Anju Chadha

Estimation of triglyceride concentration is important for the health and food industries. Use of solid state biosensors like Electrolyte Insulator Semiconductor Capacitors (EISCAP) ensures ease in operation with good accuracy and sensitivity when compared to conventional sensors. In this paper we report on packaging of miniaturized EISCAP sensors on silicon. The packaging involves glass to silicon bonding using adhesive. Since this kind of packaging is done at room temperature, it cannot damage the thin dielectric layers on the silicon wafer unlike the high temperature anodic bonding technique and can be used for sensors with immobilized enzyme without denaturing the enzyme. The packaging also involves a teflon capping arrangement which helps in easy handling of the bio-analyte solutions. The capping solves two problems. Firstly, it helps in the immobilization process where it ensures the enzyme immobilization happens only on one pit and secondly it helps with easy transport of the bio-analyte into the sensor pit for measurements. © 2010 Copyright SPIE - The International Society for Optical Engineering.

Measurement in liquid media is a major challenge in real-time detection using resonant cantilevers. This is addressed in the present study by fabricating sub-micron thick cantilevers followed by functionalization for biomolecule detection. The fabricated cantilever resonator beams of thickness 165 nm were used for measurements in two systems: (i) human immunoglobulin (HIgG) as the antibody on the cantilever sensing mouse immunoglobulin (MIgG) as corresponding antigen, and (ii) detection of triglyceride (TG) based on the enzymatic hydrolysis with lipase, using tributyrin as a model. In both cases, the beams were functionalized for covalent bonding of the protein receptor. The label-free detection was carried out by measuring the shift in resonance frequency at higher modes, using a laser Doppler vibrometer in liquid and in air. The calibration showed a linear correlation between the bioanalyte concentration and change in the resonance frequency. Notably, detection of antigen mass as low as 434 ± 59fg and triglyceride concentration in the nM range with limit of detection as 7 nM in liquid interface was achieved, greatly improving the sensitivity of bioanalyte detection in liquid samples. Although frequency-based methods are highly sensitive, the issues with measurement liquid medium limit their application. In the present report, these issues were addressed by fabricating sub-micron thick cantilever beam, choosing an appropriate functionalization method without affecting the sensitivity, and measurement at higher modes. These have resulted in circumventing issues like damping and hydrodynamic loading thus improving its potential as real-time sensor.

We report the fabrication of potentiometric electrolyteinsulator capacitor (EISCAP) biosensors based on silicon and porous silicon (PS) substrates with oxide and stacked oxidenitride dielectrics. These biosensors have been calibrated for the detection and estimation of bioanalytes like tributyrin and urea, based on enzymatic reactions and have a linear detection range from 0.1 mM to 20 mM of the bioanalyte concentration. These improved sensitivity EISCAP sensors were used for the estimation of the total acid content in rancid butter, estimation of enzyme (Lipase) activity and to estimate total triglyceride levels in blood serum. © 2009 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Cantilevers immersed in liquid experience viscous damping and hydrodynamic loading. We report on the use of such cantilevers, operating in the dynamic mode with, (i) frequency sweeping and (ii) phase locked loop methods. The solution to reliability issues such as random drift in the resonant peak values, and interference of spurious modes in the resonance frequency spectrum, are explained based on the actuation signal provided and laser spot size. The laser beam spot size and its position on the cantilever were found to have an important role, on the output signal and resonance frequency. We describe a method to distinguish the normal modes from the spurious modes for a cantilever. Uncertainties in the measurements define the lower limit of mass detection (m min). The minimum detection limits of the two measurement methods are investigated by measuring salt adsorption from phosphate buffer solution, as an example, a mass of 14 pg was measured using the 14th transverse mode of a m × 100 μm × 1 μm silicon cantilever. The optimized measurement was used to study the interaction between antibody and antigen.

