Abstract:
Chemical reactions occurring within a living cell are measured in a manner that does not affect the viability of the cell or the reaction under study. In one embodiment, one or more sensors are introduced into the cell and/or covalently associated with the exterior cell membrane. The sensor(s) emit an observable signal indicating a value of a parameter associated with the chemical reaction, e.g., the concentration of a reaction product. Because cell viability is not compromised, the cell may be stimulated (e.g., by subjection to an agonist or antagonist, a pathogen, a pharmaceutical compound, or a potential toxin) so as to affect the reaction under study.

Description:
CROSS-REFERENCE TO RELATED CASES 
       [0001]    This application claims the benefit of U.S. Provisional Patent Application No. 60/718,255, entitled “Optical Biosensor Array and Related Systems, Methods and Devices,” filed on Sep. 16, 2005, the entire disclosure of which is hereby incorporated by reference as if set forth herein in its entirety. 
     
    
     FIELD OF THE INVENTION 
       [0002]    The invention relates to monitoring chemical reactions occurring within living cells, and more particularly to methods and apparatus for measuring such reactions without compromising the viability of the cells. 
       BACKGROUND OF THE INVENTION 
       [0003]    Numerous techniques exist for monitoring the extent and time profile of a chemical reaction of interest occurring within a living cell. Many such techniques are indirect, involving an assay for reaction products outside the cell that reflect the progress of reaction(s) taking place in the cytoplasm. A disadvantage of such assays is their vulnerability to influence by the spurious presence of reaction products in the cell&#39;s environment that do not, in fact, emanate from the cell. 
         [0004]    Intracellular techniques have also been developed; these include, for example, genetically engineering cells to fluoresce in the presence of a compound of interest, radioactive binding assays, and colorimetric assays. These techniques, however, require potentially destructive modification or invasion of the cell. 
         [0005]    Intracellular and extracellular assays are often employed to track enzymatic activity by measuring the amount of reaction product through the use of coupling. Enzymatic coupling is a complicated approach that links a non-quantifiable enzymatic reaction of interest with an optically measurable (by means of a spectrophotometer) enzymatic reaction that will interact with the products of the enzymatic reaction of interest. Once again, conventional approaches suffer from a number of deficiencies. For example, coupled reactions necessarily involve at least one chemical reaction unrelated (“coupled”) to the enzymatic activity of interest, imparting potential sources of error. Enzymatic reactions require carefully controlled reaction conditions (appropriate temperatures, pHs, salt concentrations, etc.) and, as a result, monitoring efforts can be affected by rapid environmental changes or degradation of reagents, e.g., components of the enzymatic reactions (especially the enzyme itself). 
         [0006]    Accordingly, there is a need for apparatus and methods that quickly, directly and accurately measure a product of interest. 
       SUMMARY OF THE INVENTION 
       [0007]    The present invention addresses the shortcomings of the prior art by measuring chemical reactions occurring within a living cell in a manner that does not significantly affect the viability of the cell or the reaction under study. By “significantly” is meant, with respect to individual cells, that these are not killed; with respect to a population of cells, that at least the majority remains viable; and with respect to a reaction, that the variable under study is not affected beyond a predetermined threshold of precision. For example, if it is necessary to monitor the concentration of an ion to within 0.1 mole/L, the reaction is not significantly affected if the ion concentration is not shifted more than ±0.05 mole/L. 
         [0008]    In one embodiment, one or more sensors are introduced into the cell and/or covalently associated with the exterior cell membrane. The sensor(s) produce an observable signal indicating a value of a parameter associated with the chemical reaction, e.g., the concentration of a reaction product. Because cell viability is not compromised, the cell may be stimulated (e.g., by subjection to an agonist or antagonist, a pathogen, a pharmaceutical compound, or a potential toxin) so as to affect the reaction under study. By “observable” is meant visible or measurable using instrumentation. 
         [0009]    In some embodiments, the sensor is used to monitor the activity of an ion channel through the cell membrane. In such cases, the sensor may be introduced within the cell, may be attached to the exterior cell membrane, or both. For example, the external sensor may be attached to the cell membrane proximate to an ion channel of the cell, e.g., via an antibody specific to the ion channel (which typically comprises a protein). The sensor may be an ion sensor, and sensors internal and external to the cell may monitor different ions. 
         [0010]    In some embodiments, the sensor comprises an ion-selective optode. The reaction may produce an ionic product that is sequestered by the optode, or instead, the reaction may produce a non-ionic product that is ionized to facilitate monitoring using the optode. In this way, the invention can be used in numerous pharmaceutical applications involving small molecules. 
         [0011]    In some embodiments, the optode comprises an ion-selective ionophore, a source of triggering ions, and a signaling agent responsive to the triggering ions. Typically the ionophore is selective for the ionic reaction product, and upon binding of a reaction-product ion, a triggering ion is released to interact with the signaling agent and thereby produce the signal. The strength of the signal may be indicative of the concentration of the triggering ion, which itself reflects the concentration of the ionic reaction product. Different sensors, each responsive to different ions and producing differentiable signals, can be used depending on the reaction(s) being monitored. Furthermore, the signal may be produced directly by the triggering ions, or may be produced indirectly (e.g., the triggering ions may initiate a reaction cascade that results in an observable signal). 
         [0012]    In preferred embodiments, the signal is optical in nature, e.g., involving fluorescence or absorbence. The optical signal may be monitored using, for example, a spectrometer, a flurometer, or a detector for measuring absorbance. But the signal may alternatively be non-visible or may involve other forms of electromagnetic radiation. 
         [0013]    The foregoing and other features and advantages of the present invention will be made more apparent from the description, drawings, and claims that follow. 
     
    
     
       BRIEF DESCRIPTION OF DRAWINGS 
         [0014]    The advantages of the invention may be better understood by referring to the following drawings taken in conjunction with the accompanying description in which: 
           [0015]      FIG. 1  is a schematic diagram of a cell assay system according to an illustrative embodiment of the invention. 
           [0016]      FIGS. 2A-2D  are cross-sections of various optical ion sensor arrangements suitable for use in various implementations of the cell assay system of  FIG. 1 . 
           [0017]      FIG. 3  presents another embodiment of a microfluidic device in accord with the present invention. 
           [0018]      FIG. 4  depicts the embodiment of  FIG. 5  with fluorescence from the channels imaged. 
           [0019]      FIG. 5  presents a hypothetical response of yeast cells to a stimulus (Haloperidol). 
           [0020]      FIG. 6  presents data from an exemplary study using a sensor in accord with the present invention to monitor the reaction of yeast cells to Haloperidol. Sodium is shown on the left and potassium on the right. 
           [0021]      FIG. 7  illustrates an embodiment of the present invention used as an ammonia sensor. 
           [0022]      FIG. 8  depicts the calibration curve for the embodiment of  FIG. 7 . 
           [0023]      FIG. 9  presents the data from real-time monitoring of Tgase reactions using sensors in accord with one embodiment of the present invention, using 12.5 mM substrate (Z-Gln-Gly) and 1.00, 0.20, and 0.02 units/ml Transglutaminase at 30° C. Each curve represents the average reading from 5-8 reactions. 
           [0024]      FIG. 10  presents the data from real-time monitoring of TGase reactions using sensors in accord with another embodiment of the present invention, having various substrate (Z-Gln-Gly) concentrations and 0.4 units/ml Transglutaminase at 30° C. Each curve represents the average reading from 3 reactions. 
       
    
    
