Life scientists use rather bulky measurement devices to study sensitive cells. Huge pipettes or cannulas are pushed into ultra-tiny structures such as cell membranes or cytoplasm in order to measure complex processes or reactions inside cells. Rough treatment of this kind can damage cells and affect measurement results in ways that are difficult to quantify. The materials scientist Steffen Strehle from Ulm University has plans to begin development of a bionanosensor as part of an interdisciplinary collaborative project intended to provide the life sciences community with a minimally invasive tool to penetrate cells gently and carry out direct, highly sensitive measurements.
Strehle’s approach, which is funded by the German Ministry of Education and Research (BMBF) with 1.8 million euros for four years, is aimed at reducing the size of a measuring device by around several magnitudes in order to be able to facilitate the investigation of the nanoworld of molecular biology. The young scientist uses the following principle in his development project: “In semiconducting nanostructures, attaching a small number of molecules on the surface is usually enough to change the electrical conductivity of the surface structure considerably. If the surface properties only cause specific molecules to attach to the nanostructure, these molecules can be detected electrically with high sensitivity.”Strehle, who is a junior professor in the Institute of Electronic Devices and Circuits at Ulm University, will use nanowires 50 nanometres (i.e. 1 nanometre = 1 billionth of a metre) in diameter, which are 500 times thinner than a human hair. For comparison, cells have an average diameter of between one and 30 micrometres (i.e. 1 micrometre = 1 millionth of a metre) and cell membranes are between six and ten nanometres thick.
If everything goes to plan, Strehle would like to have developed a novel, commercially applicable bionanosensor system that enables cell biologists to take a close and precisely locatable look into cells and medical doctors to detect disease markers biochemically within four years. The bionanosensor will be developed by an interdisciplinary team from Ulm University, including scientists from the field of organic and analytical chemistry, biochemists, materials scientists (engineers), semiconductor physicists and electrical engineers, and two industrial partners.
During his time as postdoctoral fellow at Harvard University in Boston, Strehle was able to show that it is feasible to push silicon nanotubes into living heart muscle cells in the same way as needles and record their heartbeat, or in other words, to perform a single-cell ECG. In the laboratory of Charles Lieber, one of the most influential chemists in the world and, as Strehle puts it, the guru of silicon nanowires, Strehle explored the potential of using nanowires in the field of biology, which is his favourite subject.
According to Strehle, the advantage of using silicon nanostructures is that the researchers have recourse to a mature, 60-year-old technology in their search for new applications. Many biosensors are available, including biosensors made of silicon. They have been shown to detect a substance of interest relatively accurately, simply and reliably. In the life sciences, researchers usually use indirect methods for detecting substances, including for example fluorescence-labelling methods. However, these methods are associated with a number of drawbacks: On the one hand, fluorescent proteins need to be brought into the cell, which however disturbs the cellular system. In addition, the fluorescent label fades relatively quickly. The measurement of signals in the nanoscale cell membrane usually involves the use of the so-called patch clamp technique, a method which was developed in the 1970s. It allows high-resolution recording of the ionic current that flows through a cell's plasma membrane and thus transmembrane ion channels to be studied in detail. However, due to the size of the probes used, this technique is rather invasive and tends to damage cells.
Sense-U, as Strehle’s project is called, hopes to find what cell biologists are looking for: probes that disrupt the cells as little as possible while allowing direct measurements to be carried out. Strehle’s method has the potential to enable the study of the reactions of less stressed cells to pharmacological and toxicological substances. However, Strehle is perfectly aware of the fact that despite the considerably reduced size of the novel bionanosensor, it is still invasive in some ways.
Strehle’s U-Sense project aims to find out how the silicon surface can be functionalised in a way that enables only the molecule of interest to attach. The attachment leads to changes in the molecules' charge, which in turn can be measured as a change in the electric field on the surface. However, the problem is that these electric fields are rather small. Strehle and his colleagues have found a solution, which involves the miniaturisation of the structures, i.e. the development of nanowires of between 20 and 50 nanometres in diameter. Thanks to the wires' spherical structure, the molecules can attach on the entire wire surface, which is impossible on planar structures. The attachment of molecules to the spherical surface of the wire produces an electric field which penetrates one to several nanometres into the cell. Strehle explains that this phenomenon turns a surface effect into a volume effect, which increases the measuring sensitivity. He also points out that the detection of a molecule works best when the measurement field of the sensor has a similar range.
A major advantage of nanosensors is their high resolution: “We are able to measure more locally,” explains Strehle. A 50-nanometre sensor is able to record the currents of a single transmembrane ion channel that is around 100 nanometres away from the nearest ion channel with a greater probability than with other methods.
Strehle needs to repeat the experiment he once did at Harvard (“We have made quite good progress”) before he can take the sensor from the laboratory stage and transform it into a standardised life sciences method. This step often fails for various reasons. This is not likely to be the case in Ulm, where Strehle and his colleagues have at their disposal a well-equipped clean-room. Strehle’s task now is to further develop the functional nanosensor system using practices of modern engineering, produce the appropriate quantities, and to ensure that the system works reliably and delivers reproducible results. Strehle knows that life scientists depend on standard techniques and practicable measurement methods, and this can be a biochip for conducting biochemical analyses in ultra-small volumes or a nanowire for intracellular recordings.
The Ulm scientists are therefore working with companies like Cetoni, a German company specialised in the development and production of automated systems, micro- and nano-dosage systems, and Bruker, a global manufacturer of analytical and medical systems. However, the scientists will first have to apply innovative engineering skills to the design of the sensor. “We will learn how to manipulate nanowires, how to move them and how to get them to the target site.” One of the project priorities will be the development of new microfabrication strategies which will enable us to enter the other dimension of living things. Strehle believes that the microfabrication of the sensor should not pose a major problem. However, the researchers also need access to suitable measurement techniques, without which the sensor would be worthless. Strehle is well aware of the challenge he and his partners face; cells are complex systems where a huge number of different processes occur at different sites. He believes that the development of suitable filters is one of the project’s milestones, where the partners’ biological expertise will be put to good use for identifying molecules that can trigger similar reactions and hence lead to similar measurement results.The scientists will start with the synthesis of the nanostructure, which must be reproducible and reliable, and then concentrate on the chemical functionalisation of the surface. “During this process, we materials scientists will learn a lot about cells,” said Strehle referring to the fact that he and his fellow engineers will have to find answers to questions such as: for how long does the sensor work, do interactions with the cell occur, do the sensors clog, does the cell show signs of rejection, for how long can the measurements be carried out and what kind of signals can be expected?Once the design and the manufacture of the sensor are in place, Strehle will work with his life sciences partners in order to develop the functionalisation of the surface for one specific case. Strehle pointed out that they have quite a few suitable ideas and is fairly confident that their approach will work, especially as the silicon technology is already established and only needs to be adapted to nanostructures. He is well aware of the attraction of his approach: nanotechnology entering application and opening up new possibilities for users. It is not too far-fetched to describe Strehle as a researcher who reached out and grasped at least one of the many promises of nanotechnology.
Ruixuan Gao, Steffen Strehle, Bozhi Tian, Tzahi Cohen-Karni, Ping Xie, Xiaojie Duan, Quan Qing, and Charles M. Lieber: Outside Looking In: Nanotube Transistor Intracellular Sensors. Nano Lett. 2012, 12(6), pp. 3329-3333.