heat zones and attach the microchip to the interface manifold 307 and 407. The second would be to embed the microchip within a micro-injection molded plastic package containing microfluidic features and flexible circuitry. The microfluidic features would include reservoirs, mixing chambers, channels, valves, and waste chambers. The flexible circuitry would create the heat zones and attach the microchip to the interface manifold 307 and 407.

[0037] The detection cartridges 503 would be constructed on a continuous strip of flexible material. The circuitry that is found on the microchip would be applied to the flexible material by lithographic techniques using a conductive liquid. If the conductive liquid is not conducive to probe binding, electroplating would be used to apply a surface conducive to probe binding. The rest of the package would be created using lamination techniques. The laminated package would include microfluidic features such as reservoirs, mixing chambers, channels, valves, and waste chambers. The interface manifold 507 would connect directly to the lithographically applied circuitry.

[0038] Various methods exist for attaching the capture probes to the electrical conductors. For example, U.S. Patent No. 5,861,242 to Chee et al.; U.S. Patent No. 5,856,174 to Lipshutz et al.; U.S. Patent No. 5,856,101 to Hubbell et al.; and U.S. Patent No. 5,837,832 to Chee et al., which are hereby incorporated by reference in their entirety, disclose a method where light is shone through a mask to activate functional (for oligonucleotides, typically an -OH) groups protected with a photo-removable protecting group on a surface of a solid support. After light activation, a nucleoside building block, itself protected with a photo- removable protecting group (at the 5'-OH), is coupled to the activated areas of the support. The process can be repeated, using different masks or mask orientations and building blocks, to place probes on a substrate. Details on methods of attaching biological molecules to electrically conductive surfaces can also be found in U.S. Patent Application Serial No. 10/159,429, filed on May 30, 2002, which is hereby incorporated by reference in its entirety. [0039] Alternatively, new methods for the combinatorial chemical synthesis of peptide, polycarbamate, and oligonucleotide arrays have recently been reported (see Fodor et al., "Light-Directed, Spatially Addressable Parallel Chemical Synthesis," Science 251:767-773 (1991); Cho et al., "An Unnatural Biopolymer," Science, 261:1303-1305 (1993); and Southern et al., "Analyzing and Comparing Nucleic Acid Sequences by Hybridization to Arrays of Oligonucleotides: Evaluation Using Experimental Models," Genomics 13:1008- 10017 (1992), which are hereby incorporated by reference in their entirety).

These arrays (see Fodor et al., "Multiplexed Biochemical Assays with Biological Chips," Nature 364:555-556 (1993), which is hereby incorporated by reference in its entirety) harbor specific chemical compounds at precise locations in a high- density, information rich format, and are a powerful tool for the study of biological recognition processes.

[0040] Preferably, the probes are attached to the leads through spatially directed oligonucleotide synthesis. Spatially directed oligonucleotide synthesis may be carried out by any method of directing the synthesis of an oligonucleotide to a specific location on a substrate. Methods for spatially directed oligonucleotide synthesis include, without limitation, microlithography, light- directed oligonucleotide synthesis, application by ink jet, microchannel deposition to specific locations, and sequestration with physical barriers. In general, these methods involve generating active sites, usually by removing protective groups, and coupling to the active site a nucleotide which, optionally, has a protected active site if further nucleotide coupling is desired.

[0041] In one embodiment, the lead-bound oligonucleotides are synthesized at specific locations by light-directed oligonucleotide synthesis which is disclosed in U.S. Patent No. 5,143,854 to Pirrung et al., published PCT Application Serial No. WO 92/10092, and published PCT Application Serial No. WO 90/15070, which are hereby incorporated by reference in their entirety. In a basic strategy of this process, the surface of a solid support modified with linkers and photolabile protecting groups is illuminated through a photolithographic mask, yielding reactive hydroxyl groups in the illuminated regions. A 3'-O- phosphoramidite-activated deoxynucleoside (protected at the 5'-hydroxyl with a photolabile group) is then presented to the surface and coupling occurs at sites that were exposed to light. Following the optional capping of unreacted active sites and oxidation, the substrate is rinsed and the surface is illuminated through a second mask, to expose additional hydroxyl groups for coupling to the linker. A second 5'-protected, 3'-O-phosphoramidite-activated deoxynucleoside (C-X) is presented to the surface. The selective photodeprotection and coupling cycles are repeated until the desired set of probes are obtained. Photolabile groups are then optionally removed, and the sequence is, thereafter, optionally capped. Side chain protective groups, if present, are also removed. Since photolithography is used, the process can be miniaturized to specifically target leads in high densities on the support.