       [0025]    In the drawings, like reference characters generally refer to corresponding parts throughout the different views. The drawings are not necessarily to scale, emphasis instead being placed on the principles and concepts of the invention. 
       DETAILED DESCRIPTION OF THE INVENTION 
       [0026]    In brief overview, embodiments of the present invention provide systems, methods and devices for measuring cellular biomarkers. In exemplary embodiments, optically active polymer sensors are placed inside or outside a cell. Changes in fluorescence or absorbance indicate the ion concentrations and fluxes from the cell. The overall change in fluorescence or absorbance in response to a compound of interest provides a “fingerprint” (e.g., a spectrum) for that compound. 
         [0027]      FIG. 1  is a schematic diagram of a cell assay system  100  according to an illustrative embodiment of the invention. The cell assay system  100  includes an optical ion sensor support  102 , a biological sample holder  104 , an excitation light source  106 , a light sensor  108 , and a computing device  111 . 
         [0028]    The optical ion sensor support  102  supports an optical ion sensor  112  for positioning in the biological sample holder  104 . In various implementations, the optical ion sensor  112  is adhered to the optical ion sensor support  102  by deposition in a solution of optical ion sensor matrices dissolved or dispersed in a solvent, such as in a polar organic solvent like tetrahydrofuran (THF). In such implementations, the sensor support  102  is preferably formed from a material resistant to the solvent. Materials resistant to THF include, without limitation,  304  stainless steel;  316  stainless steel; acetal polymer (marketed as DELRIN by E. I. du Pont de Nemours and Company); bronze; carbon graphite; carbon steel; ceramic Al 2 O 3 ; a perfluoroelastomer compound, such as CHEMRAZ marketed by Greene, Tweed; epoxy; HOSTELRY Calloy (marketed by Haynes International, Inc.); KALES elastomer (marketed by DuPont Performance Elastomers); polychlorotrifluoroethylene; NYLON (marketed by E. I. du Pont de Nemours and Company); polyetherether ketone (PEEK); polyphenylene sulfide; and PTFE. 
         [0029]    The optical ion sensor  112  includes a film having a suspension of optical ion sensor matrices. The optical ion sensor matrices, in general, include an ionophore, an additive, and a chromionophore suspended in a polymer phase, for example, of polyvinyl chloride (PVC). The polymer phase also includes a plasticizer such as dioctyl sebacate (DOS). An ionophore is a substance that allows targeted ions to move across or into a membrane. Preferably the ionophore is selected to be lipid-soluble. In addition, the ionophore is preferably an electrically neutral compound that forms a complex with a target ion. The ionophore is optically inactive in the visible spectrum and does not change absorbance or fluorescence depending on its state of complexation. 
         [0030]    A chromoionophore is an ionophore that changes its optical properties in the visible spectrum depending on the state of complexation. Chromoionophores are preferably proton-sensitive dyes that change absorbance (and fluorescence in many cases) depending on its degree of hydrogen complexation (protonation). The chromionophores are preferably highly lipophilic to prevent them from leaching out of the optical ion-sensor matrix. Suitable chromionophores include Chromoionophore II and Chromionophore III. Chromionophore II exhibits light absorbance peaks at 520 nm and 660 nm and a fluorescent emission peak at 660 nm. Chromionophore III has light absorbance peaks at 500 nm and 650 nm and fluorescent emission peaks at 570 nm and 670 nm. 
         [0031]    It should be stressed that the triggering ion released by the ionophore may be observed directly, as discussed above, or indirectly. For example, the intensity of the fluorescence signal may be a trigger for an event cascade within the cell. A large calcium current may cause the sensor to fluoresce brightly, for instance, whereas low calcium does not. The fluorescence may in turn excite a secondary dye in the particle that easily generates reactive oxygen species (ROS). The ROS would then attack the cell, effectively stimulating necrosis (cell death). Alternatively, instead of a secondary component within the particle, another particle may be added. This additional particle may comprise a photo-degradable polymer. When the primary sensory fluoresces, the emitted light will break apart the secondary particle, releasing its contents. The contents may, for example, be a drug that is therapeutic or cancer-fighting (stimulating apoptosis). 
         [0032]    For optical ion sensors targeting cations, the additive can be any inert lipophilic component that has a negative charge associated with it. For optical ion sensors targeting anions, the additive is positively charged. The purpose of the additive is to embed charge sites within the polymer phase, and to help enforce charge neutrality within the optical ion sensor  112 . The additive allows the polymer phase to carry an equal amount of charged particles as additive. The concentration ratio of additive to chromoionophore is preferably 1:1, thereby allowing the chromoionphore to become completely protonated or de-protonated. One suitable additive for optical ion sensors targeting negative ions is potassium tetrakis[3,5-bis(trifluoromethyl)phenyl]borate (KTFPB). The lipophilic, anionic component TFPB molecules are retained by the polymer phase, and the potassium ions are either complexed by the ionophore or expelled into the sample solution through diffusion. In one particular implementation, the optical ion sensor film is composed of a suspension produced from about 60 mg of DOS, 30 mg of PVC, and up to about 5 mg of additive, ionophore, and chromionophore. 
         [0033]    Once the above components are dissolved into the polymer phase to form the optical ion sensor  112  and are exposed to a sample solution, the optical ion sensor  112  becomes active. It now continuously extracts or expels analyte cations (a system can work with anions as well using ion coextraction) depending on ion activity in the sample solution. With a 1:1 additive-chromoionophore ratio, and with zero target ions present in the sample solution, the optical ion sensor  112  remains completely protonated to achieve charge neutrality. As the target ion concentration increases, the ionophores in the optical ion sensor  112  extract the target ions into the optical ion sensor  112 . To maintain charge neutrality of the optical ion sensor  112 , hydrogen (hydronium) ions are stripped from the chromoionophores in the optical ion sensor  112  and expelled into the sample solution. The expelling of hydrogen ions alters the pH of the optical ion sensor  112 , thereby altering its fluorescent properties. To detect analyte anions (for example, chloride or nitrite ions), the optical ion sensor uses ion-coextraction, as opposed to proton expulsion. To detect neutral analytes, an additional agent known to interact with the target analyte to yield an ion is added to the biological sample holder  104 . An ionophore is then selected to detect the resultant ion. 
         [0034]    The following is a non-limiting, illustrative list of target ion/ionophore pairings suitable for use in the optical ion sensors: potassium/Potassium Ionophore III (BME-44), sodium/Sodium Ionophore IV, sodium/Sodium Ionophore V, sodium/Sodium Ionophore VI, calcium/Calcium Ionophore III, and calcium/Calcium ionophore IV. For target anions, illustrative target ion/ionophore pairings include chloride/Chloride Ionophore III and nitrite/Nitrite Ionophore I. 
         [0035]    The film of the optical ion sensor can be produced in various ways. In one implementation, as described above, a predetermined amount of the optical ion sensor suspension (i.e., the combined polymer phase, ionophore, additive, and chromionophore) is dissolved in a solvent, such as THF. The solution is then deposited, sprayed, or spun onto a surface. The solvent evaporates, leaving the optical ion sensor film on the surface. 
         [0036]    In another implementation, the film is formed from a deposition of optical ion sensor microspheres. To produce the microspheres, an optical ion sensor emulsion is formed by injecting an optical ion sensor suspension dissolved in THF (e.g., 16 mL THF/100 mg PVC) into a pH buffered solution. The optical ion sensor suspension includes approximately 60 mg of DOS, 30 mg of PVC, and up to approximately 5 mg of chromionophore, additive, and ionophore. The emulsion is then submerged in a sonicating water bath. Typically, 50 μL of the optical ion sensor suspension/THF solution is injected into 1,000-1,500 μL of buffered solution. The resulting emulsion contains a mixture of spherical optical ion sensor particles ranging in size from 200 nm to 20 μm. The resulting emulsion can be spun, sprayed, or evaporated onto any surface to create a porous optical ion sensor membrane. Films formed from microspheres tend to expose a greater surface area of optical ion sensor to a given sample, yielding improved performance characteristics. 
         [0037]    The biological sample holder  104  holds a biological sample for analysis by the cell assay system  100 . The biological sample can include cells adhered to the walls of the biological sample holder  104 , for example, in a monolayer, or cells suspended in a liquid buffer. The biological sample holder  104  is preferably transparent, or at least includes a transparent region through which the optical ion sensor  112  can be excited and through which the results of such excitement can be monitored. 
         [0038]    The optical ion sensor  112  is illuminated with a light source  106  to excite the chromionophores suspended therein. The light source preferably can be tuned to generate one or more predetermined wavelengths of light, desirably in the visible portion of the electromagnetic spectrum, that are selected to excite the particular chromionophore used in the optical ion sensor  112 . Alternatively, the light source may generate a wide spectrum light. In one implementation, the light source  106  is coupled to the optical ion sensor support  102 . 
         [0039]    The fluorescence of the optical ion sensor  112  is detected by a light sensor  108 . The light sensor  108  may include a charge-coupled device, a fluorometer, a photomultiplier tube, or other suitable device for measuring fluorescence. In one implementation, a spectrophotofluorometer is used to satisfy the roles of the light source  106  and the light sensor  108 . The light sensor  108  may also be coupled to the optical ion sensor support  102 . 
         [0040]    The optical ion sensor support  102  includes an agent introduction means  118 . The agent introduction means  118  can include a pipette or an electromechanical dispenser device, such as a solenoid or electrostatically driven plunger or syringe. 
         [0041]    The computing device  111  controls the various components of the cell assay system  100 . The computing device  111  may be a single computing device or multiple computing devices providing the various functionalities used to control the cell assay system. These functionalities are provided by an excitation control module  126 , an agent introduction module  130 , and an analysis module  134 . The excitation control module  126  controls the light source  108  to emit one or wavelengths of excitation light. The agent introduction module  130  controls the introduction of an agent into the biological sample holder  104  via an agent introduction means  118 . The analysis module  134  analyzes the output of the light sensor  108 , e.g., before and after an agent is introduced into the biological sample holder  104  to determine the effect of the agent on the cells in the biological sample holder  104 . The analysis module  134  may also control the other modules in the computing device, i.e., the excitation control module  126  and the agent introduction module  130 , to coordinate an assay protocol. The computing device  111  and/or devices may also include various user interface components, such as a keyboard, mouse, trackball, printer, and display. 
         [0042]    A module may be implemented as a hardware circuit comprising custom VLSI circuits or gate arrays, off-the-shelf semiconductors such as logic chips, transistors, or other discrete components. A module may also be implemented in programmable hardware devices such as field programmable gate arrays, programmable array logic, programmable logic devices or the like. 
         [0043]    Modules may also be implemented in software for execution by various types of processors. An identified module of executable code may, for instance, comprise one or more physical or logical blocks of computer instructions which may, for instance, be organized as an object, procedure, or function. Nevertheless, the executables of an identified module need not be physically located together, but may comprise disparate instructions stored in different locations which, when joined logically together, comprise the module and achieve the stated purpose for the module. A module of executable code may be a single instruction, or many instructions, and may even be distributed over several different code segments, among different programs, and across several memory devices. 
         [0044]    The various modules are in communication with the various devices they control or obtain data from. They maybe connected over a local area network, wirelessly, over a bus, or over typical cables known in the art of computer interfaces for connecting computing devices with peripherals. 
         [0045]      FIGS. 2A-2D  depict various optical ion sensor arrangements suitable for use in various implementations of the cell assay system  100  of  FIG. 1 .  FIG. 2A  shows a first optical ion sensor arrangement that includes an optical ion sensor support  202  and a biological sample holder  204 . The biological sample holder  204  includes a monolayer of cells  206  adhered to the biological sample holder  204 . Alternatively, the biological sample holder  204  holds cells suspended in a buffer. The optical ion sensor support  202  and biological sample holder  204  correspond to the optical ion sensor support  102  and biological sample holder  104  of  FIG. 1 . An optical ion sensor film  210  is coupled to the distal end of the optical ion sensor support  202 . 
         [0046]      FIG. 2B  illustrates an alternative optical ion sensor arrangement  250 , which includes a biological sample holder  254  having therein a monolayer of cells  256  adhered to the surfaces of the biological sample holder  254 , or cells suspended in a buffer. Instead of including an optical ion sensor film adhered to a support, the optical ion sensor arrangement  250  relies upon optical ion sensor particles  258  introduced into the cells  256  adhered to the biological sample holder  254 . 
         [0047]    To introduce optical ion sensors into cells, the optical ion sensors are produced as particles  258 . The optical ion sensor particles  258  are fabricated in a fashion similar to the optical ion sensor film  112  described above. One such particle  258 , the optical ion sensor nanosphere, is produced according to the following procedure. First a optical ion sensor suspension is dissolved in 500 μl of THF. The suspension preferably includes 60 mg of DOS, 30 mg of PVC and up to about 5 mg of chromoionophore, ionophore, and additive to form an optode solution. Then, 500 μl of CH2Cl2 is added to bring the total volume to 1 ml. Next, a PEG-lipid solution is prepared by dissolving a PEG-lipid (such as DSPE-PEG 550 or DSPE-PEG (2000) Folate) in 5 ml of a water, salt and buffer solution. A TAT peptide can be added to the PEG-lipid via an amine linkage to aid the resulting nanospheres in entering cells. Alternatively, it is possible to utilize a molecule that is not a PEG, e.g., a ganglioside attached to ceramide. 
         [0048]    The nanospheres are formed by adding 100 μl of optode solution dropwise to 5 ml of the PEG-lipid solution while the solution is being sonicated by a probe tip sonicator. Additional sonication is performed for approximately 2-3 minutes. The resultant nanosphere solution is sprayed through a nitrogen-feed air gun into a beaker several times to remove excess solvent. If desired, the nanosphere solution is pushed through a 0.22 μm filter to remove the larger spheres. 
         [0049]    The optical ion sensor particles  258  are introduced into the cells  256  in any suitable manner. In one method, the particles  258  are introduced into a buffer liquid deposited in the biological sample holder  254 . A voltage source then generates a voltage sufficiently strong to electroporate the cells  256 , thereby allowing the optical ion sensor particles  258  to enter directly into the cells. In another approach, the surfaces of the optical ion sensor particles  258  are first coated with a substance, for example transferrin or folate, which assist the particles  258  in crossing through cell membranes. The optical ion sensor particles  258  are introduced into a buffer in the biological sample holder  254 , and the cells  256  bring the particles  258  into their interior in vesicles via endocytosis, pinocytosis, or phagocytosis, or similar biological processes. The substance applied to the optical ion sensor particles  258  breaks down the vesicle membrane, releasing the optical ion sensor particles  258  into the cell cytoplasm. In still other approaches, the particles  258  may be introduced into cells  256  using a glass needle or through ballistic bombardment. 
         [0050]    In some embodiments, the sensor is attached to the exterior of a cell rather than introduced into the interior. If, for example, the activity of an ion channel is to be studied, the sensor may be attached to the cell proximate to the ion channel. This may be accomplished, for example, by raising antibodies against the ion-channel protein and, using conventional chemistries, covalently linking one or more such antibodies to a sensor particle as described above. The antibody-studded particles are released into a cell suspension and preferentially bind to the ion channel. This approach can be used to link ion-specific sensors to any feature on the exterior of the cell membrane to which antibodies can be raised. Alternatively, the sensors may be attached to the cell membrane by other suitable coupling chemistries, e.g., biotin-(strept)avidin complexing or polysaccharide binding. See the thesis “High Throughput Optical Sensor Arrays for Drug Screening” by Daniel I. Harjes (2006), available from the Massachusetts Institute of Technology and incorporated herein by reference. 
         [0051]      FIG. 2C  illustrates a second alternative optical ion sensor arrangement  270  that includes an optical ion sensor support  272  and a biological sample holder  274 . An optical ion sensor film  276  is coupled to the distal end of the optical ion sensor support  272 . A cell monolayer  278  adheres to the surfaces of the biological sample holder  274 . Alternatively, cells are suspended in a buffer. In addition, optical ion sensor particles  280  are introduced into the cells of the cell monolayer  278 . Preferably the chromionophores used in the optical ion sensor film  276  differ from the chromionophores used in the optical ion sensor particles  280 . In particular, the different chromionophores desirably have distinguishable fluorescence characteristics such that an analysis module analyzing the output of a light sensor monitoring the optical ion sensor arrangement  270  can differentiate between the output of the optical ion sensor film  272  and the optical ion sensor particles  280 . As a result, the analysis module can differentiate between intracellular target ion concentration and extracellular target ion concentration. In addition, the optical ion sensor film  272  may include ionophores different from those included in the optical ion sensor particles  280 . Thus, the optical ion sensor arrangement  270  can monitor the concentrations of two different target ions. 
         [0052]      FIG. 2D  illustrates a third alternative optical ion sensor arrangement  290  that includes an electrode support  292  and a biological sample holder  294 . The biological sample holder  294 , in addition to a cell monolayer  296  or cells suspended in a buffer, includes a removable optical ion sensor film  298 . The removable optical ion sensor film  298 , for example, can be a glass cover slip or other transparent surface coated with an optical ion sensor film. 
         [0053]    In still another embodiment, the optical ion sensor film is coated onto the inner surface of the biological sample holder. And in another approach, to accommodate the 96-well plate format often used in assays, one embodiment of the present invention utilizes round glass coverslips coated with the ion-exchange optode material along with the cells to be monitored. In this embodiment, each well contains a single sensor type to track a specific species of interest; the various sensor types may differ only in the ionophore employed and utilize the same or similar chromoionophores. The compound of interest is then added directly to the well. The 96-well plate is then placed in a standard plate-reader type fluorometer, and the fluorescence intensity is monitored with time. 
         [0054]    In a typical implementation, a plurality of biological sample holders holding biological samples is provided. Biological samples introduced into the holders may include cells suspended in a buffer solution, but alternatively, cells may be adhered to the walls of the biological sample holders. Next, optical ion sensors are introduced into the biological sample holders as shown in  FIGS. 2A and 2C , and/or are introduced into the cells themselves. Alternatively, the optical ion sensors can coat the walls of the biological sample holders. As described above, optical ion sensor particles can be introduced either by electroporating the cells via electrodes positioned in the biological sample holders or by the chemistry applied to the optical ion sensor particles breaching vesicle membranes within the cells. Similarly, the optical ion sensor sensors can be introduced into the cells using pico-injection, bead loading, a gene gun, or through liposomal delivery techniques known in the art. As described above, the optical ion sensors include at least one ionophore for selectively binding a predetermined ion. In some embodiments, this results in altering the pH of the optical ion sensor and a pH-sensitive chromionophore for optically indicating the concentration of the ion in a fluid surrounding the optical ion sensor. Ion concentration, in other words, is indicated by the pH of the optical ion sensor and the resulting fluorescence of the chromionophore. 
         [0055]    An agent, such as a therapeutic, toxin, biological macromolecule (such as a nucleic acid, an antibody, a protein or portion thereof, e.g., a peptide), small molecule (of 2000 amu or less, 1000 amu or less or 500 amu or less), protein, virus, bacteria, chemical compound, mixture of chemical compounds, or an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues, or other biologically active agent may be introduced into one or more of the biological sample holders. In one particular implementation using an array of biological sample holders, no agent is introduced into a first row of biological sample holders to preserve a control. A first agent is introduced into a second row of biological sample holders. Additional agents are added to additional rows of the array of biological sample holders. 
         [0056]    The fluorescence of the optical ion sensors introduced into the biological sample holders is monitored. The monitoring preferably begins prior to introduction of the agents and continues thereafter. Changes in ion concentration resulting from the introduced agents are then determined. By comparing the changes in ion concentration after adding an agent, one can determine the effect of the agent on the cells being tested. 
         [0057]      FIGS. 3 and 4  present another embodiment  300  of a sensor in accord with the present invention. The sensor comprises a well  310  and micro-channels, indicated generally at  315 , branching successively therefrom. The cells are placed in the well  310 , sensor materials with different specificities line each of the channels  315 , and a solution is flowed from the well  310  through the channels  315 . When the fluorescence from the channels  315  is imaged, the pattern formed by changes in intensity may be visualized as an optical barcode, as depicted in  FIG. 4 , indicating the presence or absence of the various sensed materials in the solution. 
       Exemplary Applications: HERG Compatibility 
       [0058]    The sensors of the present invention are suited to many different types of assays, or for the detection of compounds. In one exemplary application, a model cell system is used to screen for HERG compatibility. HERG (human ether-a-go-go) channels are implicated in the abnormal heart rhythms associated with Long QT Syndrome. It is believed that many compounds can block these channels and cause prolongation of the beat cycle of the heart. This abnormal rhythm can cause arrhythmia and can even lead to death. Accordingly, it is important that all drugs, regardless of therapeutic target, be screened for HERG compatibility. 
         [0059]    With reference to  FIG. 5 , the model cell system to be used is  Saccharomyces cerevisae,  as it contains a native ERG2 gene which is similar to the human HERG. It has been shown that known HERG inhibitors also interfere with the function of ERG2 in yeast. Much is known genetically about yeast, and gene knock-outs are commercially available, allowing for the screening of two types of yeast: the wild type (WT) containing the ERG2, and one mutated to knock-out the ERG2 gene. 
         [0060]    With reference to  FIG. 6 , using both potassium and sodium sensors according to the present invention with both yeasts stimulated with a known HERG inhibitor, Haloperidol (an anti-psychotic drug), a change is observed in sodium and potassium upon stimulation of the cells with Haloperidol. In the ERG2 knock-out, there is no response. This example utilizes two cell types and two types of sensors, providing four different data signals, and the approach can be extended to an arbitrary number of sensors and cell types. Similarly, tissue can be used instead of culture. 
       Exemplary Applications: Ammonia Screening 
       [0061]    As explained above, pH changes can be used to render neutral reaction products of interest measurable using an ion-exchange optode. For a sensor developed to monitor ammonia production in biological reactions (e.g., involving an enzyme such as transglutaminase (TGase)), for example, a low pH ionizes ammonia to ammonium, which may be extracted by a nonactin ionophore. This approach greatly expands the range of measurable species. 
         [0062]    With reference to  FIG. 7 , an ammonia sensor is made by coating a 5 mm glass disk with an ammonium/ammonia-selective cocktail containing PVC, plasticizer, THF, potassium tetrakis (p-chlorophenyl)borate (KTpClPB), Chromoionophore III, and nonactin. A second, thin layer of 5% TEFLON is optionally applied to the coated disk to protect the ammonium cocktail from the high-salt environment required for the enzymatic reaction. These disks are placed in the bottom of individual wells in 96-well plates and the enzymatic reaction is set up to occur in the well, submerging the disks. Real-time monitoring of the reaction (through measuring the change in fluorescence of the optical sensor) is performed. Although these optodes are fairly large and coated with TEFLON, their response time is still approximately 2 minutes. 
         [0063]    To utilize data monitoring in order to track the change in fluorescence of the sensors in the presence of an enzymatic reaction, a calibration curve of fluorescence readings of known concentrations of the ion of interest is first constructed. With reference to  FIG. 8 , this curve is typically sigmoidal, with the linear portion of the curve coinciding with the sensor&#39;s range of greatest accuracy. As depicted, in the case of a TGase reaction, the ammonia concentration currently susceptible to measurement is 10 −4 -10 −3  M, although sensitivity is anticipated in the range of 10 −5 -10 −3  M. 
         [0064]      FIGS. 9 and 10  present the results of monitoring of the TGase reaction. Varying enzyme and substrate concentrations of the TGase reaction demonstrate the operation of the optical sensors. In particular, these results show that the higher the enzyme concentration or the higher the substrate concentration in the reaction, the faster ammonia is produced. 
       Further Applications 
       [0065]    As discussed above, various embodiments of the present invention may be constructed to directly detect the presence of particular ions. As illustrated in Table 1 below, it is known to the art that certain diseases affect particular ion channels in a cell. Accordingly, assays for those ions utilizing the present invention may furnish a diagnostic tool to determine the presence of particular diseases. Accordingly, the scope of the present invention should be understood to also include the application of the heretofore-described subject matter to measure the ions set forth in the following tables, as well as their application to diagnose the presence of the associated diseases also appearing in the following tables. 
         [0000]    
       
