Sensor Issues pubs.acs.org/acssensors
Can Calibration-Free Sensors Be Realized? Eric Bakker* Department of Inorganic and Analytical Chemistry, University of Geneva, Quai E.-Ansermet 30, 1211 Geneva 4, Switzerland ABSTRACT: This opinion piece describes progress and issues regarding the realization of calibration-free chemical sensors for analysis in aqueous systems. A special focus of the discussion is on electrochemical methodologies, given that they are the most established. Calibration-free sensors are clearly possible, but require a multipronged approach for maximizing robustness that include the optimization of sensing materials and the use of the most adequate readout methodologies. KEYWORDS: chemical sensors, calibration-free, clinical diagnostics, electroanalysis, potentiometry, thin layer coulometry
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the time evolution of the signal, which can all be used to distinguish random fluctuations from an event that can be traced across numerous sensor points.3 Calibration requirements are clearly less stringent with such systems. For glucose biosensors, the target range for diabetic patients is 4.0−9.0 mM, about a factor 2 in concentration, while glucose test strips should be able to measure in the range of 1−30 mM. The measure of success with glucose sensors is the location of a point on an Clarke Error Grid, and therefore concentration dependent, but an error of less than 20% is typically considered acceptable.4 On the other hand, the normal range for blood electrolytes is much narrower, for potassium 3.5−5.0 mM, a 43% change in concentration, and for sodium 136−145 mM, a mere 7% change.5 As explained in a recent review on clinical diagnostics of electrolytes,5 a measurement must distinguish the electrolyte level sufficiently to allow for a meaningful diagnosis.
long lasting focus of modern chemical sensor research has been to move the measurement out of the traditional laboratory and as close as possible to the site of interest. Indeed, clinical measurements of key blood parameters are today routinely performed at the point of care. Glucose measurements are achieved by millions of people in the comfort of their own home with hand-held electrochemical biosensor devices. Remote environmental sensing is today a reality, even though the number of accessible analytes is still limited. The promise for a chemical sensor to work autonomously for long periods of time, without the need for sample preparation and without (re)calibration, remains a key selling point for research in this area. Recent work has aimed at realizing very low cost, paper-based sensing systems that can be put into the hands of low skilled operators while maintaining meaningful results.1 In the environmental sensing area, an important direction has been the development of large, autonomously operating sensor networks that provide a real time chemical map to visualize environmental contaminations and for the localization of plumes.2 Only if chemical sensors can operate with the ease and robustness demanded by the application will these directions truly become successful and have the potential to have a significant impact to our world. In this context, calibration-free sensors are primarily understood as being maintenance-free, i.e., comprising a sufficiently robust system that can be precalibrated in the factory, and used as is. This opinion piece focuses on the issues regarding calibration-free chemical sensors, with a special focus on electrochemical methodologies, since this is where most progress has historically been made.
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A CHALLENGING CASE STUDY: CLINICAL DIAGNOSTICS OF BLOOD ELECTROLYTES Given the above numbers, arguably the most challenging routine analytical sensing application in terms of required precision and accuracy is the assessment of blood electrolytes. Clinical analyzers achieve the required analytical figures of merit by careful termperature control and frequent intermittent washing and one point calibration steps and are automated, but not maintenance or calibration-free. An early study by Rumpf et al. demonstrated the absolute measurement of potassium and sodium levels in serum by ionselective electrodes using a highly symmetrical measurement cell; see Figure 1.6 The authors did not achieve a maintenancefree measurement, since high flow rates of the reference electrolyte and intermittent cleaning steps were required to renew the two reference electrodes and the side of the sensing membrane exposed to the biological fluids. But measurements without prior calibration were demonstrated, with errors of 0.02
WHAT ERROR CAN ONE TOLERATE?
The question of the required out of the box accuracy of different sensor systems must be answered by the anticipated analytical problem. An environmental contamination might increase the basal concentration by many orders of magnitude, depending on incident and type of released chemical. Moreover, the readout of a large sensor network benefits from knowledge of the behavior of neighboring sensors and of © XXXX American Chemical Society
Received: April 12, 2016 Accepted: June 22, 2016
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DOI: 10.1021/acssensors.6b00247 ACS Sens. XXXX, XXX, XXX−XXX
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ACS Sensors
an ionic liquid between the film and the sample. Buhlmann has recently explored this new type of reference electrode in paper based electrochemical sensing devices with promising results.1 The future will show whether this exciting direction will eventually result in sensors with dramatically improved robustness.
