Chemistry: molecular biology and microbiology – Apparatus – Including measuring or testing
Reexamination Certificate
1999-12-02
2004-01-06
Ponnaluri, Padmashri (Department: 1639)
Chemistry: molecular biology and microbiology
Apparatus
Including measuring or testing
C435S288200, C435S288500, C435S007100, C435S008000, C422S051000, C422S051000, C422S067000, C422S082050
Reexamination Certificate
active
06673596
ABSTRACT:
1.0 BACKGROUND OF THE INVENTION
1.1 Field of the Invention
The invention generally relates to the field of implantable diagnostic devices (i.e. devices deployed within the body of an animal) for monitoring one or more target substances, analytes, or metabolites in the animal. More particularly, the invention provides implantable biosensor devices for monitoring and regulating the level of analytes in the tissues and circulatory system of a human. In illustrative embodiments, the apparatus comprises a biosensor that is utilized to monitor the level of blood glucose in a diabetic or hypoglycemic patient. The disclosed sensors may also be used to control or regulate the delivery of a drug or other pharmaceutical agent from an external or an implantable drug delivery system. For example, the device may form part of an artificial pancreas to regulate insulin dosage in response to the level of glucose detected in situ.
1.2 Description of Related Art
1.2.1 Biosensors
Biosensors are hybrid devices combining a biological component with an analytical measuring element. The biological component reacts and/or interacts with the analyte(s) of interest to produce a response measurable by an electronic, optical, or mechanical transducer. The most common configurations presently available utilize immobilized macromolecules such as enzymes or antibodies to form the biological component. Examples of analytes and immobilized macromolecules include: glucose and immobilized glucose oxidase (e.g., Wilkins et al., 1995); nitrate and immobilized nitrate reductase (Wu et al., 1997); hydrogen peroxide and 2,3-dichlorophenoxyacetic acid and immobilized horseradish peroxidase (Rubtsova et al., 1998); and aspartate and immobilized L-aspartase (Campanella et al., 1995).
1.2.2 Whole-cell Biosensors
A further refinement for biosensors has been developed in recent years that utilizes intact living cells, such as a microorganism, or an eukaryotic cell or cell culture as an alternative to immobilized enzymes. Microbial cells are especially well suited for biosensor technologies; they are physically robust, capable of existing under extremely harsh and widely fluctuating environmental conditions, they possess an extensive repertoire of responses to their environment, and they can be genetically engineered to generate reporter systems that are highly sensitive to these environmental responses. Polynucleotide sequences that comprise specific promoter sequences are operably linked to a gene or a plurality of genes that encode the desired reporter enzyme(s) and then introduced into and maintained within the living cell. When the target analyte is present, the reporter genes are expressed, generating the enzyme(s) responsible for the production of the measured signal. Commonly used reporter systems have utilized either the &bgr;-galactosidase (lacZ) or catechol-2,3-dioxygenase (xylE) enzymes (Kricka, 1993).
Unfortunately, a limitation of these systems has been that following exposure to the target substance(s), the cells must be destructively lysed and the enzyme(s) isolated. This lysis is then followed by the addition of one or more secondary metabolites to yield a colorimetric signal that is proportional to the concentration of enzyme(s) in solution, providing a means to quantify the concentration of the original target substance.
A more recent improvement in such sensors utilizes green fluorescent protein as a reporter system, with the significant advantage that cells do not require destructive assay techniques to produce colorimetric signals. Because a substrate must be added to the green fluorescent protein constructs to first initiate the light response, however, these systems are quite complicated and offer little advantage for detection of analytes in situ (Prasher, 1995).
1.23 In Vivo Sensors
The development of an integrated in vivo implantable glucose monitor was first reported by Wilkins and Atanasov (1995). This system utilizes glucose oxidase immobilized within a micro-bioreactor. This enzyme catalyzes the oxidation of &bgr;-D-glucose by molecular oxygen to yield gluconolactone and hydrogen peroxide, with the concentration of glucose being proportional to the consumption of O
2
or the production of H
2
O
hd 2
. Unfortunately, the presence of a glucose oxidase inhibitor molecule in the human bloodstream tended to offset proportionality constants, and made the device unsatisfactorily inaccurate for precise glucose monitoring and control (Gough et al., 1997). Also limiting was the device's relatively large size (≈5×7 cm), which negated its usefulness as an implantable device.
Although several smaller needle-type and microdialysis glucose sensors have since been developed to circumvent size limitations (Gough et al., 1997, Selam, 1997), their reliance on a glucose oxidase enzyme-based system limits their overall effectiveness and reliability.
Several nonspecific electrochemical sensors have also been investigated as potential in vivo glucose sensors (e.g., Yao et al., 1994; Larger et al., 1994), but problems including limited sensitivity, instability, and limited long-term reliability have prevented their wide-spread utilization (Patzer et al., 1995). According to Atanasov et al. (1997), continuously functioning implantable glucose biosensors with long-term stability have yet to be achieved.
1.3 Deficiencies in the Prior Art
Despite a significant miniaturization of biosensors during the past decade, they are still relatively large and obtrusive to serve as ideal implantable devices. Current methodologies using mammalian bioluminescent reporter cells require cell lysis and addition of an exogenous substrate to generate a measurable response. Consequently, these cells cannot serve as continuous on-line monitoring devices.
Therefore, there remains a need for the development of a small implantable monolithic (i.e. containing both biological and electrical components constructed on a single substrate layer) bioelectronic monitor that is durable, inexpensive, wireless, and that can communicate remotely to a drug delivery system to provide the controlled delivery of a therapeutic agent such as insulin.
2.0 SUMMARY OF THE INVENTION
The present invention overcomes these and other inherent limitations in the prior art by providing implantable apparatus and methods for detecting and quantitating particular analytes in the body of an animal. In particular, the invention provides devices for the in vivo detection and quantitation of metabolites, drugs, hormones, toxins, or microorganisms such as viruses in a human or animal. In illustrative embodiments, the invention provides a BBIC device useful for the detection of glucose in a human. Such devices provide for the first time an accurate on-line detector for glucose monitoring, and offer the ability to control the administration of pharmaceutical agents via an external or implantable drug delivery system. Also disclosed are BBIC devices for detecting the concentration of signature molecules (i.e. proteins released from cancer cells, etc.), clotting factors, enzymes and the like, and other analytes present in the bloodstream or interstitial fluid. In the area of oncology, the biosensor devices find utility in both initial and remission monitoring, on-line measurement of the effectiveness of chemotherapy, and stimulation/activity of the immune system. Likewise, the biosensor devices are useful in other areas of medicine, including on-line monitoring for enzymes associated with the occurrence of blood clots (strokes, heart attacks, etc.), detection and quantitation of clotting factors (maintain level), hormone replacement, continuous drug monitoring (testing for controlled substances in prisoners, military personnel, etc.), monitoring of soldiers exposure to sub-lethal exposure to nerve agents and other debilitating agents, monitor levels of compounds affecting mental illness, and the like.
In one embodiment there is provided an implantable monolithic bioelectronic device for detecting an analyte within the body of an animal. In a general sense this device c
Applegate Bruce M.
Ripp Steven A.
Sayler Gary S.
Simpson Michael L.
Akerman & Senterfitt
Ponnaluri Padmashri
Tran My Chau
UT-Battelle LLC
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