         
               
               
               
               
               
             
               
               
               
               
               
             
           
               
                   
               
               
                 Channel 
                 Gene 
                 Channel-forming unit/ligand 
                 OMIM 
                 Disease 
               
               
                   
               
             
             
               
                   
               
             
          
           
               
                 Cation channels: 
                   
                   
                   
                   
               
               
                 CHRNA1/ACHRA 
                 CHRNAI 
                 α, ACh 
                 100690 
                 Myasthenia congenita 
               
               
                 CHRNA4 
                 CHRNA4 
                 α, ACh 
                 118504 
                 Autosomal dominant nocturnal frontal lobe epilepsy 
               
               
                 CHRNB2 
                 CHRNB2 
                 β, ACh 
                 118507 
                 Autosomal dominant nocturnal frontal lobe epilepsy 
               
               
                 Polycystin-2 
                 PKD2 
                 α 
                 173910 
                 Autosomal dominant polycystic kidney disease (ADPKD) 
               
               
                 CNGA3 
                 CNGA3 
                 α, cGMP 
                 60053 
                 Achromatopsia 2 (color blindness) 
               
               
                 CNGB1 
                 CNGB1 
                 β, cGMP 
                 600724 
                 Autosomal recessive retinitis pigmentosa 
               
               
                 CNGB3 
                 CNGB3 
                 β, cGMP 
                 605080 
                 Achromatopsia 3 
               
               
                 Sodium channels: 
               
               
                 Na.1.1 
                 SCN1A 
                 α 
                 182389 
                 Generalized epilepsy with febrile seizures (GEFS+) 
               
               
                 Na.1.2 
                 SCN2A 
                 α 
                 182390 
                 Generalized epilepsy with febrile and afebrile seizures) 
               
               
                 Na.1.4 
                 SCN4A 
                 α 
                 603967 
                 Paramyotonia congenital, potassium aggressive myotonia, 
               
               
                   
                   
                   
                   
                 hyperkalemic periodic paralysis 
               
               
                 Na.1.5 
                 SCN5a 
                 α 
                 600163 
                 Long-QT syndrome, progressive familial heart block type 1, 
               
               
                   
                   
                   
                   
                 Brugada syndrome (idiopathic ventricular arrhythmia) 
               
               
                 SCNIB 
                 SCN1B 
                 β 
                 600235 
                 Generalized epilepsy with febrile seizures (GEFS+) 
               
               
                 ENACα 
                 SCNNIA 
                 α 
                 600228 
                 Pseudohypoaldosteronism type 1 (PHA1) 
               
               
                 ENaCβ 
                 SCNN1B 
                 β 
                 600760 
                 PHA1, Liddle syndrome (dominant hypertension 
               
               
                 ENaCγ 
                 SCNN1G 
                 γ 
                 600761 
                 PHA1, Liddle syndrome 
               
               
                 Potassium channels: 
               
               
                 K, 1.1. 
                 KCNA1 
                 α 
                 176260 
                 Episodic ataxia with myokymia 
               
               
                 KCNQI/K, LQT1 
                 KCNQ1 
                 α 
                 192500 
                 Autosomal dominant long-QT syndrome (Romano-Ward) 
               
               
                   
                   
                   
                   
                 Autosomal recessive long-QT syndrome with deafness (Jervell- 
               
               
                   
                   
                   
                   
                 Lange-Nielsen) 
               
               
                 KCNQ2 
                 KCNQ2 
                 α 
                 602235 
                 BFNC (epilepsy), also with myokymia 
               
               
                 KCNQ3 
                 KCNQ3 
                 α 
                 602232 
                 BFNC (epilepsy) 
               
               
                 KCNO4 
                 KCNQ4 
                 α 
                 603537 
                 DFNA2 (dominant hearing loss) 
               
               
                 HERG/KCNH2 
                 KCNH2 
                 α 
                 152427 
                 Long-QT syndrome 
               
               
                 Kir1.1/ROMK 
                 KCNJ1 
                 α 
                 600359 
                 Bartter syndrome (renal salt loss, hypokalemic alkalosis) 
               
               
                 Kir2.1/IRK/KCNJ2 
                 KCNJ2 
                 α 
                 600681 
                 Long-QT syndrome with dysmorphic features (Andersen 
               
               
                   
                   
                   
                   
                 syndrome) 
               
               
                 Kir6.2/KATATP ATP   
                 KCNJ11 
                 α 
                 600937 
                 Persistent hyperinsulinemic hypoglycemia of infancy (PHHI) 
               
               
                 SURI 
                 SURI 
                 β 
                 600509 
                 PHHI 
               
               
                 KCNE1/Mink/ISK 
                 KCNE1 
                 β 
                 176261 
                 Autosomal dominant long-QT syndrome (Romano-Ward) 
               
