Rossana E. Madrid

Impedance microbiology: quantification of bacterial content in milk by means of capacitance growth curves.

The impedancimetric method is a technique for the rapid evaluation of milk bacterial content and also of its subproducts. Several authors have made use of culture conductance changes during bacterial growth for quantitative and qualitative assessments of microbial growth. However, interface capacitance curves, Ci, have not been used. In this paper, we quantify bacteria in cow raw milk by following their growth as the above-mentioned capacitance change time course event. With it, bigger growth variations, shorter detection times and a better coefficient of correlation with the plate count method were obtained than those yielded by conductance curves. Calibration was performed by plotting initial known concentrations, IC (CFU/ml), as a function of the time detection theshold (TDT).

Impedance bacteriometry: medium and interface contributions during bacterial growth

Impedance was measured in a cell containing culture broth inoculated with E. coli, before and during bacterial growth. The electrode interface impedance components (R/sub i/, X/sub i/) and the culture medium component R/sub m/ were separated by making use of the Warburgs model frequency dependent properties. Measurements were carried out from 18 Hz to 18 kHz with a constant current impedance bridge as growth proceeded. Growth curves for R/sub i/ and X/sub i/ showed a similar temporal pattern within the frequency range of 18-100 Hz. Dispersion was not observed in R/sub m/, meaning that the same growth response was obtained within the 18-18000-Hz range. At low frequency, the resistive and capacitive reactive components, or R/sub i/ and X/sub b/, respectively, were directly measured, where R/sub b/=(2*R/sub i/+R/sub m/) and X/sub b/=2*X/sub i/ and, above 5 kHz, R/sub m/ was obtained (for Z/sub i/ is negligible). Thus, R/sub i/ was easily discriminated from R/sub m/ by simple arithmetic. In four experiments, the maximum spread of X/sub i/, R/sub i/, and R/sub m/ was smaller than 5% indicating good repeatability. There is potential new information in dissecting out the growth curve in three separate component curves.<<ETX>>

Evaluation of chrono-impedance technique as transduction method for a carbon paste/glucose oxidase (CP/GOx) based glucose biosensor.

The chrono-impedance technique (CIT) for real time determination of glucose concentration in a first generation glucose oxidase/carbon paste electrode was implemented. The biosensor was polarized with a signal composed of 900 mV DC potential and 50 mV(RMS) AC signal at 0.4 Hz. A frequency response analyzer was used to measure the complex impedance (magnitude |Z| and phase (Φ)) of the biosensor-bulk interface. Real time measurements were performed while glucose was added to the bulk within a concentration range of 0-40 mM. The cumulative impedance dose-response curves were used to construct calibration curves, both for magnitude and phase. The best fitting was obtained with a hyperbolic equation. Four biosensors were built obtaining five calibration curves for each of them. A single test measurement (unknown glucose concentration) was also obtained after each calibration procedure. Glucose concentrations were estimated with the calibration curves and also measured by colorimetry, the latter being the reference method. Besides, one-way ANOVA test evaluated repeatability. Difference between means was not statistically significant (p>0.01) for both magnitudes (|Z| and Φ). The Students t-test assessed the differences significance, which produced in all cases p levels lower or equal than 0.44. Thus, CIT was proved to be a reliable method to measure glucose concentration in real time. Moreover, it showed high repeatability and compared well against colorimetry (r(2)=0.98).

A pH Sensor Based on a Stainless Steel Electrode Electrodeposited With Iridium Oxide

A simple procedure to make an iridium oxide (IrO2) electrodeposited pH sensor, that can be used in a chemical, biomedical, or materials laboratory, is presented here. Some exercises, based on this sensor, that can be used to teach important concepts in the field of biomedical, biochemical, tissue, or materials engineering, are also presented. This novel procedure is based on the electrodeposition of IrO2 on a stainless steel electrode, which uses a similar mechanism to an ion selective electrode (ISE) and senses changes in pH. The simplicity and cost effectiveness of this method facilitates the teaching of the concept of half-cell potential and the basics of sensors. This novel sensor has also been shown to outperform the classical glass pH-sensor. In this new methodology, students learn to build the electrode, to calibrate it, and to measure its sensitivity, repeatability, and time-response.

Real-time measurement of glucose using chrono-impedance technique on a second generation biosensor

Chrono-impedance technique (CIT) was implemented as a new transduction method for real time measurement of glucose in a biosensor system based in carbon paste (CP)/Ferrocene (FC)/glucose oxidase (GOx). The system presents high selectivity because the optimal stimulation signal composed by a 165mV DC potential and 50mV(RMS) AC signal at 0.4Hz was used. The low DC potential used decreased the interfering species effect and the biosensor showed a linear impedance response toward glucose detection at concentrations from 0mM to 20mM,with 0.9853 and 0.9945 correlation coefficient for impedance module (|Z|) and phase (Φ), respectively. The results of quadruplicate sets reveal the high repeatability and reproducibility of the measurements with a relative standard deviation (RSD) less than 10%. CIT presented good accuracy (within 10% of the actual value) and precision did not exceed 15% of RSD for high concentration values and 20% for the low concentration ones. In addition, a high correlation coefficient (R(2)=0.9954) between chrono-impedance and colorimetric methods was obtained. On the other hand, when two samples prepared at the same conditions were measured in parallel with both methods (the measurement was repeated four times), it should be noticed that students t-test produced no difference between the two mentioned methods (p=1). The biosensor system hereby presented is highly specific to glucose detection and shows a better linear range than the one reported on the previous article.

