Device and method for measuring impedance in organotypic tissues

Abstract

The present invention refers to a device for measuring impedance in organotypic tissue comprising at least one recording chamber with a liquid permeable membrane supporting the organotypic tissue, at least one bottom electrode and at least one top electrode, wherein the liquid permeable membrane divides the recording chamber into a top chamber and a bottom chamber, wherein at least the bottom chamber contains culture medium for the organotypic tissue, and the bottom electrode(s) is/are located in the bottom chamber and the top electrode(s) is/are located in the top chamber, and wherein the organotypic tissue is located between the bottom electrode(s) and the top electrode(s). The present invention also refers to the use of the device according to the present invention for measuring impedance in organotypic tissue. Furthermore, the present invention relates to a method for analyzing the effect of test compounds on pathological and non-pathological organotypic tissue by measuring the impedance of the organotypic tissue, wherein the organotypic tissue is cultured in a culture medium during the time of the analysis and the impedance of the organotypic tissue is measured at least once before and at least once after treating the organotypic tissue with the test compound or the impedance of the organotypic tissue treated with the test compound is compared to a non-treated organotypic tissue, wherein the impedance is measured using at least one electrode at each of two opposing sides of the organotypic tissue, and the electrodes are contacted with the culture medium or the tissue during measuring the impedance.

1 . A device for measuring impedance in organotypic tissue comprising at least one recording chamber with a liquid permeable membrane supporting the organotypic tissue, at least one bottom electrode and at least one top electrode, said device characterized in that:
 the liquid permeable membrane divides the recording chamber into a top chamber and a bottom chamber, wherein   the bottom electrode(s) is/are located in the bottom chamber and the top electrode(s) is/are located in the top chamber, and wherein   the organotypic tissue is located between the bottom electrode(s) and the top electrode(s).   
     
     
         2 . The device according to  claim 1 , characterized in that the top electrode(s) in the top chamber is/are movable in at least two directions so it/they can be contacted with or removed from the organotypic tissue or the culture medium. 
     
     
         3 . The device according to  claim 1 , characterized in that the electrodes are interconnected by at least one multiplexer and an impedance/gain-phase analyzer system. 
     
     
         4 . The device according to  claim 1 , characterized in that the liquid permeable membrane extends through all the recording chambers. 
     
     
         5 . The device according to  claim 1 , characterized in that the bottom electrode(s) are supported on a substrate at the bottom of the recording chamber. 
     
     
         6 . The device according to  claim 1 , characterized in that the electrodes are individually addressable. 
     
     
         7 . The device according to  claim 1 , characterized in that the recording chamber is connected to an automated liquid handling system. 
     
     
         8 . The device according to  claim 7 , characterized in that the liquid handling system can provide a humidified atmosphere in the recording chamber or the liquid handling system is placed in an CO 2  incubator. 
     
     
         9 . The device according to  claim 1 , further comprising a bottomless multiwell frame with 1-1000 wells, wherein each well defines one recording chamber. 
     
     
         10 . The device according to  claim 1 , characterized in that the device comprises a lid which contains an implemented multiplexer board. 
     
     
         11 . The device according to  claim 1 , characterized in that the bottom electrodes are connected to connection pads via conductors, wherein the conductors are isolated from each other by a passivation layer comprising silicon nitrite, silicon oxide, polyimide, or viscose polymers. 
     
     
         12 . The device according to  claim 1 , characterized in that the number of bottom electrodes in the recording chamber is 4 to 256. 
     
     
         13 . The device according to  claim 1 , characterized in that the liquid permeable membrane comprises an opening for handling liquid. 
     
     
         14 . (canceled) 
     
     
         15 . (canceled) 
     
     
         16 . (canceled) 
     
     
         17 . (canceled) 
     
     
         18 . A method for analyzing the effect of test compounds on organotypic tissue by measuring the impedance of the organotypic tissue, comprising
 (i) culturing the organotypic tissue in a culture medium during the time of the analysis;   (ii) contacting the organotypic tissue with the test compound;   (iii) optionally measuring the impedance of the organotypic tissue prior to step (iii);   (iv) measuring the impedance of the organotypic tissue at least once after step (iii), wherein
 the impedance is measured using at least one electrode at each of two opposing sides of the organotypic tissue, and 
 the electrodes are contacted with the culture medium or the tissue during measuring the impedance. 
   
     
     
         19 . The method according to  claim 18 , characterized in that the time span between the first impedance measurement prior to contacting the organotypic tissue with the test compound and the last measurement of the organotypic tissue treated with the test compound is at least 1 week. 
     
     
         20 . The method according to  claim 18 , characterized in that the impedance measurements of the organotypic tissue are performed continuously. 
     
     
         21 . The method according to  claim 18 , characterized in that the organotypic tissue represents a slice culture or explant culture derived from any mammal, vertebrate and invertebrate species of embryonic, neonatal, postnatal, and adult individuals. 
     
     
         22 . The method according to  claim 18 , characterized in that the organotypic tissue is transformed into pathological tissue by by i) introducing mutant genes by means of bacterial or viral vectors, ii) knock out genes related to specific disease, or iii) treatment with chemical agents. 
     
     
         23 . The method according to  claim 22 , characterized in that the impedance of the organotypic tissue is measured prior to and after transformation of the non-pathological organotypic tissue into pathological tissue as well as prior to and after contacting the organotypic tissue with the test compound. 
     
     
         24 . The method according to  claim 18 , characterized in that the organotypic tissue used for measuring impedance is a non-pathological tissue and is treated with test compounds to test the toxicity of the test compounds. 
     
     
         25 . The method according to  claim 18 , characterized in that the recording is performed by transient indirect electrode contact impedance recording (TIECIR), permanent indirect electrode contact impedance recording (PIECIR) or transient direct electrode contact impedance recording (TDECIR). 
     
     
         26 . The method according to  claim 18 , characterized in that the organotypic tissue is obtained from transgenic animals carrying mutation inducing properties of neurogenerative diseases selected from the group consisting of Alzheimer's disease, Parkinson's disease, Huntigton's disease, amyothrophic lateral sclerosis, prion diseases, Pick's disease, fronto-temporal dementia, progressive nuclear palsy, corticobasal degeneration, multiple system atrophy, mild-cognitive impairment, ischemic stroke, multiple sclerosis, motor neuron diseases, nerve injury and repair, age related macular degenerations, rod-cone dystrophy, cone-rod dystrophy, retinitis pigmentosa, glaucoma, and other retina associated degenerations. 
     
     
         27 . The method according to  claim 18 , characterized in that the measuring of the impedance is carried out by recording of frequency dependent impedance magnitudes and phase angles before and after application of test compounds at multiple frequencies (1 Hz-100 MHz). 
     
     
         28 . The method according to  claim 18 , characterized in that the impedance is measured by using a device comprising at least one recording chamber with a liquid permeable membrane supporting the organotypic tissue, at least one bottom electrode and at least one top electrode, said device characterized in that:
 the liquid permeable membrane divides the recording chamber into top and a bottom chamber, wherein   the bottom electrode(s) is/are located in the bottom chamber and the top electrode(s) is/are located in the top chamber, and wherein   the organotypic tissue is located between the bottom electrode(s) and the top electrode(s).