               
                   
                   
                   
                   
                 Autosomal recessive long-QT syndrome with deafness (Jervell- 
               
               
                   
                   
                   
                   
                 Lange-Nielson) 
               
               
                 KCNE2/MiRP1 
                 KCNE2 
                 β 
                 603796 
                 Long-QT syndrome 
               
               
                 KCNE3/MiRP2 
                 KCNE3 
                 β 
                 604433 
                 Periodic paralysis 
               
               
                 Calcium channels: 
               
               
                 Ca, 1.1 
                 CACNA1S 
                 α 
                 114208 
                 Hypokalemic periodic paralysis, malignant hyperthermia 
               
               
                 Ca, 1.4 
                 CACNA1F 
                 α 
                 300110 
                 X-linked congenital stationary night blindness 
               
               
                 Ca, 2.1 
                 CACNA1A 
                 α 
                 601011 
                 Familial hemiplegic migraine, episodic staxia, spinocerebella 
               
               
                   
                   
                   
                   
                 ataxia type 6 
               
               
                 RyRI 
                 RYR1 
                 α 
                 180901 
                 Malignant hyperthermia, central core disease 
               
               
                 RyR2 
                 RYR2 
                 α 
                 180902 
                 Catecholaminergic polymorphic ventricular tachycardia, 
               
               
                   
                   
                   
                   
                 arrhythmogenic right ventricular dysplasia type 2 
               
               
                 Chloride channels: 
               
               
                 CFTR 
                 ABCC7 
                 α 
                 602421 
                 Cystic fibrosis, congenital bilateral asplasia of vas deference 
               
               
                 CIC-1 
                 CLCN1 
                 α 
                 118425 
                 Autosomal recessive (Becker) or dominant (Thomsen myotonia 
               
               
                 CIC-5 
                 CLCN5 
                 α 
                 300008 
                 Dent&#39;s disease (X-linked proteinuria and kidney stones) 
               
               
                 CIC-7 
                 CLCN7 
                 α 
                 602727 
                 Osteopetrosis (recessive or dominant) 
               
               
                 CIC-Kb 
                 CLCNKB 
                 α 
                 602023 
                 Bartter syndrome type III 
               
               
                 Barttin 
                 BSND 
                 β 
                 606412 
                 Bartter syndrome type IV (associated with sensorineural 
               
               
                   
                   
                   
                   
                 deafness) 
               
               
                 GLRA1 
                 GLRA1 
                 α, glycine 
                 138491 
                 Hyperekplexin (startle desease) 
               
               
                 GABAα1 
                 GABRA1 
                 α GABA 
                 137160 
                 Juvenile myoclonus epilepsy 
               
               
                 GABAγ2 
                 GABRG2 
                 γ, GABA 
                 137164 
                 Epilepsy 
               
               
                 Gap junction channels: 
               
               
                 Cx26 
                 GJB2 
                   
                 121011 
                 DFNB3 (autosomal dominant hearing loss) 
               
               
                   
                   
                   
                   
                 DFNB1 (autosomal recessive hearing loss) 
               
               
                 Cx30 
                 GJB4 
                   
                 605425 
                 DFNA3 
               
               
                 Cx31 
                 GJB3 
                   
                 603324 
                 DFNA2 
               
               
                 Cx32 
                 GJB1 
                   
                 304040 
                 CMTX (X-linked Charcot-Mari-Tooth neuropathy) 
               
               
                 AChR α7 
                   
                   
                   
                 Inflammation 
               
               
                 ClC7 
                   
                   
                   
                 Osteoporosis 
               
               
                 Ether-a-go-go (eag, erg, elk) 
                   
                   
                   
                 Cancer 
               
               
                 Gardos channel 
                   
                   
                   
                 Sickle cell anemia 
               
               
                 P2X7 
                   
                   
                   
                 Immune disorders 
               
               
                 TRPC6 
                   
                   
                   
                 Asthma, COPD 
               
               
                 TRPM1 
                   
                   
                   
                 Melanoma 
               
               
                 TRPM2 
                   
                   
                   
                 Asthma 
               
               
                 TRPM4 
                   
                   
                   
                 Immune disorders 
               
               
                 TRPM7 
                   
                   
                   
                 Stroke 
               
               
                 TRPM8 
                   
                   
                   
                 Prostate cancer 
               
               
                 TRPV1 
                   
                   
                   
                 Urinary incontinence, pain 
               
               
                   
               
               
                 The third column classifies channel proteins into α, β, and γ subunits, where α subunits are always directly involved in pore formation, Several β subunits are only accessory (i.e., do not form pores), as is the case, for example, with SCN1B and barttin. Others (e.g. of ENaC and GABA receptors) participate in pore formation. For ligand-gated channels, the ligand is given. Note that GABA and glycine act from the extracellular side, whereas cGMP is an intracellular messenger. 
               
             
          
         
       
     
         [0000]    
       
         
               
               
               
               
               
             
           
               
                   
               
               
                 Gene 
                 Accession ID 
                 Gene Locus 
                   
                 Tissue Expression 
               
               
                   
               
             
             
               
                   
                   
                   
                 Sodium Channel Type/Disease 
                   
               
               
                 SCN1A 
                 GDB: 118870 
                 2q24 
                 SCN1, vg type 1, α-subunit (280 KDa) 
                 Brain 
               
               
                   
                 S71446 
               
               
                 SCN1B 
                 GDB: 127281 
                 19q13.1 
                 Hs.89634, vg type 1 β 1  subunit 
                 Brain, heart, skeletal 
               
               
                   
                 U12188-12104 
                   
                 (38 KDa) 
                 muscle 
               
               
                   
                 L16242, L10338 
               
               
                 SCN2A1 
                 GDB: 120367 
                 2q23 
                 SCN2A, HBSCI, vg type II, α 1 - 
                 Brain, peripheral nerve 
               
               
                   
                   
                   
                 subunit (280(KDa) 
               
               
                 SCN2A2 
                 CDB: 133727 
                 2q23-24.1 
                 HBSCH, vg type II, α 2 -subunit vg 
                 Brain 
               
               
                   
                   
                   
                 type II, β 2 -subunit (33 KDa) 
               
               
                 SCN2B 
                 GDB: 118871 
               
               
                   
                 AF019498 
               
               
                 SCN3A 
                 GDB: 132151 
                 2q24-31 
                 vg type III, α-subunit (280 kDa) 
                 Brain 
               
               
                   
                 S69887 
               
               
                 SCN4A 
                 GDB: 125181 
                 17q23.1-25.3 
                 SkM1, vg type IV α-subunit (260 kDa), 
                 Skeletal muscle 
               
               
                   
                 L04216-L04236 
                   
                 hyperkalemic periodic 
               
               
                   
                   
                   
                 paralysis, paramyotonia congentia, 
               
               
                   
                   
                   
                 potassturn-aggravated myotonia 
               
               
                 SCN4B 
                 GDB: 125182 
                 3q21 
                 vg type IV, β-subunit, 
                 Heart, fetal skeletal 
               
               
                 SCN5A 
                 GDB: 132152 
                   
                 SkM2, hH1, vg type V, α-subunit, 
                 muscle 
               
               
                   
                   
                   
                 long Q-T syndrome 3 
               
               
                 SCN6A 
                 GDB: 132153 
                 2q21-23 
                 Hs99945, vg type VI, α-subunit 
                 Heart, uterus, fetal and 
               
               
                   
                   
                   
                   
                 denervated skeletal 
               
               
                   
                   
                   
                   
                 muscle 
               
               
                 SCN7A 
                 GDB: 228137 
                 12q13 
                 vg type VII, α-subunit 
                 Brain, spinal cord 
               
               
                 SCN8A 
                 GDB: 631695 
                   
                 vg type VIII, α-subunit, motor 
               
               
                   
                   
                   
                 end-plate disease + ataxia in mice 
               
               
                 SCN9A 
                 GDB: 3750013 
                   
                 vg type IX, α-subunit 
                 Thyroid and adrenal 
               
               
                   
                   
                   
                 neuroendocrine type 
                 gland 
               
               
                 SCN10A 
                 GDB: 750014 
                 1pter-p36.3 
                 hPN3, vg type X 
                 Sensory neurons, dorsal 
               
               
                   
                   
                   
                   
                 root ganglia 
               
               
                 SCNN1A 
                 GDB: 366596 
                 12pt3 
                 SCNN1, nvg type 1 α-subunit of 
                 Kidney, lung colon 
               
               
                   
                 Z92978 
                   
                 ENaC 
               
               
                 SCNN1B 
                 GDB: 434471 
                 16p12.2-p12.1 
                 nvg 1 β-subunit, Liddle&#39;s syndrome, 
                 Kidney, lung colon 
               
               
                   
                   
                   
                 pseudohypoaldosterontsm I 
               
               
                 SCNN1D 
                 GDB: 6053678 
                 1p36.3-p36.2 
                 DnaCh, nvg 1 δ-subunit 
                 Kidney, lung, colon 
               
               
                 SCNN1G 
                 GDB: 568769 
                 16p122-p12.1 
                 nvg 1 γ-subunit, Liddle&#39;s syndrome, 
                 Kidney, lung, colon 
               
               
                   
                 X87160 
                   
                 pseudohypoaldosterontsm I 
               
               
                   
                 U53835-53853 
               
               
                   
                   
                   
                 Calcium Channel Type/Disease 
               
               
                 CACNA1A 
                 GDB: 126432 
                 19p13 
                 P/Q type α 1A- subunit, eqisodic ataxia 
                 Brain (cortex, bulbus, 
               
               
                 CACNL1A4 
                 Z80114-Z80155, 
                 19p13.1 
                 2, familial hemiplegic migraine, 
                 olfacorius, 
               
               
                   
                 X99697, U79666 
                   
                 spinocerebellar ataxia 6; tottering, 
                 hippocarnpus, 
               
               
                   
                   
                   
                 leaner, and rolling mice 
                 cerebellum, brain stem), 
               
               
                   
                   
                   
                   
                 motoneurons, kidney 
               
               
                 CACNA1B 
                 GDB: 580689 
                 9q34 
                 CACNN, N-type α 1A- subunit 
                 Central, peripheral 
               
               
                 CACNL1A5 
                 M94172, M94173 
                   
                   
                 nervous system 
               
               
                 CACNA1C 
                 GDB: 126094 
                 12p13 
                 CCHL1A1, L-type α 1A- subunit 
                 Heart, fibroblasts, lung, 
               
               
                 CACNL1A1 
                 L29636, L29634, 
                 12p13.3 
                   
                 smooth muscle (2 splice 
               
               
                   
                 L29629 
                   
                   
                 variants) 
               
               
                 CACNA1D 
                 GDB: 128872 
                 3p14.3 
                 CCHL1A2, L-type α 1D -subunit 
                 Brain, pancreas, 
               
               
                 CACNL1A2 
                   
                 3p21.3.2? 
                   
                 neuroendocrine 
               
               
                 CACNA1E 
                 GDB: 434408 
                 1q25-31 
                 R-type α 1C- subunit 
                 Brain, skeletal muscle 
               
               
                 CACNL1A6 
                   
                   
                   
                 (end plate) 
               
               
                 CACNA1F 
                 GDB: 6053864 
                 Xp11.23-11.22 
                 α 1F -Subunit 
                 Retina 
               
               
                 CACNIAG 
                 AF27964 
                 17q22 
                 T-type α 1G -subunit 
                 Brain 
               
               
                 CACNA1S 
                 GDB: 126431 
                 1q31-32 
                 L-type α 1B -subunit (5% 212, 95% 
                 Skeletal muscle (brain, 
               
               
                 CACNL1A8 
                 Z22672, L33798 
                   
                 190 kDa), malignant hyperthermia 
                 kidney) 
               
               
                   
                 U30666-U30707 
                   
                 5, hypokalernic periodic paralysis 
               
               
                 CACNA2 
                 GDB: 132010 
                 7q21-22 
                 CACNA2, CACNA2D1, α g8 -subunit 
                 α 2A ; skeletal muscle, 
               
