Barbara Enderle

Polymer-based, flexible glutamate and lactate microsensors for in vivo applications

We present a flexible microsensor, based on a polymer substrate, for multiparametric, electrochemical in vivo monitoring. The sensor strip with a microelectrode array at the tip was designed for insertion into tissue, for fast and localized online monitoring of physiological parameters. The microsystem fabrication on a wafer-level is based on a polyimide substrate and includes the patterning of platinum microelectrodes as well as epoxy and dry-film-resist insulation in a cost-effective thin-film and laminate process. A stable, electrodeposited silver/silver chloride reference electrode on-chip and a perm-selective membrane as an efficient interference rejection scheme are integrated on a wafer-level. Amperometric, electrochemical, enzyme-based biosensors for the neurotransmitter L-glutamate and the energy metabolite L-lactate have been developed. Hydrogel membranes or direct cross-linking as stable concepts for the enzyme immobilization are shown. Sensor performance including high selectivity, tailoring of sensitivity and long-term stability is discussed. For glutamate, a high sensitivity of 2.16 nAmm(-2) µM(-1) was found. For lactate, a variation in sensitivity between 2.6 and 32 nAmm(-2)mM(-1) was achieved by different membrane compositions. The in vivo application in an animal model is demonstrated by glutamate measurements in the brain of rats. Local glutamate alterations in the micromolar range and in nanoliter-range volumes can be detected and quantified with high reproducibility and temporal resolution. A novel, versatile platform for the integration of various electrochemical sensors on a small, flexible sensor strip for a variety of in vivo applications is presented.

Multiparametric, Flexible Microsensor Platform for Metabolic Monitoring \(In~Vivo\)

In this paper, we present a multiparametric microsensor platform for metabolic monitoring in vivo. In contrast to silicon or ceramic-based systems, the flexible, implantable polymer-based sensor strip is manufactured in a cost-effective hybrid of thin-film and laminate technology in a wafer-level process. Flexibility allows easy handling and placement in soft tissue. It comprises a microelectrode array for up to four electrochemical, amperometric micro-(bio)sensors, and an integrated reference electrode. The energy metabolism parameters glucose, lactate, and oxygen as well as the neurotransmitter glutamate were measured. The sensors allow dynamic, highly sensitive, localized, long-term, online measurement of up to four metabolic parameters with a single device. The reliable analytical performance of the sensors, stability of the reference electrode, and the multiparametric measurement are shown. The sensor can be inserted directly into the tissue for in vivo applications.

Sensor Access to the Cellular Microenvironment Using the Sensing Cell Culture Flask

The Sensing Cell Culture Flask (SCCF) is a cell culture monitoring system accessing the cellular microenvironment in 2D cell culture using electrochemical microsensors. The system is based on microfabricated sensor chips embedded in standard cell culture flasks. Ideally, the sensor chips could be equipped with any electrochemical sensor. Its transparency allows optical inspection of the cells during measurement. The surface of the sensor chip is in-plane with the flask surface allowing undisturbed cell growth on the sensor chip. A custom developed rack system allows easy usage of multiple flasks in parallel within an incubator. The presented data demonstrates the application of the SCCF with brain tumor (T98G) and breast cancer (T-47D) cells. Amperometric oxygen sensors were used to monitor cellular respiration with different incubation conditions. Cellular acidification was accessed with potentiometric pH sensors using electrodeposited iridium oxide films. The system itself provides the foundation for electrochemical monitoring systems in 3D cell culture.

A novel, multiparametric, flexible microsensor for metabolic monitoring in vivo

We present a new, multiparametric microsensor for metabolic monitoring in vivo. In contrast to silicon or ceramic based systems, the flexible, implantable, polymer sensor strip is manufactured in a cost-effective hybrid of thin-film and laminate technology in a wafer-level process. Flexibility allows easy handling and placement in soft tissue. It comprises electrochemical, amperometric micro-(bio)sensors for the energy metabolism parameters lactate and oxygen, the neurotransmitter glutamate as well as an integrated reference electrode. This allows dynamic, highly sensitive, localized, long-term online measurement of multiple metabolic parameters with a single device. The reliable analytical performance of the sensors has been proven. The sensor can be inserted directly into the tissue in clinical applications.

BioMEMS for the electrochemical detection of troponin I

In this paper, a foil-based BioMEMS for the electrochemical detection of troponin I, a marker for myocardial infarction is presented.

