Name: sensing of barrier tissue disruption with an organic electrochemical transistor
Uploaded: 2014-02-10T13:00:00
Description: Watch this Scientific Journal Video about Sensing of Barrier Tissue Disruption with an Organic Electrochemical Transistor at JoVE.com

The authors of this paper do not have any competing financial interests.

1. PEDOT:PSS Solution Preparation
<ol>
	<li>To 50 ml&#160;of PEDOT:PSS, add ethylene glycol (increases conductivity) in a volume ratio of 1:4 (ethylene glycol to PEDOT:PSS), 0.5 &#956;l/ml&#160;of Dodecylbenzenesulfonic acid (DBSA) as a surfactant, and 10 mg/ml 3-glycidoxypropyltrimethoxysilane (GOPS) as a cross-linker to promote adhesion of the conducting polymer to the glass slide.</li>
</ol>
2. OECT Fabrication (Figure 3)
<ol>
	<li>Define thermally evaporated gold source and drain contacts ...

<ol>
	<li>Farquhar, M. G., Palade, G. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Junctional+complexes+in+various+epithelia.">Junctional complexes in various epithelia.</a> J. Cell Biol. 17, 375-412 (1963).</li><li>Gaillard, J. L., Finlay, B. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+cell+polarization+and+differentiation+on+entry+of+Listeria+monocytogenes+into+the+enterocyte-like+Caco-2+cell+line.">Effect of cell polarization and differentiation on entry of Listeria monocytogenes into the enterocyte-like Caco-2 cell line.</a> Infect. Immun. 64, 1299-1308 (1996).</li><li>Anderson, J. M., Balda, M. S., Fanning, A. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+structure+and+regulation+of+tight+junctions.">The structure and regulation of tight junctions.</a> Curr. Opin. Cell Biol. 5, 772-778 (1993).</li><li>Guttman, J. A., Finlay, B. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tight+junctions+as+targets+of+infectious+agents.">Tight junctions as targets of infectious agents.</a> Biochim. Biophys. Acta. 1788, 832-841 (2009).</li><li>Anderson, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Molecular+structure+of+tight+junctions+and+their+role+in+epithelial+transport.">Molecular structure of tight junctions and their role in epithelial transport.</a> News. Physiol. Sci. 16, 126-130 (2001).</li><li>Anderson, J. M., Van Itallie, C. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Tight+junctions:+Closing+in+on+the+seal.">Tight junctions: Closing in on the seal.</a> Curr. Biol. 9, (1999).</li><li>Ma, T. Y., Boivin, M. A., Ye, D., Pedram, A., Said, H. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mechanism+of+TNF-{alpha}+modulation+of+Caco-2+intestinal+epithelial+tight+junction+barrier:+role+of+myosin+light-chain+kinase+protein+expression.">Mechanism of TNF-{alpha} modulation of Caco-2 intestinal epithelial tight junction barrier: role of myosin light-chain kinase protein expression.</a> Am. J. Physiol. Gastrointest. Liver Physiol. 288, 422-430 (2005).</li><li>Schulzke, J. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epithelial+tight+junctions+in+intestinal+inflammation.">Epithelial tight junctions in intestinal inflammation.</a> Ann. N.Y. Acad. Sci. 1165, 294-300 (2009).</li><li>Fisher, S. J., Swaan, P. W., Eddington, N. D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+ethanol+metabolite+acetaldehyde+increases+paracellular+drug+permeability+in+vitro+and+oral+bioavailability+in+vivo.">The ethanol metabolite acetaldehyde increases paracellular drug permeability in vitro and oral bioavailability in vivo.</a> The J. Pharmacol. Exp. Therap. 332, 326-333 (2010).</li><li>Ma, T. Y., Nguyen, D., Bui, V., Nguyen, H., Hoa, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ethanol+modulation+of+intestinal+epithelial+tight+junction+barrier.">Ethanol modulation of intestinal epithelial tight junction barrier.</a> Am. J. Physiol. 276, 965-974 (1999).</li><li>Nemeth, E., Halasz, A., Barath, A., Domokos, M., Galfi, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+hydrogen+peroxide+on+interleukin-8+synthesis+and+death+of+Caco-2+cells.">Effect of hydrogen peroxide on interleukin-8 synthesis and death of Caco-2 cells.</a> Immunopharmacol. Immunotoxicol. 29, 297-310 (2007).