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INCREASING ROBUSTNESS WITH ENZYME BIOSENSORS But what about amperometric chemical sensors, such as enzyme biosensors? Enzymes denature over time, and moreover, any direct contact of electrodes with complex samples is known to result in surface fouling. It is possible to achieve adequate robustness by controlling the rate limiting step and minimizing electrode fouling. With enzyme biosensors, a diffusion limiting membrane is placed between enzyme layer and sample.12 Besides helping to achieve adequate selectivity, the resulting current remains diffusion limited at all times and even a substantial loss of enzyme activity will not result in a deterioration of the signal. The membrane also serves to protect the active electrode surface from direct contact with the sample and hence minimizes fouling. Today, amperometric glucose sensors are mass fabricated and a subset of the sensor strips factory calibrated to describe the response of the entire batch.4 Direct film coatings onto electrodes (without enzyme) may also result in robust amperometric sensors for the detection of electrically neutral drug molecules (such as propofol) that can easily partition into a hydrophobic film and react at the underlying electrode,13 although calibration-free sensors were not the aim of the study. As typical polymeric permeation membranes exclude ionic and bulky interferences, antifouling coatings for the electrochemical detection of ionic analytes must follow alternative strategies. For example, hydrogel coatings have been demonstrated to be attractive in this regard since they act as size exclusion membranes to keep macromolecular and colloidal interferences away from the membrane.14 As they are less effective as diffusion limiting membranes, thicker coatings (hundreds of micrometers) are needed to ensure that the ions are sourced from within the gel during measurement, thereby minimizing effects of mass transport fluctuations in the contacting sample. Note that low molecular weight interferences (such as surfactants) may still permeate across the gel and can therefore result in chemical interference and/or electrode fouling.
Figure 1. Symmetrical measuring cell for the sensing of potassium and sodium ions in undiluted serum without prior calibration.6 Two reference electrodes with free-flowing liquid junctions were used in this arrangement. While this approach was not maintenance-free, a residual error of just a fraction of a millivolt was found.
mM for potassium and 1.4 mM for sodium (tolerable errors: 0.10 and 0.5 mM, respectively). Indeed, the signal from a symmetrical cell containing a reference solution of known composition carries information and can be considered an internal one step calibration. This sensing approach is similar to the Kodak Ektachem electrolyte sensor (today the Johnson & Johnson Vitros system),7 which uses thick film technology in a highly symmetrical arrangement where the signal is generated from applying the sample and a reference solution to the chip. Recent work on paper based ion-selective electrodes equally aimed to use a cell of high symmetry in which the sample and a reference solution were applied to two different zones on the paper (no interdevice repeatabilities were reported).1 In other work, mass fabricated all solid state ion sensors have been shown to give out of the box reproducibilities of just 0.2 mV at millimolar sample concentrations2b and demonstrating “fit for purpose” environmental trace detection as well,8 suggesting that these systems can be batch calibrated. While this looks great, adsorption and surface fouling by biological, macromolecular and chemical constituents of the sample are today still major practical limitation to achieve maintenance-free sensors that are free of signal drift.5 Important efforts in materials durability and compatibility with complex samples have been achieved, for example with hydrophilic surface coatings9 and nitric oxide release strategies for active biocompatibility control on appropriate sensing materials.10 Unfortunately, liquid junction reference electrodes are fundamentally unattractive choices for achieving maintenancefree electrochemical sensors. Recently, important progress has been made in introducing fundamentally sound reference elements that no longer require a liquid junction, which tends to be the major source of error in potentiometric experiments. While not yet established in clinical diagnostics applications, a most promising candidate appears to be based on Kakiuchi’s ionic liquid reference electrode.11 Rather than relying on a liquid junction potential between the sample and a salt solution, the potential at this film is based on equilibrium partitioning of
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USING ROBUST METHODOLOGIES: FROM AMPEROMETRY TO COULOMETRY Still, f urther progress will likely remain limited if one continues to use detection principles that are highly sensitive to chemical changes of the sensing surface. Despite the important advances described above, amperometry is an inherently kinetic technique and the signal remains very sensitive to fluctuations in temperature and other processes that influence the kinetics of the rate limiting step. On the other hand, direct potentiometry gives a signal that is dependent on the logarithm of the ion activity change, which places important demands on the tolerable error in potential. Let us therefore step back in time to the 1830s with Michael Faraday, who in an electrolysis experiment measured the mass of deposited metal and the corresponding amount of charge.15 The latter was measured by the volume of gas (hydrogen or B
DOI: 10.1021/acssensors.6b00247 ACS Sens. XXXX, XXX, XXX−XXX