Foil-based BioMEMS for electrochemical capillary immunoassay for hepatitis serology

A foil-based BioMEMS for electrochemical capillary immunoassay for hepatitis serology is presented. Immobilization of the hepatitis antigen utilizing capillary forces and special stopping barriers in foil based microsystems and the application of it in fast immunoassays will be shown. The measurement starts with the addition of the antibody containing human serum sample. Subsequently the signal antibody labelled with glucose oxidase binds to the hepatitis B antibody and converts glucose to hydrogen peroxide, which is oxidized at the electrode in the measurement cell. The assay is very sensitive and designed to differentiate between hepatitis B negative (below 10 U/l) and positive serum. The present limit of detection of immunoassays in the BioMEMS is 20 atto-moles.

Amperometric micro-immunosensor for rapid Substance-P quantification in biological fluids

A capillary immunosensor-chip has been developed for fast determination of substance-p (SP) levels in biological fluids. The microdevice approach combines the advantages of the microfluidic capillary for implementation of a competitive immunoassay retaining sensitivity and specificity, with the highly sensitive electrochemical detection of GOD-tracer by flow-injection analysis. The assay exhibits a detection limit of 10 pg/mL. The immunosensor has been successfully used for SP quantification in blood plasma and saliva of healthy human volunteers: 110 pg/mL in plasma and 200 pg/mL in saliva were measured. The developed miniaturized immunoassay showed good precision, high sensitivity, acceptable stability and reproducibility with consistent results comparable to those obtained by the commercial assay kit. According to this, the proposed on-chip immunosensing method may have a potential applicability in medical diagnostics of major diseases.

Interstitial Glucose and Lactate Levels Are Inversely Correlated With the Body Mass Index: Need for In Vivo Calibration of Glucose Sensor Results With Blood Values in Obese Patients

Background: Continuously measured glucose and lactate levels in interstitial fluid (ISF) may markedly differ from their respective blood levels. Methods: Combining microdialysis with a bioanalytical microsystem, the interstitial glucose and lactate concentrations of eight male volunteers with different body mass index (BMI) were monitored during a 2-fold glucose tolerance test over the period of three hours. Results: Significant correlations were found between abdominally measured sensor results and reference measurements (R2 = .967 for glucose and R2 = .936 for lactate, P < .05). The physiological delay of the abdominally observed glucose appearance in the ISF correlated positively with the BMI (R2 = .787, P < .05). The relative in vivo recovery of glucose and lactate was inversely proportional to the BMI of the volunteers (R2 = .540 for glucose, R2 = .609 for lactate, P < .05). One subject with a BMI of > 34 kg/m2 showed abdominally as well as the antebrachially significantly reduced tissue glucose values compared to blood glucose values (P < .001). Conclusions: A very good correlation between abdominally measured sensor results and the results of the reference method verified the reliability of the BioMEMS. The abdominally measured glucose level in ISF decreased significantly with increasing BMI. Therefore, an in vivo calibration of glucose levels in ISF with blood levels seems to be necessary especially in markedly obese subjects.

Amperometric Monitoring of Substance-P Levels in Biological Fluids

In this paper, a foil-based microfluidic chip (Fig. 1) for rapid determination of the neuropeptide Substance-P (SP) level in biological fluids is presented. Compared with standard ELISA methods, the miniaturization allows reducing the assay preparation and the measurement time. Due to the high variability levels and the extremely poor stability of SP in blood plasma, such improvement is crucial in point of care diagnostics of SP-induced inflammatory and immune diseases. The measurement is realized by a competitive immunoassay based on the sequenced immobilization of primary and secondary capture antibody into the capillary channel of the sensor (Fig. 2). Subsequent oxidation of hydrogen peroxide generated by the SP-labeled glucose oxidase mixed with the plasma sample resulted in a current signal inversely proportional to SP concentration. The measured 160 pg/ml in human sample (Fig. 3) was repeatable and reproducible by the corresponding optical ELISA kit on microtiter plates. Combined sensitive detection limit of 10 pg/ml and two minutes response demonstrates that this on-chip immunosensing technique is ideally suited for clinical application.

Bioanalytische Mikrosysteme für die klinische Chemie

Für den Schnelltest von Lebertransaminaseaktivitäten wurde ein miniaturisiertes Analysesystem mittels Hybridtechnologie hergestellt. Dabei wurde die Mikrofluidik mittels Leiterplattentechnologie hergestellt und mit in Dünnschichttechnologie hergestellten Glutamatbiosensoren kombiniert. Die Funktion des Systems wurde in wäßrigen GOT (AST)und OFT (ALT)-Lösungen demonstriert. Das erforderliche Probenund Reagenzvolumen war jeweils 70 fxl, die Dauer eines Tests betrug 4,5 Minuten. Das miniaturisierte System hat äußere Abmessungen von 42 x 22 x 1,5 mm^ und ein inneres Gesamtvolumen von 11 ßl.