Glucose biosensor based on functionalized ZnO nanowire/graphite films dispersed on a Pt electrode.

We present a glucose biosensor based on ZnO nanowire self-sustained films grown on compacted graphite flakes by the vapor transport method. Nanowire/graphite films were fragmented in water, filtered to form a colloidal suspension, subsequently functionalized with glucose oxidase and finally transferred to a metal electrode (Pt). The obtained devices were evaluated using scanning electron microscopy, energy-dispersive x-ray spectroscopy, cyclic voltammetry and chronoamperometry. The electrochemical responses of the devices were determined in buffer solutions with successive glucose aggregates using a tripolar electrode system. The nanostructured biosensors showed excellent analytical performance, with linear response to glucose concentrations, high sensitivity of up to ≈17 μA cm(-2) mM(-1) in the 0.03-1.52 mM glucose concentration range, relatively low Michaelis-Menten constant, excellent reproducibility and a fast response. The detection limits are more than an order of magnitude lower than those achievable in commercial biosensors for glucose control, which is promising for the development of glucose monitoring methods that do not require blood extraction from potentially diabetic patients. The strong detection enhancements provided by the functionalized nanostructures are much larger than the electrode surface-area increase and are discussed in terms of the physical and chemical mechanisms involved in the detection and transduction processes.

Evaluation of Impedance Spectroscopy as a Transduction Method for Bacterial Biosensors

Biosensors are simple, feasible and cost-effective devices where biological specificity and selectivity, and electronic miniaturization are combined. Degrading microorganisms, such as the M7 species of Streptomyces genus, can specifically be used as biorecognition element, for lindane detection and quantification. Furthermore, electrochemical impedance spectroscopy is a non-destructive technique that allows evaluating bacterial activity by measuring conductivity changes in a culture medium. In this work, instrumental conditions were optimized to apply this method as transduction principle in bacterial biosensors. By means of electrochemical impedance spectroscopy, concentrations of chloride ions close to the environmental lindane values were measured. This is a suitable, simple and economical technique for use as a transduction method in biorecognition devices for organochlorine pesticides detection, particularly lindane.

Amplifier spurious input current components in electrode-electrolyte interface impedance measurements

BackgroundIn Impedance Microbiology, the time during which the measuring equipment is connected to the bipolar cells is rather long, usually between 6 to 24 hrs for microorganisms with duplication times in the order of less than one hour and concentrations ranging from 101 to 107 [CFU/ml]. Under these conditions, the electrode-electrolyte interface impedance may show a slow drift of about 2%/hr. By and large, growth curves superimposed on such drift do not stabilize, are less reproducible, and keep on distorting all over the measurement of the temporal reactive or resistive records due to interface changes, in turn originated in bacterial activity. This problem has been found when growth curves were obtained by means of impedance analyzers or with impedance bridges using different types of operational amplifiers.MethodsSuspecting that the input circuitry was the culprit of the deleterious effect, we used for that matter (a) ultra-low bias current amplifiers, (b) isolating relays for the selection of cells, and (c) a shorter connection time, so that the relays were maintained opened after the readings, to bring down such spurious drift to a negligible value. Bacterial growth curves were obtained in order to test their quality.ResultsIt was demonstrated that the drift decreases ten fold when the circuit remained connected to the cell for a short time between measurements, so that the distortion became truly negligible. Improvement due to better-input amplifiers was not as good as by reducing the connection time. Moreover, temperature effects were insignificant with a regulation of ± 0.2 [°C]. Frequency did not influence either.ConclusionThe drift originated either at the dc input bias offset current (Ios) of the integrated circuits, or in discrete transistors connected directly to the electrodes immersed in the cells, depending on the particular circuit arrangement. Reduction of the connection time was the best countermeasure.

Electronic continuous growth follow-up of microbial cultures

An apparatus for the measurement of bacterial growth is described. This instrument applies an alternate adequate current of two different frequencies through a pair of electrodes immersed in a cultured medium. It monitors, detects and quantifies the growth of microorganisms based on the measurement of the impedance across the two electrodes, from which medium conductivity and interface electrode impedance changes can be dissected out. The variation of these parameters are proportional to bacterial growth. They are processed by this computer controlled apparatus and displayed on a monitor showing bulk resistance and electrode impedance variations as time course events.

A polymer chip-integrable piezoelectric micropump with low backpressure dependence

We describe a piezoelectric micropump constructed in polymers with conventional machining methods. The micropump is self-contained and can be built as an independent device or as an on-chip module within laminated microfluidic chips. We demonstrate on-chip integrability by the fabrication and testing of an active micromixer with two pumps. Average flow rates from sub-μl min−1 to 300 μl min−1 can be obtained with low influence from the backpressure up to approximately 10 kPa. The micropump design allows potential use in low-cost disposable polymeric Lab on a Chip devices.