[0042] The protective groups can, themselves, be photolabile.

Alternatively, the protective groups can be labile under certain chemical conditions, e.g., acid. In this example, the surface of the solid support can contain a composition that generates acids upon exposure to light. Thus, exposure of a region of the substrate to light generates acids in that region that remove the protective groups in the exposed region. In addition, the synthesis method can use 3'-protected 5'-O-phosphoramidite-activated deoxynucleoside. In this case, the oligonucleotide is synthesized in the 5' to 3' direction, which results in a free 5' end.

[0043] The general process of removing protective groups by exposure to light, coupling nucleotides (optionally competent for further coupling) to the exposed active sites, and optionally capping unreacted sites is referred to herein as "light-directed nucleotide coupling."

[0044] The probes may be targeted to the electrically separated conductors by using a chemical reaction for attaching the probe or nucleotide to the conductor which preferably binds the probe or nucleotide to the conductor rather than the support material. Alternatively, the probe or nucleotide may be targeted to the conductor by building up a charge on the conductor which electrostatically attracts the probe or nucleotide.

[0045] Nucleases can be used to remove probes which are attached to the wrong conductor. More particularly, a target nucleic acid molecule may be added to the probes. Targets which bind at both ends to probes, one end to each conductor, will have no free ends and will be resistant to exonuclease digestion. However, probes which are positioned so that the target cannot contact both conductors will be bound at only one end, leaving the molecule subject to digestion. Thus, improperly located probes can be removed while protecting the properly located probes. After the protease is removed or inactivated, the target nucleic acid molecule can be removed and the device is ready for use. The capture probes can be formed from natural nucleotides, chemically modified nucleotides, or nucleotide analogs, as long as they have activated hydroxyl groups compatible with the linking chemistry. Such RNA or DNA analogs include, but are not limited to, 2'-O-alkyl sugar modifications, methylphosphonate, phosphorothioate, phosphorodithioate, formacetal, 3'-thioformacetal, sulfone, sulfamate, and nitroxide backbone modifications, amides, and analogs, where the base moieties have been modified. In addition, analogs of oligomers may be polymers in which the sugar moiety has been modified or replaced by another suitable moiety, resulting in polymers which include, but are not limited to, polyvinyl backbones (Pitha et al., "Preparation and Properties of Poly (I- vinylcytosine)," Biochim. Biophvs. Acta 204:381-8 (1970); Pitha et al., "Poly(l- vinyluracil): The Preparation and Interactions with Adenosine Derivatives," Biochim. Biophys. Acta 204:39-48 (1970), which are hereby incorporated by reference in their entirety), morpholino backbones (Summerton et al., "Morpholino Antisense Oligomers: Design, Preparation, and Properties," Antisense Nucleic Acid Drug Dev. 7:187-9 (1997), which is hereby incorporated by reference in its entirety) and peptide nucleic acid (PNA) analogs (Stein et al., "A Specificity Comparison of Four Antisense Types: Morpholino, 2'-O-methyl RNA, DNA, and Phosphorothioate DNA," J. Antisense Nucleic Acid Drug Dev. 7:151-7 (1997); Egholm et al., "Peptide Nucleic Acids (PNA)-Oligonucleotide Analogues with an Achiral Peptide Backbone," J. Am . Chem Soc. 114: 1895- 1897 (1992); Faruqi et al., "Peptide Nucleic Acid-Targeted Mutagenesis of a Chromosomal Gene in Mouse Cells." Proc. Natl. Acad. Sci. USA 95:1398-403 (1998); Christensen et al., "Solid-Phase Synthesis of Peptide Nucleic Acids," 1 Pent. Sci. 1:175-83 (1995); Nielsen et al., "Peptide Nucleic Acid (PNA). A DNA Mimic with a Peptide Backbone," Bioconjug. Chem. 5:3-7 (1994), which are hereby incorporated by reference in their entirety).