               
                 CACNL2A 
                 Z28613, Z28609 
                   
                 (175 kDa), MHS3 
                 heart, brain, ileum; α 2B ; 
               
               
                   
                 Z28605, Z28602 
                   
                   
                 brain; α 2CVD ; aorta 
               
               
                   
                 Z28699, M76559 
               
               
                 CACNB1 
                 GDB: 132012 
                 17q21-22 
                 β 1 -Subunit (524 aa, 54 kDa) 
                 β 1 A/M; skeletal muscle 
               
               
                 CACNLB1 
                 GDB: 1073281 
                   
                   
                 β 1 B/C; brain, heart, 
               
               
                   
                 U86952-U86961 
                   
                   
                 spleen 
               
               
                   
                 M76560, L06111 
               
               
                   
                 GDB: 193328 
               
               
                 CACNB2 
                 GDB: 132014 
                 10p12 
                 MYSB, β 2 -subunit 
                 β 2 A/B/E; brain, heart, 
               
               
                 CACNLB2 
                 Q08289 
                   
                   
                 lung, aorta 
               
               
                 CACNB3 
                 GDB: 341023 
                 12q13 
                 β 2 -subunit (482 aa) 
                 Brain, heart, lung, spleen, 
               
               
                 CACNLB3 
                 L27584 
                   
                   
                 skeletal and smooth 
               
               
                   
                   
                   
                   
                 muscle, aorta, trachea, 
               
               
                   
                   
                   
                   
                 ovary, colon 
               
               
                 CACNB4 
                 GDB: 6028693 
                 2q22-23 
                 β 2 -subunit, lethargic mice 
                 Brain, kidney 
               
               
                 CACNG 
                 GDB: 132015 
                 17q24 
                 γ-Subunit (222 aa, 30 kDa) 
                 Skeletal muscle, lung 
               
               
                 CACNLG 
                 L07738 
               
               
                 CACNG2 
                   
                   
                 γ2-Subunit, stargazin, absence 
                 Brain 
               
               
                   
                   
                   
                 epilepsy stargazer, waggler mice 
               
               
                 RYR1 
                 GDB: 120359 
                 19q13.1 
                 Ryanodine receptor 1, Ca release 
                 Skeletal muscle, testis, 
               
               
                   
                   
                   
                 channel, 3 splice variants, 
                 brain, submaxillary and 
               
               
                   
                   
                   
                 malignant hyperthermia 1, central 
                 adrenal glands, spleen 
               
               
                   
                   
                   
                 core disease 
               
               
                 RYR2 
                 GDB: 125278 
                 1pter-qter 
                 RYR2, calcium release channel 
                 Heart, smooth muscle 
               
               
                   
                   
                 1q42.1-43 
               
               
                 RYR3 
                 GDB: 138451 
                 15q14 
                 RYR3, calcium release channel 
                 Brain, neonatal skeletal 
               
               
                   
                   
                 15q14-15 
                   
                 muscle, adult diaphragm 
               
               
                   
                   
                   
                 Potassium Channel Type/Disease 
               
               
                 KCNA1 
                 GDB: 127903 
                 12p13 
                 RBK1, HUK1, MBK1, AEMK, 
                 Brain, nerve, heart, 
               
               
                   
                 LO2750 
                   
                 Kv1.1, Shaker homolog 1, Shaker, 
                 skeletal muscle, retina, 
               
               
                   
                   
                   
                 episodic ataxia 1 (with myokymia) 
                 pancreatic islet 
               
               
                 KCNA1B 
                   
                 3q26.1 
                 Kvβ1.1, Kvβ1.3 (splice product), 
               
               
                   
                   
                   
                 β-subunit 
               
               
                 KCNA2 
                 GDB: 128062 
                 12pter-qter 
                 HK4, Kv1.2, Shaker homolog 2 
                 Brain, nerve, heart, 
               
               
                   
                 X17622 
                   
                   
                 pancreatic islet 
               
               
                 KCNA2B 
                   
                 1p36.3 
                 Kvβ1.2, β-subunit 
               
               
                 KCNA8 
                 GDB: 128079 
                 1p13.3 
                 Hs.1750, MK3, HLK3, HPCN3, 
                 Skeletal muscle, 
               
               
                   
                 L23499 
                   
                 Kv1.3, Shaker homolog 3 
                 lymphocytes (brain, 
               
               
                   
                   
                   
                   
                 lung, thymus, spleen) 
               
               
                 KCNA4 
                 GDB: 126730 
                 11p14 
                 Hs.89647, Hs.1854, HK1, HPCN2, 
                 Brain, nerve, heart, fetal 
               
               
                   
                 M60450 
                   
                 Kv1.4, Shaker homolog 4 
                 skeletal muscle, 
               
               
                   
                 M55514 
                   
                   
                 pancreatic islet 
               
               
                 KCNA4L 
                 GDB: 386059 
                 11q14 
                 Shaker homolog type 4-like 
               
               
                 KCNA5 
                 GDB: 127904 
                 12p13.3-13.2 
                 Hs.89509, HK2, HPCNI, Kv1.5 
                 Brain, heart, kidney, lung, 
               
               
                   
                 M83254 
                 12p13 
                 Shaker homolog 5 
                 skeletal muscle, 
               
               
                   
                 M60451 
                 12p13.33-12.31 
                   
                 pancreatic islet 
               
               
                 KCNA6 
                 GDB: 128080 
                 12p13 
                 HBK2, Kv1.6, Shaker homolog 6 
                 Brain, pancreatic islet 
               
               
                   
                 X17622 
               
               
                 KCNA7 
                 GDB: 127905 
                 19q13.3 
                 HAK6, Kv1.7 Shaker homolog 7 
               
               
                 KCNA8 
                   
                   
                 see KCNQ1 
               
               
                 KCNA9 
                   
                   
                 see KCNQ1 
               
               
                 KCNA10 
                 GDB: 5885822 
                   
                 Shaker homolog type 10, cGMP 
               
               
                   
                   
                   
                 activated 
               
               
                 KCNB1 
                 GDB: 128081 
                 20q13.2 
                 Kv2.1, Shab homolog 1 
                 Brain, heart, kidney, 
               
               
                   
                   
                   
                   
                 retina, skeletal muscle 
               
               
                 KCNB2 
                   
                   
                 Kv2.2, Shab homolog 2 
                 Brain, heart, retina 
               
               
                 KCNC1 
                 GDB: 128082 
                 11p15.1 
                 Kv3.1, Shaw homolog 1 
                 Brain, skeletal muscle, 
               
               
                   
                 S56770 
                   
                   
                 spleen, lymphocytes 
               
               
                   
                 M96747 
               
               
                 KCNC2 
                 GDB: 127906 
                 19q13.3-13.4 
                 Kv3.2, Shaw homolog 2 
                 Brain 
               
               
                 KCNC3 
                 GDB: 127907 
                 19q13.3 
                 Kv3.3, Shaw homolog 3 
                 Brain, liver 
               
               
                 KCNC4 
                 GDB: 127908 
                 1p21 
                 Kv3.4, HKSHIIIC, Shaw homolog 4 
                 Brain, skeletal muscle 
               
               
                 KCND1 
                 GDB: 128083 
                   
                 Kv4.1, Shal hormolog 1 
                 Brain 
               
               
                 KCND2 
                 GDB: 134771 
                   
                 RK5, Kv4.2, Shal homolog 2 
                 Brain, heart, aorta 
               
               
                 KCND3 
                 GDB: 134772 
                   
                 Kv4.3, KSHIVB, Shal homolog 3 
               
               
                 KCNE1 
                 GDB: 127909 
                 21q22.1-22.2 
                 MinK, ISK, vg Isk homolog 1 (129 
                 Kidney, submandibular 
               
               
                   
                   
                   
                 aa), long Q-T syndrome 5 
                 gland, uterus, heart, 
               
               
                   
                   
                   
                   
                 cochlea, retina 
               
               
                 KCNMA1 
                 GDB: 386031 
                 10pter-qter 
                 SLO, Hs.62679, α-subunit member 
                 Fetal skeletal muscle 
               
               
                   
                 U09383-4 
                 7q32.1 
                 1, α-subunit of maxiK or BK 
               
               
                   
                 U02632 
                   
                 channel 
               
               
                 KCNMB1 
                 GDB: 6099615 
                 5q34 
                 hSLO-β, β-subunit member 1 (191 
                 Smooth, fetal skeletal 
               
               
                   
                 U42600 
                   
                 aa), β-subunit of max1K or BK 
                 muscle, brain 
               
               
                   
                   
                   
                 channel 
                 (hippocampus, corpus 
               
               
                   
                   
                   
                   
                 callosum) 
               
               
                 KCNN1 
                 U69883 
                   
                 SK(Ca)1, small-conductance Ca- 
                 Brain, heart 
               
               
                   
                   
                   
                 activated K channel, apamin- 
               
               
                   
                   
                   
                 insensitive 
               
               
                 KCNN2 
                   
                   
                 SK(Ca)2, apamin sensitive 
                 Brain, adrenal gland 
               
               
                 KCNN3 
                 Y08263 
                 1q? 
                 SK(Ca)3, small-conductance Ca- 
                 Brain, heart, (human 
               
               
                   
                 AA285078 
                   
                 activated K channel, intermediate 
                 embryonic) skeletal 
               
               
                   
                   
                   
                 apamin sensitivity 
                 muscle, liver 
               
               
                 KCNN4 
                 AF022150 
                 19q13.2 
                 IK1, intermediate-conductance 
                 T lymphocytes, colon, 
               
               
                   
                 AF022797 
                   
                 Ca-activated K channel, KCa4, 
                 smooth muscles, 
               
               
                   
                 AF033021 
                   
                 SK4, Gantos channel 
                 prostata, red blood cells, 
               
               
                   
                 AF000972 
                   
                   
                 neurons 
               
               
                 KCNQ1 
                 GDB: 741244 
                 11p15.5 
                 KCNA9, (KV)LQT1, KQT-like 
                 Heart, cochlea, kidney, 
               
               
                   
                 U40990 
                   
                 subfamily member 1, long Q-T 
                 lung, placenta, colon 
               
               
                   
                   
                   
                 syndrome 1 
               
               
                 KCNQ2 
                 GDB: 9787229, 
                 20q13.3 
                 KQT-like subfamily member 2 (872 
                 Brain 
               
               
                   
                 Y15065, 
                   
                 aa) 
               
               
                   
                 AF033348 
               
               
                 KCNQ3 
                 GDB: 9787230 
                 8q24.22-24.3 
                 KQT-like subfamily member 3 (825 
                 Brain 
               
               
                   
                 AF033347 
                   
                 aa) 
               
               
                 HERG 
                 GDB: 407638 
                 7q35-36 
                 HERG, similar to ether-a-go go 
                 Brain, heart 
               
               
                   
                   
                   
                 (eag), IKr, long Q-T syndrome 2 
               
               
                 KCNJ1 
                 GDB: 204206 
                 11q24 
                 ROMK1, Kirl.1, Hs.463, 
                 Kidney, pancreatic islets 
               
               
                   
                 U65406, U12541 
                   
                 Bartter/hyperprostaglandin 
               
               
                   
                   
                   
                 E syndrome 
               
               
                 KCNJ2 
                 GDB: 278964 
                 17pter-qter 
                 IRK1, Kir2.1, Hs.1547 
                 Muscle, neural tissue, 
               
               
                   
                 U12507 
                   
                   
                 heart 
               
               
                 KCNJ3 
                 GDB: 278325 
                 2q24.1 
                 GIRK1, Kir3.1 
                 Heart, cerebellum 
               
               
                   
                 U50964 
               
               
                 KCNJ4 
                 GDB: 374080 
                 22q13.1 
                 HIR, HIRK1, HIRK2, Kir2.3 
                 Heart, skeletal muscle, 
               
               
                   
                 Z97056 
                   
                   
                 brain 
               
               
                 KCNJ5 
                 GDB: 547948 
                 11q24 
                 CIR, KATP1, GIRK4, Kir3.4 
                 Heart, pancreas 
               