</li><li>Vogelmann, R., Amieva, M. R., Falkow, S., Nelson, W. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Breaking+into+the+epithelial+apical-junctional+complex--news+from+pathogen+hackers.">Breaking into the epithelial apical-junctional complex--news from pathogen hackers.</a> Curr. Opin. Cell Biol. 16, 86-93 (2004).</li><li>Nusrat, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Clostridium+difficile+toxins+disrupt+epithelial+barrier+function+by+altering+membrane+microdomain+localization+of+tight+junction+proteins.">Clostridium difficile toxins disrupt epithelial barrier function by altering membrane microdomain localization of tight junction proteins.</a> Infect. Immun. 69, 1329-1336 (2001).</li><li>Obert, G., Peiffer, I., Servin, A. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rotavirus-induced+structural+and+functional+alterations+in+tight+junctions+of+polarized+intestinal+Caco-2+cell+monolayers.">Rotavirus-induced structural and functional alterations in tight junctions of polarized intestinal Caco-2 cell monolayers.</a> J. Virol. 74, 4645-4651 (2000).</li><li>Nagar, B., Overduin, M., Ikura, M., Rini, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Structural+basis+of+calcium-induced+E-cadherin+rigidification+and+dimerization.">Structural basis of calcium-induced E-cadherin rigidification and dimerization.</a> Nature. 380, 360-364 (1996).</li><li>Boulenc, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Importance+of+the+paracellular+pathway+for+the+transport+of+a+new+bisphosphonate+using+the+human+Caco-2+monolayers+model.">Importance of the paracellular pathway for the transport of a new bisphosphonate using the human Caco-2 monolayers model.</a> Biochem. Pharmacol. 46, 1591-1600 (1993).</li><li>Artursson, P., Magnusson, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epithelial+transport+of+drugs+in+cell+culture.+II:+Effect+of+extracellular+calcium+concentration+on+the+paracellular+transport+of+drugs+of+different+lipophilicities+across+monolayers+of+intestinal+epithelial+(Caco-2)+cells.">Epithelial transport of drugs in cell culture. II: Effect of extracellular calcium concentration on the paracellular transport of drugs of different lipophilicities across monolayers of intestinal epithelial (Caco-2) cells.</a> J. Pharm. Sci. 79, 595-600 (1990).</li><li>Artursson, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epithelial+transport+of+drugs+in+cell+culture.+I:+A+model+for+studying+the+passive+diffusion+of+drugs+over+intestinal+absorptive+(Caco-2)+cells.">Epithelial transport of drugs in cell culture. I: A model for studying the passive diffusion of drugs over intestinal absorptive (Caco-2) cells.</a> J. Pharm. Sci. 79, 476-482 (1990).</li><li>Artursson, P., Karlsson, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Correlation+between+oral+drug+absorption+in+humans+and+apparent+drug+permeability+coefficients+in+human+intestinal+epithelial+(Caco-2)+cells.">Correlation between oral drug absorption in humans and apparent drug permeability coefficients in human intestinal epithelial (Caco-2) cells.</a> Biochem. Biophys. Res. Commun. 175, 880-885 (1991).</li><li>Balda, M. S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+dissociation+of+paracellular+permeability+and+transepithelial+electrical+resistance+and+disruption+of+the+apical-basolateral+intramembrane+diffusion+barrier+by+expression+of+a+mutant+tight+junction+membrane+protein.">Functional dissociation of paracellular permeability and transepithelial electrical resistance and disruption of the apical-basolateral intramembrane diffusion barrier by expression of a mutant tight junction membrane protein.</a> J. Cell Biol. 134, 1031-1049 (1996).</li><li>Hubatsch, I., Ragnarsson, E. G. E., Artursson, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Determination+of+drug+permeability+and+prediction+of+drug+absorption+in+Caco-2+monolayers.">Determination of drug permeability and prediction of drug absorption in Caco-2 monolayers.</a> Nat. Protoc. 2, 2111-2119 (2007).