Measurement of the transepithelial electrical impedance has been used since the 1980s to determine confluency and barrier function of epithelial monolayers in cell culture. The underlying technique is a four-terminal sensing which uses different pairs of current-carrying and voltage-sensing electrodes to make more accurate measurements. There are several commercially available devices to measure transepithelial impedance, but despite the ease of use and the high reliability, there are also some disadvantages as the non-translatable output frequency and their expensiveness.
Thus, we represent how to build a cost-effective and programmable volt-ammeter. At first, we want to show how TEER measurement with commercially available devices works. To do so, we had cultured a cell layer of choroid plexus epithelial papilloma cells on Transwell filters with a pore size of three micrometer.
This setting has been described by Schroten et al. as in vitro model of the blood-cerebrospinal fluid barrier. Now, a chopstick electrode is connected to an epithelial volt-ammeter.
The device is switched on and set to measure resistance. The electrode is sterilized in 80%ethanol and subsequently equilibrated in an appropriate medium. The impedance to be measured will serve as reference value in order to evaluate the reliability of the volt-ammeter that will be assembled afterwards.
In this example, we record an impedance of 680 ohms. Let's start with the assembly of a low-budget volt-ammeter with programmable output frequency. First of all, you will need a standard USB charger, and an USB extension cord as five-volt direct current power supply.
An 8-bit microcontroller on a USB development board will then be used to generate a square wave current. Four cables with banana plugs are connected to two standard multimeters to measure voltage and current. Please ensure that the multimeters are capable of measuring current in a range of some microamperes with TrueRMS.
A female RJ14 connector can be found on standard telephone cables. Just make sure, that the connector has six pins of which at least the inner four are wired. Finally, you will need some standard equipment, as our cables, a luster terminal, a 120 kiloohm resistor, and some tools as insulation strippers, a crimping tool, wire end ferrules, and soldering iron.
The device is assembled exactly as illustrated in the layout diagram. At first, the USB extension is connected to the microcontroller. During normal operation, it is powered by a five-volt DC USB charger, that can readily be connected to a personal computer for programming.
Two cables are stripped and crimped with wire end ferrules on one side. The other side is soldered either directly to pin zero and two of the microcontroller or to soldering lugs, which in turn are clipped on the respective pins. Next, the power-delivering cables are connected to a luster terminal.
The first multimeter will be used to measure current and is connected in series with 120 kiloohm resistor and a current-passing chopstick electrodes. This arrangement ensures that the output current is limited, so the measurement will have no impact on cell viability. The four conductors of the telephone extension cord are dismantled and crimped to ferrules as shown before.
Once the cable is prepared, you will have to test continuity of conductors and pins. In our example, pin three to six are connected to the white, brown, green and yellow conductor. Now pin five and six, that is green and yellow, are connected to the luster terminal to apply voltage to the outer electrode pair.
Finally, you will have to connect the second multimeter, which will be used to measure the transepithelial voltage drop, to pins three and four, that is, in our example, to the white and brown conductor. We decided to mount installation in a cheap plastic chassis. Before the first use, the microcontroller has to be programmed.
The source code is written in C+and can be uploaded by a USB. In brief, pin zero into a center output mode. When switched on, the function loop will start to alternate as pins between ground and plus five volts with a variable delay.
In our example, we used a theoretical half-oscillation time of 40 milliseconds. Let's see how the measurement results compare to the reference values we had obtained before. The chopstick electrode is repositioned to the recently assembled volt-ammeter.
The device is being powered on in three steps. That is, plugging in a USB charger, switching the left multimeter to AC voltage measurement, and switching the second multimeter to microampere. Pay attention, that alternating current has to be selected explicitly.
In this example, the potential drop across the Transwell filter system is measured as approximately 25 millivolts, while we record a current of 37.1 microamperes. According to Ohm's Law, electric impedance can be easily calculated at 674 ohms, which is very close to the reference value of 680. We've shown that measurement values are reliable over a range from zero to 1.8 kiloohms.
Thus, the described volt-ammeter can be used both for initial experiments and for further studies. However, if your results shall be published, you might always want to support your data, by measuring molecule flux across the respective cell layer.
