1. A device for monitoring cell-substrate impedance, comprising:
a) a nonconducting substrate;
b) two or more electrode arrays fabricated on said substrate, wherein each of said two or more electrode arrays comprises two electrode structures;
c) two or more fluid containers on said substrate, wherein each of said two or more arrays is associated with one of said two or more fluid containers; and
d) at least two connection pads, each of which is located on an edge of said substrate;
wherein for each of said two or more electrode arrays, each of said two electrode structures comprises multiple electrode elements and the first of said two electrode structures of each of said at least two electrode arrays is connected to one of said at least two connection pads, and the second of said two electrode structures of each of said at least two electrode arrays is connected to another of said at least two connection pads;
further wherein at least two of said two or more electrode arrays share one common connection pad;
further wherein each electrode array has an approximately uniform electrode resistance distribution across the entire array; and
further wherein said substrate has a surface suitable for cell attachment or growth; wherein said cell attachment or growth on said substrate can result in a detectable change in impedance between or among said electrode structures within each electrode array.
2. A system for monitoring cell-substrate impedance, comprising:
a) one or more devices of claim 1, wherein said at least two fluid containers are two or more wells, wherein at least two of said two or more wells of said device comprise an electrode array at the bottom of the well, wherein said electrode array is individually addressed, further wherein said device can be used to measure differences in impedance values that relate to cell behavior;
b) an impedance analyzer;
c) a device station comprising electronic circuitry that can engage said device and selectively connect said two or more electrode arrays of said device to said impedance analyzer; and
d) a software program that can control said device station and record and analyze data obtained from said impedance analyzer.
3-57. (canceled)
58. A method of performing an assay to evaluate the effect of one or more test compounds on cells, comprising:
providing the cell-substrate impedance monitoring system of claim 2;
adding cells to two or more wells of said system;
adding different concentrations of a test compound to said two or more of said three or more wells comprising cells to form twor more compound ls comprising different concentrations of said test compound;
providing at least one additional well as a control well to which cells are added that does not receive test compound,
monitoring impedance from said two or more test compound wells and said at least one control well to obtain impedance measurements from said two or more compound wells and said at least one control well at at least one time point before the addition of different concentrations of a test compound to said compound wells and at three or more time points after adding said different concentrations of a test compound to said three or more compound wells;
analyzing said impedance measurements from said two or more compound wells and said at least one control well to evaluate the effect of said test compound on cells.
59. The method of claim 58, wherein said monitoring impedance comprises monitoring impedance at regular or irregular time intervals for an assay period of from minutes to days.
60. The method of claim 58, wherein said analyzing comprises plotting said impedance measurements from said three or more compound wells that comprise different concentrations of a compound versus time to obtain three or more compound concentration impedance curves and plotting said impedance measurements for said at least one control well versus time to obtain at least one control impedance curve.
61. The method of claim 60, further comprising comparing at least one of said three or more compound concentration impedance curves with said control impedance curve to evaluate an effect of said test compound on cells.
62. The method of claim 58, wherein said analyzing comprises deriving a cell index, a normalized cell index, or a delta cell index from said impedance measurements from said three or more compound wells that comprise different concentrations of a compound and from said at least one control well.
63. The method of claim 62, further comprising plotting said cell index, said normalized cell index, or said delta cell index of said at least three compound wells that comprise different concentrations of a compound versus time to provide three or more compound concentration CI curves and plotting said cell index, said normalized cell index, or said delta cell index of said at least one control well versus time to provide at least one control CI curve.
64. The method of claim 63, further comprising comparing at least one of said compound concentration CI with said at least one control CI curve to evaluate the effect of said test compound on cells.
65. The method of claim 64, wherein said assay is an assay for cell viability, cell adhesion, apoptosis, cell differentiation, cell proliferation, cytotoxicity, cell morphology change, cell cycle change, IgE-mediated cell activation or stimulation, receptor-ligand binding, cell number quantification, cell quality control, time-dependent cell cycling monitoring, detection or quantification of neutralizing antibodies, specific T-cell mediated cytotoxic effect, viral or bacterial toxin mediated cell pathologic changes, or cell death; and
wherein evaluating the effect of said test compound on cells comprises determining a change in cell status in response to at least one of said different concentrations of said test compound, wherein a change in cell status comprises at least one of: cell attachment or adhesion status, cell growth or proliferation status; the number of viable cells or dead cells; cytoskeleton change or re-organization; or the number of cells going through apoptosis or necrosis.