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ACS Sensors oxygen, depending on the direction of current flow) produced from an acidified water solution (the so-called volta-electrometer, later voltameter or coulometer). This allowed him, without prior calibration, to relate amount (number of moles) to the charge passed across the cell, which formed the basis for Faraday’s first law of electrolysis (the second law simply states that chemical and electrochemical equivalences are the same). This was an alternative and infinitely more elegant manner of determining the molar mass of metals than the laborious procedures developed earlier by Berzelius. Sadly, Berzelius did not appear capable of understanding Faraday’s approach and apparently made sure to use his influence to suppress its acceptance within the wider scientific community during his lifetime. We realize now that Faraday’s early experiments were very close in spirit to today’s aspirations by the sensor community for simple, low cost devices that show an easily visualized analog signal (here, volume of produced gas or mass of plated metal) and that require no calibration (by using coulometry)! Indeed, conceptual improvements of glucose biosensors, while allowing for a significant reduction in sample size, have been achieved with an exhaustive, coulometric sensing mode.16 If the sample glucose level can be quantitatively and selectively depleted by the biosensor, the current may be integrated to give total charge, which is related to the amount of analyte by Faraday’s law. To reduce analysis time, the sample layer thickness should normally not exceed that of the diffusion layer (typically less than 100 μm). If, and only if, the entire current can be ascribed to the reaction of interest (excellent selectivity and nearly 100% coulometric efficiency) and the sample volume is fixed by a suitable microfluidic channel, one may achieve a calibration-free sensor. Can such a device still be called a sensor? Perhaps not in the strictest sense, since the sample solution is altered by the measurement. But if this perturbation involves only a very small portion of the total sample that can easily be replenished by diffusion or fluidic delivery, perturbation is no more severe than with other amperometric sensors where the composition of the diffusion layer close to the electrode is altered in a similar way. A thin layer coulometric glucose biosensor (FreeStyle) by Abbott/Therasens is commercially established and requires a sample volume of just 300 nL.16 A subcutaneously implantable glucose monitoring system has also been developed based on the technology, although a calibration step is here still required to correct for matrix effects. Coulometric sensors with thin layer sample layers are also promising for the detection of ionic species. In one recent example, shown in Figure 2, the three halides chloride, bromide and iodide were detected with high precision by exhaustive oxidative plating onto a silver electrode.17 The thin layer sample was defined by the space between the silver electrode and a Donnan exclusion cation-exchange membrane, outside of which was an electrolyte solution and the reference/counter electrode. Interdevice reproducibility was found to be 2.5% (N = 6), even though the cells were made by hand. Note that a comparable reproducibility in direct potentiometry would need to ensure variations of less than 0.5 and 0.3 mV for monovalent and divalent ions, respectively. This is already quite difficult. One can also make use of the coulometric thin layer sample concept to electrochemically deliver ions. This may result in a family of sensing concepts that essentially behave the same way as volumetric titrations, but without the hassle of taking aliquots, preparing standards and doing the volumetric delivery.
Figure 2. Thin layer coulometric halide detection as a more robust readout principle compared to potentiometry. Top: experimental arrangement and relevant reactions.17 Shown here is the exhaustive anodic deposition of chloride onto a silver element, while the countercation migrates across a Donnan exclusion cation-exchange membrane into the outer solution. There, reduction at the counter electrode results in a dissolution of halide ions. Bottom: when interrogated with a linear potential sweep at an appropriate scan rate, the integrated current gives the total charge, which is translated to concentration with Faraday’s law.
In one such recent example, a total alkalinity sensor was developed that delivers hydrogen ions by electrochemical control across a pH responsive polymeric membrane of high mobility.18 A pH probe placed directly opposite across the thin layer sample gap of ca. 100 μm measures the resulting pH. No pumps or moving parts were required to achieve this coulometric alkalinity probe. While the interdevice reproducibility for this system has not yet been rigorously characterized, mass fabrication should result in excellent repeatabilities, as demonstrated by the earlier work of van der Schoot et al.,19 who developed a dynamically operating alkalinity probe that was fully mass produced.
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CONCLUSIONS AND OUTLOOK Recent years have seen significant progress with optimized materials and methodologies to make maintenance-free electrochemical sensors a real possibility. In many instances, the recording of fluctuations with time, or the identification of point sources with large arrays are more important than absolute values, which places lesser requirements on the accuracy of sensor outputs. In all cases, however, one must ask: what is the intended application? Obviously, a sensor should be evaluated in view of that application, using adequate materials that are compatible with the matrix of interest. Moreover, there exist analytical methodologies, such as exhaustive coulometry, that are inherently more robust than others. This should be taken into account from the start of the research to achieve the important goal of sensors that no longer need to be calibrated in the field. This is in fact the answer to the question of what error can one tolerate? In all cases the answer is, the system must be as good as required and only that good! C
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(15) Ehl, R. G.; Ihde, A. J. Faraday’s Electrochemical Laws and the Determination of Equivalent Weights. J. Chem. Educ. 1954, 31, 226− 232. (16) Heller, A.; Feldman, B. Electrochemistry in Diabetes Management. Acc. Chem. Res. 2010, 43, 963−973. (17) Cuartero, M.; Crespo, G. A.; Ghahraman Afshar, M.; Bakker, E. Exhaustive Thin-Layer Cyclic Voltammetry for Absolute Multianalyte Halide Detection. Anal. Chem. 2014, 86, 11387−11395. (18) Ghahraman Afshar, M.; Crespo, G. A.; Bakker, E. Thin-Layer Chemical Modulations by a Combined Selective Proton Pump and pH Probe for Direct Alkalinity Detection. Angew. Chem. 2015, 127, 8228− 8231. (19) van der Schoot, B.; van der Wal, P.; de Rooij, N.; West, S. Titration-on-a-Chip, Chemical Sensor−Actuator Systems from Idea to Commercial Product. Sens. Actuators, B 2005, 105, 88−95.
AUTHOR INFORMATION
Corresponding Author
*E-mail:
[email protected]. Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS The author thanks the Swiss National Science Foundation and the European Union (FP7-GA 614002-SCHeMA project).
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REFERENCES
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