               
                 KCNJ6 
                 GDB: 547949 
                 21q22.1 
                 KCNJ7, GIRK2, KATP2, BIR1, 
                 Cerebellum, pancreatic 
               
               
                   
                 U24660 
                   
                 Kir3.2, ataxia, weaver mice 
                 islet 
               
               
                 KCNJ8 
                 GDB: 633096 
                 12p11.23] 
                 Kir6.1, uKATP, ubiquitious K ATP   
                 Brain, heart, skeletal, 
               
               
                   
                   
                   
                 α-subunit 
                 smooth muscle, others 
               
               
                 KCNJ10 
                 GDB: 3750203 
                 1q22-23] 
                 Kir1.2, Kir4.1 
                 Glia 
               
               
                 KCNJ11 
                 GDB: 7009893 
                 [11p15.1] 
                 Kir6.2, BIR, K(ATP) α-subunit, 
                 Pancreatic islets 
               
               
                   
                   
                   
                 hyperinsulinemic hypoglycemia 
               
               
                 KCNJ12 
                 GDB: 4583927 
                 [17p11.1] 
                 Kir2.2 
               
               
                 KCNJ15 
                 GDB: 6275865 
                 [21q22.2] 
                 Kir4.2 
               
               
                 KCNJN1 
                 GDB: 6108062 
                 [ ] 
                 Kir2.2v, subfamily inhibitor 1 
               
               
                 SUR1 
                 GDB: 591970 
                 [11p15.1] 
                 SUR(1), sulfonylurea receptor, 
                 Pancreatic islets 
               
               
                   
                   
                   
                 K(ATP) β-subunit, 
               
               
                   
                   
                   
                 hyperinsulinemic hypoglycemia 
               
               
                 SUR2 
                   
                 12p12.1] 
                 SUR2, SUR2A, B, sulfonylurea 
                 2A: heart, 2B: brain, liver, 
               
               
                   
                   
                   
                 receptor 2 (1545-aa), β-subunit of 
                 skeletal, smooth muscle, 
               
               
                   
                   
                   
                 K(ATP) 
                 urinary bladder 
               
               
                 KCNK1 
                 GDB: 6045446 
                 1q42-43 
                 DPK, TWIK1 
                 Kidney 
               
               
                 KCNK2 
                   
                 1q41 
                 TREK1 
                 Brain 
               
               
                 KCNK3 
                 GDB: 9773281 
                 2p23 
                 TASK 
                 Kidney 
               
               
                   
               
             
          
         
       
     
         [0000]    
       
         
               
               
               
             
           
               
                   
               
               
                 Therapeutic Target 
                 Enzyme Family 
                 Assay 
               
               
                   
               
             
             
               
                 Alzheimer&#39;s 
                 CMGC 
                 ERK2 (P42mapk) 
               
               
                 Alzheimer&#39;s 
                 Phospholipase 
                 PLA2 
               
               
                 Alzheimer&#39;s 
                 Cyclooxygenases 
                 COX2 
               
               
                 Alzheimer&#39;s 
                 CaMK 
                 MARK1 
               
               
                 Alzheimer&#39;s 
                 CaMK 
                 MARK2 
               
               
                 Alzheimer&#39;s 
                 AGC 
                 PKCalpha 
               
               
                 Alzheimer&#39;s 
                 AGC 
                 PKCgamma 
               
               
                 Alzheimer&#39;s 
                 AGC 
                 PKCgamma 
               
               
                 Alzheimer&#39;s 
                 Cysteine proteases 
                 caspase-3 
               
               
                 Alzheimer&#39;s 
                 Cysteine proteases 
                 caspase-6 
               
               
                 Alzheimer&#39;s 
                 Aspartic proteases 
                 BACE-1 (beta-secretase) 
               
               
                 Alzheimer&#39;s 
                 Aspartic proteases 
                 cathepsin D 
               
               
                 Alzheimer&#39;s 
                 Aspartic proteases 
                 cathepsin F 
               
               
                 Alzheimer&#39;s 
                 Metalloproteases 
                 ACE 
               
               
                 Alzheimer&#39;s 
                 Metalloproteases 
                 ACE 
               
               
                 Alzheimer&#39;s 
                 Metalloproteases 
                 TACE 
               
               
                 Alzheimer&#39;s 
                 NO synthases 
                 constitutive NOS (cerebellar) 
               
               
                 Alzheimer&#39;s 
                 Monoamine &amp; neurotransmitter 
                 acetylcholinesterase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Alzheimer&#39;s 
                 Monoamine &amp; neurotransmitter 
                 COMT (catechol-O-methyl transferase) 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Alzheimer&#39;s 
                 Monoamine &amp; neurotransmitter 
                 MAO-A 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Alzheimer&#39;s 
                 Monoamine &amp; neurotransmitter 
                 MAO-B 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Alzheimer&#39;s 
                 Monoamine &amp; neurotransmitter 
                 tyrosine hydroxylase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Alzheimer&#39;s 
                 Phospholipase C 
                 PLC 
               
               
                 Alzheimer&#39;s 
                 Miscellaneous enzymes 
                 xanthine oxidase/superoxide O2-scavenging 
               
               
                 Dependence/Addiction 
                 AGC 
                 PKA 
               
               
                 Dependence/Addiction 
                 AGC 
                 PKCalpha 
               
               
                 Dependence/Addiction 
                 AGC 
                 PKCbeta 1 
               
               
                 Dependence/Addiction 
                 AGC 
                 PKCbeta 2 
               
               
                 Dependence/Addiction 
                 AGC 
                 PKCdelta 
               
               
                 Dependence/Addiction 
                 Monoamine &amp; neurotransmitter 
                 GABA transaminase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Dependence/Addiction 
                 Cyclases 
                 adenylyl cyclase (stimulated) 
               
               
                 Dependence/Addiction 
                 Phospholipase C 
                 PLC 
               
               
                 Dependence/Addiction 
                 ATPase 
                 ATPase (Na+/K+) 
               
               
                 Inflammation/Arthritis/Allergy 
                 RTK 
                 EGFR kinase 
               
               
                 Inflammation/Arthritis/Allergy 
                 RTK 
                 FLT-1 kinase (VEGFR1) 
               
               
                 Inflammation/Arthritis/Allergy 
                 RTK 
                 KDR kinase (VEGFR2) 
               
               
                 Inflammation/Arthritis/Allergy 
                 CTK 
                 Fyn kinase 
               
               
                 Inflammation/Arthritis/Allergy 
                 CTK 
                 HCK 
               
               
                 Inflammation/Arthritis/Allergy 
                 CTK 
                 Lck kinase 
               
               
                 Inflammation/Arthritis/Allergy 
                 CTK 
                 Lyn kinase 
               
               
                 Inflammation/Arthritis/Allergy 
                 CTK 
                 ZAP70 kinase 
               
               
                 Inflammation/Arthritis/Allergy 
                 CMGC 
                 ERK2 (P42mapk) 
               
               
                 Inflammation/Arthritis/Allergy 
                 CMGC 
                 JNK 1 
               
               
                 Inflammation/Arthritis/Allergy 
                 CMGC 
                 JNK 2 
               
               
                 Inflammation/Arthritis/Allergy 
                 CMGC 
                 P38alpha kinase 
               
               
                 Inflammation/Arthritis/Allergy 
                 Phospholipase 
                 PLA2 
               
               
                 Inflammation/Arthritis/Allergy 
                 Cyclooxygenases 
                 COX1 
               
               
                 Inflammation/Arthritis/Allergy 
                 Cyclooxygenases 
                 COX2 
               
               
                 Inflammation/Arthritis/Allergy 
                 TXA2 synthetase 
                 TXA2 synthetase 
               
               
                 Inflammation/Arthritis/Allergy 
                 CaMK 
                 MAPKAPK2 
               
               
                 Inflammation/Arthritis/Allergy 
                 AGC 
                 PKA 
               
               
                 Inflammation/Arthritis/Allergy 
                 Lipoxygenases 
                 12-lipoxygenase 
               
               
                 Inflammation/Arthritis/Allergy 
                 Lipoxygenases 
                 15-lipoxygenase 
               
               
                 Inflammation/Arthritis/Allergy 
                 Serine proteases 
                 elastase 
               
               
                 Inflammation/Arthritis/Allergy 
                 Serine proteases 
                 cathepsin G 
               
               
                 Inflammation/Arthritis/Allergy 
                 Serine proteases 
                 kallikrein 
               
               
                 Inflammation/Arthritis/Allergy 
                 Serine proteases 
                 tryptase 
               
               
                 Inflammation/Arthritis/Allergy 
                 Cysteine proteases 
                 caspase-1 
               
               
                 Inflammation/Arthritis/Allergy 
                 Cysteine proteases 
                 caspase-4 
               
               
                 Inflammation/Arthritis/Allergy 
                 Cysteine proteases 
                 caspase-5 
               
               
                 Inflammation/Arthritis/Allergy 
                 Cysteine proteases 
                 cathepsin B 
               
               
                 Inflammation/Arthritis/Allergy 
                 Cysteine proteases 
                 cathepsin X 
               
               
                 Inflammation/Arthritis/Allergy 
                 Aspartic proteases 
                 cathepsin F 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 MMP-1 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 MMP-2 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 MMP-3 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 MMP-7 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 MMP-8 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 MMP-9 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 MMP-13 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 MT1-MMP (MMP-14) 
               
               
                 Inflammation/Arthritis/Allergy 
                 Metalloproteases 
                 TACE 
               
               
                 Inflammation/Arthritis/Allergy 
                 Phosphatases 
                 phosphatase CD45 
               
               
                 Inflammation/Arthritis/Allergy 
                 Phosphodiesterases 
                 PDE2 
               
               
                 Inflammation/Arthritis/Allergy 
                 Phosphodiesterases 
                 PDE4 
               
               
                 Inflammation/Arthritis/Allergy 
                 Phosphodiesterases 
                 acid sphingomyelinase 
               
               
                 Inflammation/Arthritis/Allergy 
                 Monoamine &amp; neurotransmitter 
                 HNMT (histamine N-methyltransferase) 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Inflammation/Arthritis/Allergy 
                 Miscellaneous enzymes 
                 myeloperoxidase 
               
               
                 Inflammation/Arthritis/Allergy 
                 Miscellaneous enzymes 
                 xanthine oxidase/superoxide O2-scavenging 
               
               
                 Neuroprotection 
                 RTK 
                 TRKB 
               
               
                 Neuroprotection 
                 CMGC 
                 CDK5 
               
               
                 Neuroprotection 
                 CMGC 
                 DYRK1a 
               
               
                 Neuroprotection 
                 CMGC 
                 ERK1 
               
               
                 Neuroprotection 
                 CMGC 
                 ERK2 (P42mapk) 
               
               
                 Neuroprotection 
                 MCGC 
                 JCK 3 
               
               
                 Neuroprotection 
                 Cyclooxygenases 
                 COX1 
               
               
                 Neuroprotection 
                 Cyclooxygenases 
                 COX2 
               
               
                 Neuroprotection 
                 CaMK 
                 CaMK2alpha 
               
               
                 Neuroprotection 
                 AGC 
                 PKA 
               
               
                 Neuroprotection 
                 Cysteine proteases 
                 caspase-3 
               
               
                 Neuroprotection 
                 Phosphodiesterases 
                 PDE1 
               
               
                 Neuroprotection 
                 Phosphodiesterases 
                 PDE6 
               
               
                 Neuroprotection 
                 NO synthases 
                 constitutive NOS (endothelial) 
               
               
                 Neuroprotection 
                 NO synthases 
                 constitutive NOS (cerebellar) 
               
               
                 Neuroprotection 
                 Monoamine &amp; neurotransmitter 
                 acetylcholinesterase 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Neuroprotection 
                 Monoamine &amp; neurotransmitter 
                 COMT (catechol-O-methyl transferase) 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Neuroprotection 
                 Monoamine &amp; neurotransmitter 
                 GABA transaminase 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Neuroprotection 
                 Monoamine &amp; neurotransmitter 
                 HNMT (histamine N-methyltransferase) 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Neuroprotection 
                 Monoamine &amp; neurotransmitter 
                 MAO-A 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Neuroprotection 
                 Monoamine &amp; neurotransmitter 
                 MAO-A 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Neuroprotection 
                 Monoamine &amp; neurotransmitter 
                 PNMT (phenylethanoiamine-N-methyl 
               