66. The method of claim 62, further plotting said cell index, said normalized cell index, or said delta change index versus concentration for at least one time point of said assay to generate one or more dose response curves for one or more time points at which impedance is measured.
67. The method of claim 166, further comprising using said one or more dose-response curves to calculate a time-dependent IC5, IC10, IC20, IC30, IC40, IC50, IC60, IC70, IC80, IC90, or IC95 at one or more time points for said compound.
68. The method of claim 67, comprising using said one or more dose-response curves to calculate one or more time-dependent IC50 for said compound for one or more time points.
69. The method of claim 62, further comprising deriving a cell change index or a second order derivative of a cell index from said impedance measurements from at least one of said three or more compound wells that comprise different concentrations of a test compound and from said at least one control well.
70. The method of claim 69, wherein said cell change index or a second order derivative of a cell index is a cell change index (CCI).
71. The method of claim 70, further comprising determining the CCI at a given time point for at least one of said three or more compound wells that comprise different concentrations of a test compound and said at least one control well as being either: approximately equal to 0.7, much greater than 0.7, greater than zero and less than 0.7, approximately equal to zero, less than zero, or much less than zero;
wherein a CCI approximately equal to 0.7 indicates log rate growth, a CCI much greater than 0.7 indicates faster than log rate growth, a CCI greater than zero and less than 0.7 indicates slower than log rate growth, a CCI approximately equal to zero indicates no growth (a constant cell index), a CCI less than zero indicates cells are detaching from the substrate, and a CCI much less than zero indicates cell are detaching rapidly from the substrate.
72. The method of claim 70,
further comprising plotting said cell change index from said at least one of said three or more compound wells that comprise different concentrations of a compound and from said at least one control well versus time to obtain at least one test compound CCI curve and at least one control CCI curve; and
further comprising comparing said at least one test compound CCI curve with said at least one control CCI curve.
73. The method of claim 72, wherein evaluating the effect of said test compound on cells comprises determining a change in cell status in response to at least one of said different concentrations of said test compound, wherein a change in cell status comprises at least one of: cell attachment or adhesion status, cell growth or proliferation status; the number of viable cells or dead cells; cytoskeleton change or re-organization; or the number of cells going through apoptosis or necrosis.
74. The method of claim 73, wherein said assay is an assay for cell viability, cell adhesion, apoptosis, cell differentiation, cell proliferation, cytotoxicity, cell morphology change, cell cycle changes, IgE-mediated cell activation or stimulation, receptor-ligand binding, cell number quantification, cell quality control, time-dependent cell cycling monitoring, detection or quantification of neutralizing antibodies, specific T-cell mediated cytotoxic effect, viral or bacterial toxin mediated cell pathologic changes, or cell death.
75-84. (canceled)
The claims below are in addition to those above.
All refrences to claim(s) which appear below refer to the numbering after this setence.
1. A seed of the corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859.
2. A population of seed of the corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859.
3. An essentially homogeneous population of corn seeds consisting essentially of the inbred corn seed of claim 1.
4. A population of corn seeds consisting essentially of the inbred corn seed of claim 1, and essentially free from hybrid seed.
5. A corn plant of corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859.
6. A plant part of the corn plant of claim 5.
7. The plant part of claim 6, further defined as pollen, an ovule or a cell.
8. An essentially homogeneous population of corn plants of the corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859.
9. A corn plant expressing all of the physiological and morphological characteristics of the corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859.
10. The corn plant of claim 5, further comprising a nuclear or cytoplasmic gene conferring male sterility.
11. A tissue culture of cells of a plant of corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859.
12. The tissue culture of claim 11, wherein the cells comprise cells derived from embryos, immature embryos, meristematic cells, immature tassels, microspores, pollen, leaves, anthers, roots, root tips, silk, flowers, kernels, ears, cobs, husks, or stalks.
13. The tissue culture of claim 12, wherein the cells comprise protoplasts or callus cells.
14. A corn plant regenerated from the tissue culture of claim 11, wherein the corn plant expresses all of the physiological and morphological characteristics of the corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859.