[0046] Capture probes suitable for the detection cartridge of the present invention include, without limitation, oligonucleotide, peptide nucleic acids, or peptide nucleic acids analogs. The capture probes can contain the following exemplary modifications: pendant moieties, such as proteins (e.g., nucleases, toxins, antibodies, signal peptides, and poly-L-lysine), intercalators (e.g., acridine and psoralen), chelators (e.g., metals, radioactive metals, boron, and oxidative metals), alkylators, and other modified linkages (e.g., alpha anomeric nucleic acids). Such analogs include various combinations of the above-mentioned modifications involving linkage groups and/or structural modifications of the sugar or base for the purpose of improving RNaseH-mediated destruction of the targeted RNA, binding affinity, nuclease resistance, and or target specificity. [0047] The present invention relates to testing ambient environment for biological agents. Airborne samples can be collected by passing air over a filter for a constant time. The filter can be washed with lysis buffer. Alternatively, the filter can be placed directly into the lysis buffer. Waterborne samples can be collected by passing a constant amount of water over a filter. The filter can then be washed with lysis buffer or soaked directly in the lysis buffer. Dry samples can be directly deposited into lysis buffer for removal of the organism of interest.

[0048] When whole cells, such as bacterial cells or virus samples are being analyzed, it is typically necessary to extract the nucleic acids from the cells or viruses, prior to continuing with the various sample preparation operations. Accordingly, following sample collection, nucleic acids may be liberated from the collected cells, viral coat, etc., into a crude extract, followed by additional treatments to prepare the sample for subsequent operations such as denaturation of contaminating (DNA binding) proteins, purification, filtration, and desalting. [0049] Liberation of nucleic acids from the sample cells or viruses and denaturation of DNA binding proteins may generally be performed by physical or chemical methods. For example, chemical methods generally employ lysing agents to disrupt the cells and extract the nucleic acids from the cells, followed by treatment of the extract with chaotropic salts such as guanidinium isothiocyanate or urea to denature any contaminating and potentially interfering proteins. Generally, where chemical extraction and/or denaturation methods are used, the appropriate reagents may be incorporated within the extraction chamber or a separate accessible chamber, or may be externally introduced. [0050] Alternatively, physical methods may be used to extract the nucleic acids and denature DNA binding proteins. U.S. Patent No. 5,304,487 to Wilding et al., which is hereby incorporated by reference in its entirety, discusses the use of physical protrusions within microchannels or sharp edged particles within a chamber or channel to pierce cell membranes and extract their contents. More traditional methods of cell extraction may also be used, e.g., employing a channel with restricted cross-sectional dimension which causes cell lysis when the sample is passed through the channel with sufficient flow pressure. Alternatively, cell extraction and denaturing of contaminating proteins may be carried out by applying an alternating electrical current to the sample. More specifically, the sample of cells is flowed through a microtubular array while an alternating electric current is applied across the fluid flow. A variety of other methods may be utilized within the device of the present invention to effect cell lysis/extraction, including, e.g., subjecting cells to ultrasonic agitation, or forcing cells through microgeometry apertures, thereby subjecting the cells to high shear stress resulting in rupture. [0051] Following extraction, it is often desirable to separate the nucleic acids from other elements of the crude extract, e.g., denatured proteins, cell membrane particles, and the like. Removal of particulate matter is generally accomplished by filtration, flocculation, or the like. Ideally, the sample is concentrated by filtration, which is more rapid and does not require special reagents. A variety of filter types may be readily incorporated into the device. Samples can be forced through filters that will allow only the cellular material to pass through, trapping whole organisms and broken cell debris. Further, where chemical denaturing methods are used, it may be desirable to desalt the sample prior to proceeding to the next step. Desalting of the sample, and isolation of the nucleic acid may generally be carried out in a single step, e.g., by binding the nucleic acids to a solid phase and washing away the contaminating salts or performing gel filtration chromatography on the sample. Suitable solid supports for nucleic acid binding include, e.g., diatomaceous earth, silica, or the like. Suitable gel exclusion media is also well known in the art and is commercially available from, e.g., Pharmacia (Piscataway, NJ ) and Sigma Chemical (St. Louis, MO). This isolation and/or gel filtration desalting may be carried out in an additional chamber, or alternatively, the particular chromatographic media may be incorporated in a channel or fluid passage leading to a subsequent reaction chamber.