</li><li>Uchida, M., Fukazawa, T., Yamazaki, Y., Hashimoto, H., Miyamoto, Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+modified+fast+(4+day)+96-well+plate+Caco-2+permeability+assay.">A modified fast (4 day) 96-well plate Caco-2 permeability assay.</a> J. Pharmacol. Toxicol. Methods. 59, 39-43 (2008).</li><li>Krug, S. M., Fromm, M., Gunzel, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two-Path+Impedance+Spectroscopy+for+Measuring+Paracellular+and+Transcellular+Epithelial+Resistance.">Two-Path Impedance Spectroscopy for Measuring Paracellular and Transcellular Epithelial Resistance.</a> Biophys. J. 97, 2202-2211 (2009).</li><li>Wegener, J., Abrams, D., Willenbrink, W., Galla, H. J., Janshoff, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+multi-well+device+to+measure+transepithelial+electrical+resistances+under+physiological+conditions.">Automated multi-well device to measure transepithelial electrical resistances under physiological conditions.</a> BioTechniques. 37, 592-594 (2004).</li><li>Weber, C. R., Shen, L., Wu, L., Wang, Y., Turner, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Occludin+is+Required+for+Tumor+Necrosis+Factor+(TNF)-Mediated+Regulation+of+Tight+Junction+(TJ)+Barrier+Function.">Occludin is Required for Tumor Necrosis Factor (TNF)-Mediated Regulation of Tight Junction (TJ) Barrier Function.</a> Gastroenterology. 140, (2011).</li><li>Owens, R. M., Malliaras, G. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Organic+electronics+at+the+interface+with+biology.">Organic electronics at the interface with biology.</a> MRS Bull. , (2010).</li><li>Lin, P., Yan, F., Yu, J. J., Chan, H. L. W., Yang, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Application+of+Organic+Electrochemical+Transistors+in+Cell-Based+Biosensors.">The Application of Organic Electrochemical Transistors in Cell-Based Biosensors.</a> Adv. Mater. 22, 3655-3660 (2010).</li><li>White, H. S., Kittlesen, G. P., Wrighton, M. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chemical+Derivatization+of+an+Array+of+3+Gold+Microelectrodes+with+Polypyrrole+-+Fabrication+of+a+Molecule-Based+Transistor.">Chemical Derivatization of an Array of 3 Gold Microelectrodes with Polypyrrole - Fabrication of a Molecule-Based Transistor.</a> J. Am. Chem. Soc. 106, 5375-5377 (1984).</li><li>Bernards, D. A., Malliaras, G. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Steady-state+and+transient+behavior+of+organic+electrochemical+transistors.">Steady-state and transient behavior of organic electrochemical transistors.</a> Adv. Funct. Mater. 17, 3538-3544 (2007).</li><li>Jimison, L. H., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measurement+of+Barrier+Tissue+Integrity+with+an+Organic+Electrochemical+Transistor.">Measurement of Barrier Tissue Integrity with an Organic Electrochemical Transistor.</a> Adv. Mater. 24, 5919-5923 (2012).</li><li>Tria, S., Jimison, L. H., Hama, A., Bongo, M., Owens, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sensing+of+EGTA+Mediated+Barrier+Tissue+Disruption+with+an+Organic+Transistor.">Sensing of EGTA Mediated Barrier Tissue Disruption with an Organic Transistor.</a> Biosensors. 3, 44-57 (2013).</li><li>Tria, S. A., Jimison, L. H., Hama, A., Bongo, M., Owens, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Validation+of+the+organic+electrochemical+transistor+for+in+vitro+toxicology.">Validation of the organic electrochemical transistor for in vitro toxicology.</a> Biochim. Biophys. Acta. 1830, 4381-4390 (2013).</li><li>Zhu, Z. T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+simple+poly(3,4-ethylene+dioxythiophene)/poly(styrene+sulfonic+acid)+transistor+for+glucose+sensing+at+neutral+pH.">A simple poly(3,4-ethylene dioxythiophene)/poly(styrene sulfonic acid) transistor for glucose sensing at neutral pH.</a> Chem. Commun. , 1556-1557 (2004).</li><li>Lin, P., Yan, F., Yu, J., Chan, H. L., Yang, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+application+of+organic+electrochemical+transistors+in+cell-based+biosensors.">The application of organic electrochemical transistors in cell-based biosensors.</a> Adv. Mater. 22, 3655-3660 (2010).</li></ol>