A modified electrolyte-insulator-semiconductor-capacitor (EISCAP) developed in-house was used for the determination of triglycerides and urea in blood, in addition to urea in milk. The sensitivity of the sensor was 55 ± 0.5 mV per unit change of pH. The EISCAP was optimized to operate at 25 °C in 0.25 mM phosphate buffer (pH 6), 1 M KCl and 1 mg lipase for triglycerides and in 1 mM Tris-HCl buffer (pH 7.4), 0.5 M KCl and 0.5 U urease for urea. Comparisons of the results obtained from the developed biosensor and clinical records gave correlation coefficients of 0.979 and 0.974 for blood triglycerides and blood urea respectively. Similarly, estimation of milk urea determined using the sensor in comparison with the spectrophotometric method gave a correlation coefficient of 0.992. The sensor gave reproducible and reliable results up to six months. © 2011 Elsevier B.V. All rights reserved.

We report fabrication of potentiometric biosensors with silicon for the estimation of triglycerides and urea based on enzymatic reactions. The sensor is an Electrolyte-Insulator-Semiconductor capacitor (EISCAP) that shows a shift in the measured CV with changes in the pH of the electrolyte. Enzyme mediated biological reactions involve changes in the pH of the electrolyte and an EISCAP can be effectively used for detection of biological compounds. Optimization of the conditions for the enzymatic reaction and calibration of the sensor are included. Effect of replacing silicon with porous silicon is discussed.

A novel, enzymatic, porous silicon (PS) based potentiometric method for estimating triglycerides is reported. Lipase, an enzyme, which hydrolyses triglycerides was immobilised on PS which was prepared from p-type (1 0 0) crystalline silicon and was thermally oxidized. On hydrolysis the triglycerides result in the formation of fatty acids which causes a change in the pH of the solution. Enzyme solution-oxidized PS-crystalline silicon structure was used to detect changes in pH during the hydrolysis of tributyrin as a shift in the capacitance-voltage (C-V) characteristics. Optimisation of the conditions for the enzymatic reaction and calibration of the sensor are included. © 2002 Elsevier Science B.V. All rights reserved.

A novel composite porous silicon/polysilicon microcantilever for biosensing applications with enhanced sensitivity is reported. It is fabricated by surface micromachining of polysilicon cantilevers followed by the formation of the surface porous layer after release by Reaction Induced Vapor Phase Stain Etch. The microcantilevers with porous surface layer are characterized by their morphology that exhibits a dual macro and nanostructure for very effective immobilization of biomolecules. The current work focuses on the fabrication of composite porous silicon/polysilicon microcantilevers, characterization of their morphology and resonance frequency, as well as demonstration of improved immobilization of enzyme resulting in enhanced sensing of triglycerides. © 2009 IEEE.

Lipase from Pseudomonas cepacia was covalently immobilized on crystalline silicon, porous silicon and silicon nitride surfaces. The various stages of immobilization were characterized using FTIR (Fourier transform infrared) spectroscopy. The surface topography of the enzyme immobilized surfaces was investigated using scanning electron microscopy (SEM). The quantity of the immobilized active enzyme was estimated by the para-nitrophenyl palmitate (pNPP) assay. The immobilized lipase was used for triglyceride hydrolysis and the acid produced was detected by a pH sensitive silicon nitride surface as a shift in the C-V (capacitance-voltage) characteristics of an electrolyte-insulator-semiconductor capacitor (EISCAP) thus validating the immobilization method for use as a biosensor. © 2008 Elsevier B.V. All rights reserved.

A novel method for estimating triglycerides is reported. Porous silicon, prepared from p-type (100) crystalline silicon was thermally oxidized and used to immobilise lipase, an enzyme, which hydrolyses triglycerides resulting in the formation of fatty acids. This causes a change in the pH of the solution. Enzyme solution-oxidized porous silicon-crystalline silicon structure was used to detect changes in pH during the hydrolysis of tributyrin as a shift in the capacitance-voltage (C-V) characteristics. Detailed calibration of the sensor is included. © 2001 Elsevier Science B.V.