[0052] The probes are preferably selected to bind with the target such that they have approximately the same melting temperature. This can be done by varying the lengths of the hybridization region. A-T rich regions may have longer target sequences, whereas G-C rich regions would have shorter target sequences. [0053] Hybridization assays on substrate-bound oligonucleotide arrays involve a hybridization step and a detection step. In the hybridization step, the sample potentially containing the target and an isostabilizing agent, denaturing agent or renaturation accelerant is brought into contact with the probes of the array and incubated at a temperature and for a time appropriate to allow hybridization between the target and any complementary probes. [0054] Including a hybridization optimizing agent in the hybridization mixture significantly improves signal discrimination between perfectly matched targets and single-base mismatches. As used herein, the term "hybridization optimizing agent" refers to a composition that decreases hybridization between mismatched nucleic acid molecules, i.e., nucleic acid molecules whose sequences are not exactly complementary.

[0055] An isostabilizing agent is a composition that reduces the base-pair composition dependence of DNA thermal melting transitions. More particularly, the term refers to compounds that, in proper concentration, result in a differential melting temperature of no more than about 1°C for double stranded DNA oligonucleotides composed of AT or GC, respectively. Isostabilizing agents preferably are used at a concentration between 1 M and 10 M, more preferably between 2 M and 6 M, most preferably between 4 M and 6 M, and, optimally, at about 5 M. For example, a 5 M agent in 2X SSPE (sodium chloride/sodium phosphate/EDTA solution) is suitable. Betaines and lower tetraalkyl ammonium salts are examples of suitable isostabilizing agents. [0056] Betaine (N,N,N,-trimethylglycine (Rees et al., "Betaine Can

Eliminate the Base Pair Composition Dependence of DNA Melting," Biochem. 32:137-144 (1993), which is hereby incorporated by reference in its entirety) can eliminate the base pair composition dependence of DNA thermal stability. Unlike teframethylammonium chloride ("TMACl"), betaine is zwitterionic at neutral pH and does not alter the polyelectrolyte behavior of nucleic acids while it does alter the composition-dependent stability of nucleic acids. Inclusion of betaine at about 5 M can lower the average hybridization signal, but increases the discrimination between matched and mismatched probes. [0057] A denaturing agent is a composition that lowers the melting temperature of double stranded nucleic acid molecules by interfering with hydrogen bonding between bases in a double-stranded nucleic acid or the hydration of nucleic acid molecules. Denaturing agents can be included in hybridization buffers at concentrations of about 1 M to about 6 M and, preferably, about 3 M to about 5.5 M.

[0058] Denaturing agents include formamide, formaldehyde, dimethylsulfoxide ("DMSO"), tetraethyl acetate, urea, guanid ne thiocyanate ("GuSCN"), glycerol and chaotropic salts. As used herein, the term "chaotropic salt" refers to salts that function to disrupt van der Waal's attractions between atoms in nucleic acid molecules. Chaotropic salts include, for example, sodium trifluoroacetate, sodium tricholoroacetate, sodium perchlorate, and potassium thiocyanate.