This technique provides a novel method to integrate an organic electrochemical transistor with live cells to measure barrier tissue integrity. The main advantages of this technique are the rapidity and sensitivity, but also the low cost of the device for dynamic monitoring of barrier tissue.
As this method uses live cells, a critical point is to be sure to use a monolayer, which represents an intact barrier layer. The parameters of the barrier should be defined during the characterization of t...

The gastrointestinal epithelium is an example of barrier tissue, which controls the passage of molecules between different compartments of the body. The epithelium consists of elongated columnar cells joined together by complexes of proteins that provide a physical barrier1 against pathogens and toxins, while allowing the passage of water and nutrients required to sustain the body. This selectivity is due to the polarization of the epithelial cells, which creates two different membrane domains: the apical side of the cells exposed to the lumen and the basal side of the cells anchored on the underlying tissues2,3. Tight junctions (TJ) are complexes of proteins present at the apical portion of epithelial cells and are part of a larger complex known as the apical junction4. Ion flow across the barrier tissue may either go via the transcellular (through the cell) or via a paracellular (between two adjacent cells) pathway. The sum of the transport through both pathways is known as the transepithelial resistance. The apical junction is responsible for the regulation of ions and molecules passing across the barrier5,6 via a specific opening and closing function. A dysfunction or disruption of these protein complexes is often related to disease7-11. In addition, many enteric pathogens/toxins are known to specifically target this complex, thereby entering the body and leading to diarrhea, most likely as a consequence of massive dysregulation of ion/water flow across the barrier12-14. Barrier tissue may also be modified by changing the extracellular microenvironment. Cadherin is a critical protein for cell-cell adhesion, and is involved in the formation of the apical junction. Calcium is required for the correct structural conformation of Cadherin, and a decrease in extracellular calcium has been shown to result in the destruction of the cell-cell junction and a subsequent opening of the paracellular pathway between the cells15. In this study, EGTA (Ethylene glycol-bis(beta-aminoethyl ether)-N,N,N&#39;,N&#39;-tetra acetic acid), a specific calcium chelator, was used to induce a breach in barrier tissue, as it has been shown to have a rapid and drastic effect on paracellular ion flow16,17. This calcium chelator was used on a confluent and differentiated monolayer of the Caco-2 cell-line. Cultured in cell culture inserts, this cell line is known to develop the characteristics of the gastrointestinal tract and is widely used by the pharmaceutical industry to test the absorption of drugs18,19.
Methods to monitor barrier tissue integrity are plentiful. These methods are often optical, relying on immunofluorescence staining of particular proteins known to be at the apical junction20, or relying on the quantification of a fluorescent tracer molecule that is normally impermeable to the barrier tissue21,22. However, label-free methods (i.e. without a fluorophore/chromophore) are preferable as the use of a label can incur artifacts, and often increases cost and assay time. Electrical, label-free monitoring of barrier tissue&#160;has recently emerged as a dynamic monitoring method23. For example recent technological advances in electrical impedance spectroscopy have allowed the development of a commercially available scanning apparatus24,25 that can measure transepithelial resistance (TER), a measurement of the ion conductance across the cell layer.
Organic electronics has created a unique opportunity to interface the world of electronics and biology26,27 28,29 by using conducting polymers that&#160;can conduct both electronic and ionic carriers. A new technique to detect breaches in barrier tissue using the OECT30-32 was recently introduced. This device was validated against existing techniques used to assess barrier tissue integrity, including immunofluorescence, permeability assays using Lucifer yellow, and impedance spectroscopy using the Cellzscope. In the case of all toxic compounds tested, the OECT was found to operate with equal or better sensitivity, and with increased temporal resolution, compared to the above techniques. In this device, PEDOT:PSS, a conducting polymer that has been shown to be stable and biocompatible33,34, is used as the active material in the transistor channel. The OECT is composed of drain and source electrodes on either side of a conducting polymer channel. This is then placed in contact with an electrolyte, which forms an integral part of the device. A gate electrode is immersed in the electrolyte (Figure 1), and when a positive gate voltage is applied at the gate, cations from the electrolyte are forced into the channel, thus dedoping the conducting polymer and resulting in a change in the source-drain current. The device is thus extremely sensitive to minute changes in ionic flux due to amplification by the transistor. A cell layer grown on a cell culture insert was placed between the gate electrode and the conducting polymer channel. The presence of an intact cell layer acts as barrier for the cations entering into the conducting polymer, therefore, in the presence of an intact monolayer, the drain current decreases (Figure 2: transition from region a to b). In the presence of a toxic compound, the barrier tissue will progressively lose its integrity, letting the cations enter into the polymer film and increasing the drain current (Figure 2: region c). With this technique, the breach in barrier tissue is seen by the modulation of the drain current, corresponding to the modulation of the flux across the monolayer. This device is able to measure minute variations in ionic flux with unprecedented temporal resolution and sensitivity in real time. This technology will be of interest in the domain of toxicology for drug testing, disease diagnostics or basic research as the barrier model can be easily adapted. This method will also help to reduce animal experimentation, as it allows the validation of in vitro models to replace in vivo testing.