               
                   
                 syntheses &amp; metabolism 
                 transferase) 
               
               
                 Neuroprotection 
                 Monoamine &amp; neurotransmitter 
                 tyrosine hydroxylase 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Neuroprotection 
                 Cyclases 
                 guanylyl cyclase (basal) 
               
               
                 Neuroprotection 
                 Cyclases 
                 guanylyl cyclase (stimulated) 
               
               
                 Neuroprotection 
                 ATPase 
                 ATPase (Na+/K+) 
               
               
                 Neuroprotection 
                 Miscellaneous enzymes 
                 xanthine oxidase/superoxide O2-scavenging 
               
               
                 Parkinson 
                 CMGC 
                 JNK 1 
               
               
                 Parkinson 
                 Phospholipase 
                 PLA2 
               
               
                 Parkinson 
                 Cyclooxygenases 
                 COX2 
               
               
                 Parkinson 
                 Cysteine proteases 
                 caspase-3 
               
               
                 Parkinson 
                 NO synthases 
                 constitutive NOS (cerebellar) 
               
               
                 Parkinson 
                 Monoamine &amp; neurotransmitter 
                 acetylcholinesterase 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Parkinson 
                 Monoamine &amp; neurotransmitter 
                 COMT (catechol-O-methyl transferase 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Parkinson 
                 Monoamine &amp; neurotransmitter 
                 MAO-A 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Parkinson 
                 Monoamine &amp; neurotransmitter 
                 MAO-B 
               
               
                   
                 syntheses &amp; metabolism 
               
               
                 Cancer 
                 RTK 
                 Axl kinase 
               
               
                 Cancer 
                 RTK 
                 c-kit kinase 
               
               
                 Cancer 
                 RTK 
                 c-kit kinase 
               
               
                 Cancer 
                 RTK 
                 EGFR kinase 
               
               
                 Cancer 
                 RTK 
                 EphA1 kinase 
               
               
                 Cancer 
                 RTK 
                 EphA3 kinase 
               
               
                 Cancer 
                 RTK 
                 EphA4 kinase 
               
               
                 Cancer 
                 RTK 
                 EphB2 kinase 
               
               
                 Cancer 
                 RTK 
                 FGFR1 kinase 
               
               
                 Cancer 
                 RTK 
                 FGFR2 kinase 
               
               
                 Cancer 
                 RTK 
                 FGFR3 kinase 
               
               
                 Cancer 
                 RTK 
                 FGFR4 kinase 
               
               
                 Cancer 
                 RTK 
                 FLT-1 kinase (VEGFR1) 
               
               
                 Cancer 
                 RTK 
                 FLT-3 kinase 
               
               
                 Cancer 
                 RTK 
                 FLT-4 kinase (VEGFR3) 
               
               
                 Cancer 
                 RTK 
                 Fms/CSFR kinase 
               
               
                 Cancer 
                 RTK 
                 HER2/ErbB2 kinase 
               
               
                 Cancer 
                 RTK 
                 HER4/ErbB4 kinase 
               
               
                 Cancer 
                 RTK 
                 KDR kinase (VEGFR2) 
               
               
                 Cancer 
                 RTK 
                 PDGFRalpha kinase 
               
               
                 Cancer 
                 RTK 
                 PDGFRbeta kinase 
               
               
                 Cancer 
                 RTK 
                 Ret kinase 
               
               
                 Cancer 
                 RTK 
                 TIE2 kinase 
               
               
                 Cancer 
                 RTK 
                 TRKA 
               
               
                 Cancer 
                 CTK 
                 Abl kinase 
               
               
                 Cancer 
                 CTK 
                 BLK 
               
               
                 Cancer 
                 CTK 
                 BMX (Bk) kinase 
               
               
                 Cancer 
                 CTK 
                 BRK 
               
               
                 Cancer 
                 CTK 
                 BTK 
               
               
                 Cancer 
                 CTK 
                 CSK 
               
               
                 Cancer 
                 CTK 
                 FAK 
               
               
                 Cancer 
                 CTK 
                 Fes kinase 
               
               
                 Cancer 
                 CTK 
                 Fyn kinase 
               
               
                 Cancer 
                 CTK 
                 JAK2 
               
               
                 Cancer 
                 CTK 
                 JAK3 
               
               
                 Cancer 
                 CTK 
                 Lck kinase 
               
               
                 Cancer 
                 CTK 
                 PYK2 
               
               
                 Cancer 
                 CTK 
                 Src kinase 
               
               
                 Cancer 
                 CTK 
                 Syk 
               
               
                 Cancer 
                 CTK 
                 Yes kinase 
               
               
                 Cancer 
                 CMGC 
                 CDC2/CDK1 (cycB) 
               
               
                 Cancer 
                 CMGC 
                 CDK2 (cycE) 
               
               
                 Cancer 
                 CMGC 
                 CDK4 (cycD1) 
               
               
                 Cancer 
                 CMGC 
                 CDK5 
               
               
                 Cancer 
                 CMGC 
                 CK2 (casein kinase 2) 
               
               
                 Cancer 
                 CMGC 
                 DYRKla 
               
               
                 Cancer 
                 CMGC 
                 ERK1 
               
               
                 Cancer 
                 CMGC 
                 ERK2 (P42mapk) 
               
               
                 Cancer 
                 CMGC 
                 HIPK2 
               
               
                 Cancer 
                 CMGC 
                 IKKalpha 
               
               
                 Cancer 
                 CMGC 
                 IKKbeta 
               
               
                 Cancer 
                 CMGC 
                 JNK 1 
               
               
                 Cancer 
                 CMGC 
                 JNK 2 
               
               
                 Cancer 
                 CMGC 
                 NEK1 
               
               
                 Cancer 
                 CMGC 
                 NEK2 
               
               
                 Cancer 
                 CMGC 
                 NEK4 
               
               
                 Cancer 
                 CMGC 
                 p38alpha kinase 
               
               
                 Cancer 
                 CMGC 
                 p38beta 2 kinase (SAPK2b2) 
               
               
                 Cancer 
                 CMGC 
                 p38delta kinase 
               
               
                 Cancer 
                 CMGC 
                 p38ganuna kinase 
               
               
                 Cancer 
                 Cyclooxygenases 
                 COX2 
               
               
                 Cancer 
                 CaMK 
                 CaMK1delta 
               
               
                 Cancer 
                 CaMK 
                 CaMK 
               
               
                 Cancer 
                 CaMK 
                 CHK1 
               
               
                 Cancer 
                 CaMK 
                 CHK2 
               
               
                 Cancer 
                 CaMK 
                 DAPK1 
               
               
                 Cancer 
                 CaMK 
                 DAPK2 
               
               
                 Cancer 
                 CaMK 
                 MAPKAPK2 
               
               
                 Cancer 
                 CaMK 
                 MAPKAPK3 
               
               
                 Cancer 
                 CaMK 
                 MAPKAPK5 (PRAK0 
               
               
                 Cancer 
                 CaMK 
                 MAARK1 
               
               
                 Cancer 
                 CaMK 
                 MARK2 
               
               
                 Cancer 
                 CaMK 
                 MARK4 
               
               
                 Cancer 
                 CaMK 
                 Pim 1 kinase 
               
               
                 Cancer 
                 CaMK 
                 Pirn2 kinase 
               
               
                 Cancer 
                 AGC 
                 Akt1/PKBalpha 
               
               
                 Cancer 
                 AGC 
                 Akt2/PKBbeta 
               
               
                 Cancer 
                 AGC 
                 Akt3/PKBgamma 
               
               
                 Cancer 
                 AGC 
                 AurA/Aur2 kinase 
               
               
                 Cancer 
                 AGC 
                 AurB/Aur1 kinase 
               
               
                 Cancer 
                 AGC 
                 AurC/Aur3 kinase 
               
               
                 Cancer 
                 AGC 
                 P70S6Ke 
               
               
                 Cancer 
                 AGC 
                 PDK1 
               
               
                 Cancer 
                 AGC 
                 PKA 
               
               
                 Cancer 
                 AGC 
                 PKCalpha 
               
               
                 Cancer 
                 AGC 
                 PKCbeta 1 
               
               
                 Cancer 
                 AGC 
                 PKCbeta 2 
               
               
                 Cancer 
                 AGC 
                 PKCdelta 
               
               
                 Cancer 
                 AGC 
                 PKCgamma 
               
               
                 Cancer 
                 AGC 
                 PKG2 
               
               
                 Cancer 
                 AGC 
                 ROCK1 
               
               
                 Cancer 
                 AGC 
                 ROCK2 
               
               
                 Cancer 
                 AGC 
                 RSK2 
               
               
                 Cancer 
                 AGC 
                 SGK1 
               
               
                 Cancer 
                 Lipoxygenases 
                 12-lipoxygenase 
               
               
                 Cancer 
                 TKL 
                 RAF-1 kinase 
               
               
                 Cancer 
                 STE 
                 MEK1/MAP2K1 
               
               
                 Cancer 
                 STE 
                 MKK4/JNK1 
               
               
                 Cancer 
                 STE 
                 MKK6 
               
               
                 Cancer 
                 STE 
                 PAK1 
               
               
                 Cancer 
                 STE 
                 PAK2 
               
               
                 Cancer 
                 Serine proteases 
                 elastase 
               
               
                 Cancer 
                 Serine proteases 
                 cathepsin G 
               
               
                 Cancer 
                 Cysteine proteases 
                 caspase-2 
               
               
                 Cancer 
                 Cysteine proteases 
                 caspase-3 
               
               
                 Cancer 
                 Cysteine proteases 
                 caspase-8 
               
               
                 Cancer 
                 Cysteine proteases 
                 caspase-9 
               
               
                 Cancer 
                 Cysteine proteases 
                 cathepin B 
               
               
                 Cancer 
                 Cysteine proteases 
                 cathepsin H 
               
               
                 Cancer 
                 Cysteine proteases 
                 cathepsin L 
               
               
                 Cancer 
                 Cysteine proteases 
                 cathepsin X 
               
               
                 Cancer 
                 Aspartic proteases 
                 cathepsin D 
               
               
                 Cancer 
                 Aspartic proteases 
                 cathepsin E 
               
               
                 Cancer 
                 Metalloproteases 
                 MMP-1 
               
               
                 Cancer 
                 Metalloproteases 
                 MMP-2 
               
               
                 Cancer 
                 Metalloproteases 
                 MMP-3 
               
               
                 Cancer 
                 Metalloproteases 
                 MMP-7 
               
               
                 Cancer 
                 Metalloproteases 
                 MMP-8 
               
               
                 Cancer 
                 Metalloproteases 
                 MMP-9 
               
               
                 Cancer 
                 Metalloproteases 
                 MMP-12 
               
               
                 Cancer 
                 Metalloproteases 
                 MMP-13 
               
               
                 Cancer 
                 Metalloproteases 
                 MT1-MMP (MMP-14) 
               
               
                 Cancer 
                 Metalloproteases 
                 TACE 
               
               
                 Cancer&#39; 
                 Metalloproteases 
                 MMP-1 
               
               
                 Cancer 
                 Phosphatases 
                 phosphatase 1B 
               
               
                 Cancer 
                 Phosphatases 
                 phosphatase 2B 
               
               
                 Cancer 
                 Phosphodiesterases 
                 PDE2 
               
               
                 Cancer 
                 Phosphodiesterases 
                 PDE4 
               
               
                 Cancer 
                 Phosphodiesterases 
                 PDE5 
               
               
                 Cancer 
                 Phosphodiesterases 
                 acid spingomyelinase 
               
               
                 Cancer 
                 NO synthases 
                 constitutive NOS (endothelial) 
               
               
                 Cancer 
                 NO synthases 
                 constitutive NOS (cerebellar) 
               
               
                 Cancer 
                 Cyclases 
                 adenylyl cyclase (basal) 
               