[0059] A renaturation accelerant is a compound that increases the speed of renaturation of nucleic acids by at least 100-fold. They generally have relatively unstructured polymeric domains that weakly associate with nucleic acid molecules. Accelerants include heterogeneous nuclear ribonucleoprotein ("hnRP") Al and cationic detergents such as, preferably, cetyltrimethylammonium bromide ("CTAB") and dodecyl trimethylammonium bromide ("DTAB"), and, also, polylysine, spermine, spermidine, single stranded binding protein ("SSB"), phage T4 gene 32 protein, and a mixture of ammonium acetate and ethanol. Renaturation accelerants can be included in hybridization mixtures at concentrations of about 1 μM to about 10 mM and, preferably, 1 μM to about 1 mM. The CTAB buffers work well at concentrations as low as 0.1 mM. [0060] Addition of small amounts of ionic detergents (such as N-lauroyl- sarkosine) to the hybridization buffers can also be useful. LiCl is preferred to NaCl. Hybridization can be at 20°-65°C, usually 37°C to 45°C for probes of about 14 nucleotides. Additional examples of hybridization conditions are provided in several sources, including: Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd Ed., Cold Spring Harbor, N.Y. (1989); and Berger and Kimmel, "Guide to Molecular Cloning Techniques," Methods in Enzymology Vol. 152, Academic Press, Inc., San Diego, Calif. (1987); Young et al., "Efficient Isolation of Genes by Using Antibody Probes," Proc. Natl. Acad. Sci. USA 80:1194 (1983), which are hereby incorporated by reference in their entirety.

[0061] In addition to aqueous buffers, non-aqueous buffers may also be used. In particular, non-aqueous buffers which facilitate hybridization but have low electrical conductivity are preferred. [0062] The sample and hybridization reagents are placed in contact with the probe array and incubated. Generally, incubation will be at temperatures normally used for hybridization of nucleic acids, for example, between about 20°C and about 75°C, e.g., about 25°C, about 30°C, about 35°C, about 40°C, about 45°C, about 50°C, about 55°C, about 60°C, or about 65°C. For probes longer than about 14 nucleotides, 37-45°C is preferred. For shorter probes, 55-65°C is preferred. More specific hybridization conditions can be calculated using formulae for determining the melting point of the hybridized region. Preferably, hybridization is carried out at a temperature at or between ten degrees below the melting temperature and the melting temperature. More preferred, hybridization is carried out at a temperature at or between five degrees below the melting temperature and the melting temperature. The target is incubated with the capture probes for a time sufficient to allow the desired level of hybridization between the target and any complementary capture probes. After incubation with the hybridization mixture, the electrically separated conductors are washed with the hybridization buffer, which also can include the hybridization optimizing agent. These agents can be included in the same range of amounts as for the hybridization step, or they can be eliminated altogether. [0063] Processing of the sample within first chamber of the detection cartridge can involve neutralizing the sample, contacting the neutralized sample with a buffer, treating the sample with conductive ions, and treating the sample with an enhancer, all which are stored in a plurality of reagent containers within the detection cartridge which are positioned so as to discharge into the first chamber of the cartridge as needed. Molecules that are not captured are expelled from the first chamber through a second conduit and into a second chamber. The detection system can be programmed by a series of operation buttons on the front of the device and the results can be seen on a visual display, or alternatively, sent to a central monitoring station or one or more remote monitoring stations. [0064] In one embodiment, the present invention can be used for real time detection of biological warfare agents. With the recent concerns over the use of biological weapons in a theater of war and in terrorist attacks, the device could be configured into a personal sensor for the combat soldier, attached to combat or reconnaissance vehicles, or placed into a remote sensor for advanced warnings of a biological threat. The device, which can be used to specifically identify the agent, can be coupled with a modem to send the information to another location. Mobile devices may also include a global positioning system to provide both location and pathogen information.

[0065] In this aspect, suitable capture probes include, without limitation, those able to identify bacteria such as Bacillus spp., Pseudomonas spp., E. coli spp. or Enterobacteria spp. Because more than one agent may be present in a given sample, one embodiment of the present invention provides detection chips having multiple test sites and multiple capture probes.