15. A process of producing corn seed, comprising crossing a first parent corn plant with a second parent corn plant, wherein one or both of the first or the second parent corn plant is a plant of the corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859, wherein seed is allowed to form.
16. The process of claim 15, further defined as a process of producing hybrid corn seed, comprising crossing a first inbred corn plant with a second, distinct inbred corn plant, wherein the first or second inbred corn plant is a plant of the corn variety I294213, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859.
17. The process of claim 16, wherein crossing comprises the steps of:
(a) planting the seeds of first and second inbred corn plants;
(b) cultivating the seeds of said first and second inbred corn plants into plants that bear flowers;
(c) preventing self pollination of at least one of the first or second inbred corn plant;
(d) allowing cross-pollination to occur between the first and second inbred corn plants; and
(e) harvesting seeds on at least one of the first or second inbred corn plants, said seeds resulting from said cross-pollination.
18. The corn plant of claim 5, further comprising a transgene introduced by genetic transformation.
19. A method of producing an inbred corn plant derived from the corn variety I294213, the method comprising the steps of:
(a) preparing a progeny plant derived from corn variety I294213 by crossing a plant of the corn variety I294213 with a second corn plant, wherein a sample of the seed of the corn variety I294213 was deposited under ATCC Accession No. PTA-7859;
(b) crossing the progeny plant with itself or a second plant to produce a seed of a progeny plant of a subsequent generation;
(c) growing a progeny plant of a subsequent generation from said seed and crossing the progeny plant of a subsequent generation with itself or a second plant; and
(d) repeating steps (b) and (c) for an additional 2\u201310 generations to produce an inbred corn plant derived from the corn variety I294213.
20. A method of producing a conversion of the corn variety I294213 to express at least one new trait, the method comprising the steps of:
(a) crossing a first corn plant having a first diploid genome comprising a plurality of paired chromosomes comprising a plurality of mappable genetic loci with a pair of alleles at each locus, and further comprising a conversion that confers at least one new trait, with a second plant of the corn variety I294213, a sample of the seed of the corn variety I294213 having been deposited under ATCC Accession No. PTA-7859, the plant of the corn variety I294213 having a second diploid genome comprising a plurality of paired chromosomes comprising a plurality of mappable genetic loci with a pair of alleles at each locus, to produce seed comprising a diploid genome having a plurality of paired chromosomes comprising a plurality of mappable genetic loci with a pair of alleles at each locus, wherein one of the alleles is contributed by the first corn plant and the other is contributed by the plant of the corn variety I294213, said genome further comprising the conversion that confers the new trait;
(b) harvesting and planting the seed thereby produced to produce at least one progeny plant of the first filial generation, said progeny plant comprising a diploid genome comprising the conversion;
(c) crossing said progeny plant with a plant of the corn variety I294213 to produce seed of a subsequent filial generation, the seed comprising a diploid genome having a plurality of paired chromosomes comprising a plurality of mappable genetic loci with a pair of alleles at each locus, wherein one of the alleles is contributed by the progeny plant and the other is contributed by the plant of the corn variety I294213, and further comprising the conversion that confers the new trait;
(d) growing at least one progeny plant of the subsequent filial generation from the seed produced in step (c), said progeny plant comprising a genome comprising the conversion that confers the new trait;
(e) repeating steps (c) and (d) for at least three additional generations to produce a converted plant of the corn variety I294213 wherein the plant comprises a diploid genome having a plurality of paired chromosomes comprising a plurality of mappable genetic loci with a pair of alleles at each locus, wherein both alleles at substantially all of the loci consist essentially of the allele found at the same locus in corn variety I294213, the genome further comprising the conversion that confers the new trait; and
(f) harvesting the seed of the converted plant.
21. The method of claim 20, wherein the conversion was conferred by a transgene.
22. The method of claim 20, wherein the conversion is selected from the group consisting of a dominant allele and a recessive allele.
23. The method of claim 20, wherein the new trait is selected from the group consisting of herbicide tolerance; insect resistance; resistance to bacterial, fungal, nematode or viral disease; waxy starch; male sterility and restoration of male fertility.
24. A converted plant of the corn variety I294213 produced by the method of claim 20.