               
                 Cancer 
                 Cyclases 
                 adenylyl cyclase (stimulated) 
               
               
                 Cancer 
                 Phospholipase C 
                 PLC 
               
               
                 Cancer 
                 Miscellaneous enzymes 
                 myeloperoxidase 
               
               
                 Cancer 
                 Miscellaneous enzymes 
                 xanthine oxidase/superoxide O2-scavenging 
               
               
                 Diabetes 
                 RTK 
                 Ax1 kinase 
               
               
                 Diabetes 
                 RTK 
                 EGFR kinase 
               
               
                 Diabetes 
                 RTK 
                 IGFIR kinase 
               
               
                 Diabetes 
                 CMGC 
                 ERK2 (P42mapk) 
               
               
                 Diabetes 
                 CMGC 
                 Jnk1 
               
               
                 Diabetes 
                 Cyclooxygenases 
                 COX2 
               
               
                 Diabetes 
                 TXA2 synthetase 
                 TXA2 synthetase 
               
               
                 Diabetes 
                 CaMK 
                 AMPKalpha 
               
               
                 Diabetes 
                 AGC 
                 Akt1/PKBalpha 
               
               
                 Diabetes 
                 AGC 
                 Akt2/PKBbeta 
               
               
                 Diabetes 
                 AGC 
                 Akt3/PKBgamma 
               
               
                 Diabetes 
                 AGC 
                 PDK1 
               
               
                 Diabetes 
                 AGC 
                 PKA 
               
               
                 Diabetes 
                 AGC 
                 PKCalpha 
               
               
                 Diabetes 
                 AGC 
                 PKCbeta 1 
               
               
                 Diabetes 
                 AGC 
                 PKCbeta 2 
               
               
                 Diabetes 
                 AGC 
                 PKCgamma 
               
               
                 Diabetes 
                 AGC 
                 SGK2 
               
               
                 Diabetes 
                 Metalloproteases 
                 ACE 
               
               
                 Diabetes 
                 Metalloproteases 
                 MMP-1 
               
               
                 Diabetes 
                 Metalloproteases 
                 MMP-2 
               
               
                 Diabetes 
                 Metalloproteases 
                 MMP-3 
               
               
                 Diabetes 
                 Metalloproteases 
                 MMP-7 
               
               
                 Diabetes 
                 Metalloproteases 
                 MMP-8 
               
               
                 Diabetes 
                 Metalloproteases 
                 MMP-9 
               
               
                 Diabetes 
                 Metalloproteases 
                 MT1-MMP (MMP-14) 
               
               
                 Diabetes 
                 Metalloproteases 
                 TACE 
               
               
                 Diabetes 
                 Phosphodiesterases 
                 PDE3 
               
               
                 Diabetes 
                 Phosphodiesterases 
                 PDE4 
               
               
                 Diabetes 
                 Phosphodiesterases 
                 PDE5 
               
               
                 Diabetes 
                 NO synthases 
                 constitutive NOS (endothelial) 
               
               
                 Diabetes 
                 Monoamine &amp; neurotransmitter 
                 acetylcholinesterase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Diabetes 
                 Monoamine &amp; neurotransmitter 
                 GABA transaminase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Diabetes 
                 Monoamine &amp; neurotransmitter 
                 MAO-B 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Diabetes 
                 Cyclases 
                 adenylyl cyclase (basal) 
               
               
                 Diabetes 
                 Miscellaneous enzymes 
                 acetylCoA synthetase 
               
               
                 Diabetes 
                 Miscellaneous enzymes 
                 HMG-CoA reductase 
               
               
                 Diabetes 
                 Miscellaneous enzymes 
                 xanthine oxidase/superoxide O2-scavenging 
               
               
                 Metabolic Diseases 
                 Cyclooxygenases 
                 COX2 
               
               
                 Metabolic Diseases 
                 AGC 
                 PKA 
               
               
                 Metabolic Diseases 
                 Metalloproteases 
                 ACE 
               
               
                 Metabolic Diseases 
                 Phosphodiesterases 
                 PDE3 
               
               
                 Metabolic Diseases 
                 Phosphodiesterases 
                 PDE4 
               
               
                 Metabolic Diseases 
                 NO synthases 
                 constitutive NOS (endothelial) 
               
               
                 Metabolic Diseases 
                 Miscellaneous enzymes 
                 acetylCoA synthetase 
               
               
                 Metabolic Diseases 
                 Miscellaneous enzymes 
                 HMG-CoA reductase 
               
               
                 Metabolic Diseases 
                 Miscellaneous enzymes 
                 xanthine oxidase/superoxide 02-scavenging 
               
               
                 Obesity 
                 CTK 
                 PYK2 
               
               
                 Obesity 
                 CMGC 
                 JNK1 
               
               
                 Obesity 
                 CaMK 
                 AMPJakoga 
               
               
                 Obesity 
                 AGC 
                 PKA 
               
               
                 Obesity 
                 Metalloproteases 
                 ACE 
               
               
                 Obesity 
                 Metalloproteases 
                 ACE 
               
               
                 Obesity 
                 Phosphatases 
                 phosphatase 1B 
               
               
                 Obesity 
                 Phosphodiesterases 
                 PDE2 
               
               
                 Obesity 
                 Phosphodiesterases 
                 PDE3 
               
               
                 Obesity 
                 Monoamine &amp; neurotransmitter 
                 acetylcholinesterase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Obesity 
                 ATPase 
                 ATPase (Na+/K+) 
               
               
                 Obesity 
                 Miscellaneous enzymes 
                 HMG-CoA reductase 
               
               
                 Reproduction 
                 Phospholipase 
                 PLA2 
               
               
                 Reproduction 
                 Cyclooxygenases 
                 COX1 
               
               
                 Reproduction 
                 Cyclooxygenases 
                 COX2 
               
               
                 Reproduction 
                 Phosphodiesterases 
                 PDE5 
               
               
                 Reproduction 
                 NO synthases 
                 constitutive NOS (endothelial) 
               
               
                 Reproduction 
                 Cyclases 
                 guanylyl cyclase (stimulated) 
               
               
                 Cystic Fibrosis 
                 Phospholipase 
                 PLA2 
               
               
                 Cystic Fibrosis 
                 TXA2 synthetase 
                 TXA2 synthetase 
               
               
                 Cystic Fibrosis 
                 AGC 
                 PKA 
               
               
                 Cystic Fibrosis 
                 AGC 
                 PKCbeta 1 
               
               
                 Cystic Fibrosis 
                 AGC 
                 PKCbeta 2 
               
               
                 Cystic Fibrosis 
                 Serine proteases 
                 elastase 
               
               
                 Cystic Fibrosis 
                 Serine proteases 
                 cathepsin G 
               
               
                 Cystic Fibrosis 
                 Metalloproteases 
                 MMP-2 
               
               
                 Cystic Fibrosis 
                 Phosphodiesterases 
                 PDE3 
               
               
                 Cystic Fibrosis 
                 Phosphodiesterases 
                 PDE5 
               
               
                 Cystic Fibrosis 
                 Cyclases 
                 adenylyl cyclase (stimulated) 
               
               
                 Cystic Fibrosis 
                 Phospholipase C 
                 PLC 
               
               
                 Cystic Fibrosis 
                 Miscellaneous enzymes 
                 myeloperoxidase 
               
               
                 Immunosuppression Profile 
                 RTK 
                 EGFR kinase 
               
               
                 Immunosuppression Profile 
                 CTK 
                 JAK3 
               
               
                 Immunosuppression Profile 
                 CMGC 
                 ERK2 (P42mapk) 
               
               
                 Immunosuppression Profile 
                 Cyclooxygenases 
                 COX1 
               
               
                 Immunosuppression Profile 
                 Cyclooxygenases 
                 COX2 
               
               
                 Immunosuppression Profile 
                 Serine proteases 
                 elastase 
               
               
                 Immunosuppression Profile 
                 Serine proteases 
                 cathepsin G 
               
               
                 Immunosuppression Profile 
                 Serine proteases 
                 tryptase 
               
               
                 Immunosuppression Profile 
                 Cysteine proteases 
                 cathepsin B 
               
               
                 Immunosuppression Profile 
                 Metalloproteases 
                 ECE-1 
               
               
                 Immunosuppression Profile 
                 Metalloproteases 
                 ECE-1 
               
               
                 Immunosuppression Profile 
                 Metalloproteases 
                 MMP-1 
               
               
                 Immunosuppression Profile 
                 Metalloproteases 
                 MMP-2 
               
               
                 Immunosuppression Profile 
                 Metalloproteases 
                 MMP-9 
               
               
                 Immunosuppression Profile 
                 Phosphatases 
                 phosphatase CD45 
               
               
                 Immunosuppression Profile 
                 Phosphodiesterases 
                 PDE4 
               
               
                 Immunosuppression Profile 
                 Phosphodiesterases 
                 acid spingomyelinase 
               
               
                 Immunosuppression Profile 
                 Cyclases 
                 adenylyl cyclase (basal) 
               
               
                 Immunosuppression Profile 
                 Cyclases 
                 adenylyl cyclase (stimulated) 
               
               
                 Migraine 
                 Cyclooxygenases 
                 COX2 
               
               
                 Migraine 
                 NO synthases 
                 constitutive NOS (cerebellar) 
               
               
                 Migraine 
                 Monoamine &amp; neurotransmitter 
                 GABA transaminase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Migraine 
                 Cyclases 
                 guanylyl cyclase (stimulated) 
               
               
                 Pain 
                 CMGC 
                 ERK2 (42mapk) 
               
               
                 Pain 
                 Phospholipase 
                 PLA2 
               
               
                 Pain 
                 Cyclooxygenases 
                 COX1 
               
               
                 Pain 
                 Cyclooxygenases 
                 COX2 
               
               
                 Pain 
                 AGC 
                 PKA 
               
               
                 Pain 
                 Serine proteases 
                 elastase 
               
               
                 Pain 
                 Metalloproteases 
                 MMP-1 
               
               
                 Pain 
                 Metalloproteases 
                 MMP-2 
               
               
                 Pain 
                 Metalloproteases 
                 MMP-3 
               
               
                 Pain 
                 Metalloproteases 
                 MMP-7 
               
               
                 Pain 
                 Phosphodiesterases 
                 PDE4 
               
               
                 Pain 
                 NO synthases 
                 constitutive NOS (endothelial) 
               
               
                 Pain 
                 NO synthases 
                 constitutive NOS (cerebellar) 
               
               
                 Pain 
                 Monoamine &amp; neurotransmitter 
                 GABA transaminase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Pain 
                 Monoamine &amp; neurotransmitter 
                 MAO-A 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Pain 
                 Monoamine &amp; neurotransmitter 
                 MAO-B 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Pain 
                 Monoamine &amp; neurotransmitter 
                 tyrosine hydroxylase 
               
               
                   
                 synthesis &amp; metabolism 
               
               
                 Pain 
                 Miscellaneous enzymes 
                 xanthine oxidase/superoxide 02-scavenging 
               
               
                   
               
             
          
         
       
     
         [0066]    Thus, according to various features, the systems, methods and devices of the invention provide, without limitation, a novel approach for using optodes and cells for measurement, including drug discovery and the monitoring of enzymatic reactions; the coating of an ion-selective PVC cocktail with TEFLON to prevent salt interference; an approach for using the optodes for extracellular measurement in a 96-well plate format, and for HERG and ammonia screening. The invention also provides a microdevice for combining optodes with cell measurements, sensor inserts for a well plate (such as PVC-coated glass disks), and bio-compatible coatings for optodes. Commercial applications for the invention include, without limitation, drug discovery, clinical monitoring, toxicity studies, chemical and biological detection, and the like. 
         [0067]    Additional applications and protocols for analyzing ion-channel activity are described in U.S. Pat. No. 6,969,449, the entirety of which is incorporated by reference. Such protocols can be readily adapted for use with the optical ion sensors and cell assay systems described herein. 
         [0068]    It will therefore be seen that the foregoing represents a highly advantageous approach to measurements using optical biosensor arrays. The terms and expressions employed herein are used as terms of description and not of limitation and there is no intention, in the use of such terms and expressions, of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the invention claimed.