[0066] Although preferred embodiments have been depicted and described in detail herein, it will be apparent to those skilled in the relevant art that various modifications, additions, substitutions, and the like can be made without departing from the spirit of the invention and these are therefore considered to be within the scope of the invention as defined in the claims which follow.

Claims

WHAT IS CLAIMED:
1. A method for detecting a target molecule comprising: providing a detection system comprising: a plurality of detection cartridges, where each detection cartridge comprises: a housing defining a first chamber and a detection chip within the first chamber defined by said housing, wherein said detection chip comprises: two or more electrically separated conductors fabricated on a substrate and capture probes attached to the conductors such that a gap exists between the capture probes on the electrically separated conductors, wherein a sample, potentially containing a target molecule, can be analyzed for the presence of the target molecule by determining whether the conductors are electrically connected; an electrical connector extending through the housing and coupled to the electrically separated conductors so that the presence of a target molecule connecting the capture probes on the electrically separated conductors can be detected; a support unit into which the detection cartridge can be positioned to carry out a procedure for detecting the target molecule in the sample, wherein said support unit has an electrical coupler suitable for electrical communication with the electrical connector of the detection cartridge, whereby the presence of the target molecule in the sample can be detected and communicated to the support unit; and a detection cartridge manipulator suitable for holding the plurality of detection cartridges and positioned to insert unused detection cartridges individually into the support unit for detection of a target molecule in a sample and for removing detection cartridges individually from the support unit after use; inserting an unused detection cartridge into the support unit; processing a sample within the first chamber of the detection cartridge under conditions effective to permit any target molecules present in the sample to bind to the capture probes and thereby connect the capture probes; and determining whether the electrical conductors are electrically connected, thereby detecting the presence of the target molecule in the sample.
2. The method according to claim 1, wherein the detection cartridge further comprises: a sample pre-treatment chamber positioned upstream of the first chamber.
3. The method according to claim 1, wherein the detection cartridge manipulator is a magazine comprising a plurality of separate numbered slots.
4. The method according to claim 1 , wherein the detection cartridge manipulator is a rotatable carousel comprising a plurality of separate numbered slots, each slot housing an individual detection cartridge.
5. The method according to claim 1 , wherein the detection cartridge manipulator is a tape reel structure comprising a plurality of slots, each slot housing an individual detection cartridge.
6. The method according to claim 1 further comprising: returning the individual detection cartridge to the detection cartridge manipulator after said determining.
7. The method according to claim 1 , wherein said processing comprises: neutralizing the sample; contacting the neutralized sample with a buffer; treating the sample with a conductive ion solution after said contacting with a buffer; and treating the sample with an enhancer after said treating with a conductive ion solution.
8. The method according to claim 1, wherein the support unit further comprises: a controller in communication with the electrical connector to enable electrical communication between the electrical conductors of the detection cartridge and the controller, whereby the presence of the target molecule in the sample can be detected and communicated to the controller.
9. The method according to claim 8, wherein the support unit further comprises: digital coupling to permit the controller to communicate data to a monitoring station external to the support unit.
10. The method according to claim 9, wherein the data is the result of said determining.
11. The method according to claim 9, wherein the data is the number of procedures detecting the target molecule that have been carried out by the detection system.
12. The method according to claim 9, wherein the monitoring station provides notice for replacement of the detection cartridges in the detection cartridge manipulator.
13. The method according to claim 9, wherein the monitoring station is a central monitoring station.
14. The method according to claim 9, wherein the monitoring station is a remote monitoring station.
15. The method according to claim 1 , wherein the two or more electrically separated conductors are in the form of spaced apart conductive fingers.
16. The method according to claim 15, wherein a plurality of pairs of spaced apart conductive fingers are present on the detection chip.
17. The method according to claim 1 , wherein the capture probes are oligonucleotides.
18. The method according to claim 1 , wherein the capture probes are peptide nucleic acid analogs.
19. The method according to claim 1 , wherein the detection chip comprises multiple separated conductors and multiple capture probes.