<table border="0" cellpadding="0" cellspacing="0" width="589">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">CLEVIOS PH 1000</td>
			<td style="width:87px;">HERAUS CLEVIOS</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:216px;">AZ9260 resin</td>
			<td style="width:87px;">CIPEC SPECIALITIES</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">Dodecylbenzenesulfonic acid (DBSA)</td>
			<td style="width:87px;">Acros Organic</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:216px;">3-glycidoxypropyltrimethoxysilane (GOPS)</td>
			<td style="width:87px;">Sigma Aldrich</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:216px;">24-well Suspended cell Culture insert Millicell&#160; PET 0.4 &#956;m Millipore</td>
			<td style="width:87px;">Dominique dutscher</td>
			<td style="width:119px;">51705</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">24-well cell culture plate BD Falcon</td>
			<td style="width:87px;">Dominique dutscher</td>
			<td style="width:119px;">51705</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">STERICUP-GP PES 0.22 &#956;M</td>
			<td style="width:87px;">Dominique dutscher</td>
			<td style="width:119px;">51246</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">ADVANCED DMEM Marque GIBCO</td>
			<td style="width:87px;">Fisher scientific</td>
			<td style="width:119px;">E3434T</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">FBS HEAT INACT. S.AMERICAN</td>
			<td style="width:87px;">Fisher scientific</td>
			<td style="width:119px;">E3387M</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">PENICILLIN STREPTOMYCIN</td>
			<td style="width:87px;">Fisher scientific</td>
			<td style="width:119px;">E3470C</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">GLUTAMAX</td>
			<td style="width:87px;">Fisher scientific</td>
			<td style="width:119px;">E3524T</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">TRYPSIN 0.05% EDTA</td>
			<td style="width:87px;">Fisher scientific</td>
			<td style="width:119px;">E3513N</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:216px;">EGTA (Ethylene glycol-bis(2-aminoethylether)-N,N,N&#8242;,N&#8242;-tetraacetic acid)</td>
			<td style="width:87px;">Sigma Aldrich</td>
			<td style="width:119px;">E4378</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">ETHYLENE GLYCOL, ANHYDROUS, 99.8%,</td>
			<td style="width:87px;">Sigma aldrich</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Caco-2 cells</td>
			<td style="width:87px;">ATCC</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:216px;">PDMS</td>
			<td style="width:87px;">Dow corning</td>
			<td style="width:119px;">SYLGARD 184 SILICONE ELASTOMER</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Au (99.99%)</td>
			<td style="width:87px;">NEYCO</td>
			<td style="width:119px;">AU3X6</td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="22">
			<td height="22" style="height:22px;width:216px;">Chromium (99.95%)</td>
			<td style="width:87px;">NEYCO</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:216px;">Parylene C</td>
			<td style="width:87px;">Specialty Coating Systems</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="43">
			<td height="43" style="height:43px;width:216px;">Ag/AgCl wire</td>
			<td style="width:87px;">HARVARD APPARATUS</td>
			<td style="width:119px;"></td>
			<td style="width:167px;"></td>
		</tr>
		<tr height="64">
			<td height="64" style="height:64px;width:216px;">Photoresist</td>
			<td style="width:87px;">CIPEC SPECIALITIES</td>
			<td style="width:119px;">R&#233;sine AZ9260</td>
			<td style="width:167px;"></td>
		</tr>
	</tbody>
</table>

sensing of barrier tissue disruption with an organic electrochemical transistor

The gastrointestinal tract is an example of barrier tissue that provides a physical barrier against entry of pathogens and toxins, while allowing the passage of necessary ions and molecules. A breach in this barrier can be caused by a reduction in the extracellular calcium concentration. This reduction in calcium concentration causes a conformational change in proteins involved in the sealing of the barrier, leading to an increase of the paracellular flux. To mimic this effect the calcium chelator ethylene glycol-bis(beta-aminoethyl ether)-N,N,N&#39;,N&#39;-tetra acetic acid (EGTA) was used on a monolayer of cells known to be representative of the gastrointestinal tract. Different methods to detect the disruption of the barrier tissue already exist, such as immunofluorescence and&#160;permeability assays. However, these methods are time-consuming and costly and not suited to dynamic or high-throughput measurements. Electronic methods for measuring barrier tissue integrity also exist for measurement of the transepithelial resistance (TER), however these are often costly and complex. The development of rapid, cheap, and sensitive methods is urgently needed as the integrity of barrier tissue is a key parameter in drug discovery and pathogen/toxin diagnostics. The organic electrochemical transistor (OECT) integrated with barrier tissue forming cells has been shown as a new device capable of dynamically monitoring barrier tissue integrity. The device is able to measure minute variations in ionic flux with unprecedented temporal resolution and sensitivity, in real time, as an indicator of barrier tissue integrity. This new method is based on a simple device that can be compatible with high throughput screening applications and fabricated at low cost.

During the first step of measurement, the drain current may vary somewhat, but in most cases it should remain stable (Figure 2, section a). If the signal is not stable, the transistor should be discarded and replaced. This stability check also ensures that any initial losses in conductivity of the device do not affect the subsequent measurement. After several minutes of measurement, the insert with cells forming barrier tissue is placed on top of the channel. The drain current should immediately decrease...

Watch this Scientific Journal Video about Sensing of Barrier Tissue Disruption with an Organic Electrochemical Transistor at JoVE.com

Sensing of Barrier Tissue Disruption with an Organic Electrochemical Transistor

The Organic Electrochemical Transistor is integrated with live cells and used to monitor ion flux across the gastrointestinal epithelial barrier. In this study, an increase in ion flux, related to disruption of tight junctions, induced by the presence of the calcium chelator EGTA (ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetra acetic acid), is measured.

The gastrointestinal tract is an example of barrier tissue that provides a physical barrier against entry of pathogens and toxins, while allowing the ...

Research

JoVE Journal

Bioengineering

Hollow Microneedle-based Sensor for Multiplexed Transdermal Electrochemical Sensing

The development of a minimally invasive multiplexed monitoring system for rapid analysis of biologically-relevant molecules could offer individuals ...

Bacterial Detection &amp; Identification Using Electrochemical Sensors

Bacterial Detection & Identification Using Electrochemical Sensors

Electrochemical  sensors are widely used for rapid and accurate measurement of blood glucose and  can be adapted for detection of a wide variety of ...

Electric Cell-substrate Impedance Sensing for the Quantification of Endothelial Proliferation, Barrier Function, and Motility

Electric Cell-substrate Impedance Sensing (ECIS) is an in vitro impedance measuring system to quantify the behavior of cells within adherent cell layers. ...

Preparation of Silicon Nanowire Field-effect Transistor for Chemical and Biosensing Applications

Surveillance using biomarkers is critical for the early detection, rapid intervention, and reduction in the incidence of diseases. In this study, we ...

Tissue Engineering by Intrinsic Vascularization in an In Vivo Tissue Engineering Chamber

Tissue Engineering by Intrinsic Vascularization in an In Vivo Tissue Engineering Chamber

In reconstructive surgery, there is a clinical need for an alternative to the current methods of autologous reconstruction which are complex, costly and ...

Wet Chemistry and Peptide Immobilization on Polytetrafluoroethylene for Improved Cell-adhesion

Endowing materials surface with cell-adhesive properties is a common strategy in biomaterial research and tissue engineering. This is particularly ...

Development of an Electrochemical DNA Biosensor to Detect a Foodborne Pathogen

Vibrio parahaemolyticus (V. parahaemolyticus) is a common foodborne pathogen that contributes to a large proportion of public health problems globally, ...

In Vitro Multiparametric Cellular Analysis by Micro Organic Charge-modulated Field-effect Transistor Arrays

In Vitro Multiparametric Cellular Analysis by Micro Organic Charge-modulated Field-effect Transistor Arrays

Modern electrophysiology has been constantly fueled by the parallel development of increasingly sophisticated tools and materials. In turn, discoveries in ...

Disruption of the Blood-Spinal Cord Barrier Using Low-Intensity Focused Ultrasound in a Rat Model

Low-intensity focused ultrasound (LIFU) uses ultrasonic pulsations at lower intensities than ultrasound and is being tested as a reversible and precise ...

Evaluating MOF-Induced Cell Behavior Changes by Electric Cell-Substrate Sensing

Electric Cell-Substrate Sensing for Real-Time Evaluation of Metal-Organic Framework Toxicological Profiles

Author Spotlight: Advances in Evaluating Human Lung Epithelial Cells' Response to Metal-Organic Frameworks 

Metal-organic frameworks (MOFs) are hybrids formed through the coordination of metal ions and organic linkers in organic solvents. The implementation of ...