20. The method according to claim 1 , wherein the capture probes are specific for a biological warfare agent.
21. The method according to claim 20, wherein the biological warfare agent is a bacteria selected from the group consisting of Bacillus spp., Pseudomonas spp.. E. coli spp., and Enterobacteria spp.
22. The method according to claim 1, wherein said method is used to detect a target molecule in an environmental region, said method further comprising: scanning the environmental region with a continuous particle- monitoring device coupled to the detection system, wherein the continuous particle monitoring device determines particle levels in the environmental region.
23. The method according to claim 22 further comprising: activating an aerosol collector comprising a filter and a sample concentrator when a preset value is exceeded.
24. The method according to claim 23, wherein said activating comprises: taking in an air sample through the aerosol collector; filtering the air sample; wetting the filtered air sample; and transferring the air sample to the detection system.
25. A device for detecting a target molecule comprising: a detection system comprising: a plurality of detection cartridges, where each detection cartridge comprises: a housing defining a first chamber and a detection chip within the first chamber defined by said housing, wherein said detection chip comprises: two or more electrically separated conductors fabricated on a substrate and capture probes attached to the conductors such that a gap exists between the capture probes on the electrically separated conductors, wherein a sample, potentially containing a target molecule, can be analyzed for the presence of the target molecule by determining whether the conductors are electrically connected; an electrical connector extending through the housing and coupled to the electrically separated conductors so that the presence of a target molecule connecting the capture probes on the electrically separated conductors can be detected; a support unit into which the detection cartridge can be positioned to carry out a procedure for detecting the target molecule in the sample, wherein said support unit has an electrical coupler suitable for electrical communication with the electrical connector of the detection cartridge, whereby the presence of the target molecule in the sample can be detected and communicated to the support unit; and a detection cartridge manipulator suitable for holding the plurality of detection cartridges and positioned to insert unused detection cartridges individually into the support unit for detection of a target molecule in a sample and for removing detection cartridges individually from the support unit after use.
26. The device according to claim 25, wherein the detection cartridge further comprises: a sample pre-treatment chamber positioned upstream of the first chamber.
27. The device according to claim 25, wherein the detection cartridge manipulator is a magazine comprising a plurality of separate numbered slots.
28. The device according to claim 25, wherein the detection cartridge manipulator is a rotatable carousel comprising a plurality of separate numbered slots, each slot housing an individual detection cartridge.
29. The device according to claim 25, wherein the detection cartridge manipulator is a tape reel structure comprising a plurality of slots, each slot housing an individual detection cartridge.
30. The device according to claim 25, wherein the detection system further comprises: a controller in communication with the electrical connector of the detection cartridge to enable electrical communication between the electrical conductors of the detection cartridge and the controller, whereby the presence of the target molecule in the sample can be detected and communicated to the controller.
31. The device according to claim 25, wherein the support unit further comprises: a digital coupling to permit the controller to communicate data to a monitoring station external to the support unit.
32. The device according to claim 31 , wherein the monitoring station is a central monitoring station.
33. The device according to claim 31 , wherein the monitoring station is a remote monitoring station.
34. The device according to claim 25, wherein the two or more electrically separated conductors are in the form of spaced apart conductive fingers.
35. The device according to claim 34, wherein a plurality of pairs of spaced apart conductive fingers are present on the detection chip.
36. The device according to claim 25, wherein the capture probes are oligonucleotides. ,
37. The device according to claim 25, wherein the capture probes are peptide nucleic acid analogs.
38. The device according to claim 25, wherein the detection chip comprises multiple separated conductors and multiple capture probes.
39. The device according to claim 25 further comprising: a continuous particle-monitoring device coupled to the detection system, wherein the continuous particle monitoring device determines particle level in the environmental region.
40. The device according to claim 39 further comprising an aerosol collector comprising: a filter and a sample concentrator.
PCT/US2004/018374 2003-06-12 2004-06-09 A low maintenance detector for biological agents WO2005019413A3 (en)

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