An Introduction to Chemical and Biological Sensors Denise Wilson, Associate Professor Department of Electrical Engineeri

1. An Introduction to Chemical and Biological Sensors Denise Wilson, Associate Professor Department of Electrical Engineering University of Washington NSF/RISE Workshop/Short Course Development and Study of Advanced Sensors and Sensor Materials July 10, 2006

2. Outline • Classes of Chemical and Biological Sensors • Single Stage Transduction (Solid State I) • Multiple Stage Transduction (Solid State II) • Single Stage Transduction (Optical I) • Multiple Stage Transduction (Optical II) • Performance Metrics • Steady-state, short term • Steady-state, long term • Transient

3. Single Stage Transduction (Solid State I) • Class Definition: Biochemical Activity/Energy is directly converted to Electrical Energy • Electrochemical Sensors: Ion interaction (chemical signal) with a functionalized (partially or fully selective) material induces a separation of charges (electrical signal) • Basic Electrochemical Cell: • Potentiometric: voltage is measured at zero current draw • Amperometric: current is measured at constant voltage • Variations on the Electrochemical Cell: • FET (Field Effect Transistor) Based Devices • LAPs (Light addressable potentiometric sensors) • Conductivity Based Sensors (Chemiresistors): adsorption or absorption of molecules onto a functionalized (partially or fully selective) material induces a change in resistance

4. Measurement Circuit Working Electrode Reference Electrode Single Stage Transduction (Solid State I) • Basic Electrochemical Cell • Basic Operation: • Ions exchange electrons with the metal electrode: • With the reference electrode: in a controlled and constant way • With the working electrode: in a manner dependent on analyte concentration • The separation of charges is the electrical signal that represents the (bio) chemical signal at the electrode(s)

5. Single Stage Transduction (Solid State I) • Basic Electrochemical Cell: Signal Measurement • Potentiometric: In the presence of zero current, the voltage induced by the separation of charges (between working and reference electrode) is measured. • Governed by the Nernst Equation • Voltage = E1o - E2o + (RT/nF) ln ([C1]/[C2]) • E1o = standard electrode potential of ion interacting with electrode 1 • E2o = standard electrode potential of ion interacting with electrode 2 • C1 = concentration of ion 1 at electrode 1 • C2 = concentration of ion 2 at electrode 2 • Demonstrates no scaling penalty (same sensitivity for smaller devices) • Reference electrode materials and surrounding solutions are chosen for stability (eg. Ag/AgCl) • Working electrode materials are designed or chosen for analyte selectivity • Selectivity controlled through: electrode design or permeable membranes

6. Single Stage Transduction (Solid State I) • Basic Electrochemical Cell: Signal Measurement • Amperometric: In the presence of a controlled voltage, the current between working and reference electrodes is measured. • Ions are consumed in the measurement process (non-equilibrium) • Current: • Dependent on conductivity of solution • Responds only to ions whose redox potential < applied potential • Possesses inherent scaling (miniaturization) penalty that reduces sensitivity and sensor performance. • Reference and working electrode materials chosen on same basis as potentiometric operation • Selectivity controlled through: • Electrode design or permeable membranes or applied voltage Current Applied Potential

7. Single Stage Transduction (Solid State I) • Variations on the Basic Electrochemical Cell: ChemFET • Potentiometric measurement is inherent in FET structure (zero gate current) • Ions interact with interface between gate and insulator layer • Induced charge is “seen” as field effect in underlying FET channel • Conductive gate is used to provide an “electrical” reference electrode • Example: • NMOSFET • Analyte of interest = negative ion • Increased concentration = higher induced negative surface charge which must be overcome for FET to conduct current = higher threshold voltage Conductive Gate Negative Ions FET Insulator n+ + + + + n+ Source Drain p Positive (induced) charge

8. Single Stage Transduction (Solid State I) • Variations on the Basic Electrochemical Cell: ISFET • Gate/Insulator combination layers that provide desired selectivity are limited to a few analytes (e.g. palladium/oxide for measuring hydrogen) • A wider selection of insulator layers with desired selectivity is possible. • Removal of the “gate” reintroduces need for a reference electrode • Reference electrode in same sensing solution introduces instability • Vulnerability of insulator layer to trapped charge introduces drift. • Example: • N-ISFET with a negative ion of interest (same basic operation as ChemFET) Reference Electrode Negative Ions FET Insulator n+ + + + + n+ Source Drain p Positive (induced) charge

9. Single Stage Transduction (Solid State I) • Variations on the Basic Electrochemical Cell: ENFET • Immobilized enzyme attached to surface of insulator in ISFET structure: • Increases sensitivity (through catalytic enzyme activity) • Increases selectivity (through inherent functionalization of selected enzyme) • Insulator is partially protected from trapped charge (by enzyme layer) • ENFET retains many of the disadvantages of the ISFET structure Reference Electrode Ions of interest Immobilized Enzyme FET Insulator n+ + + + + n+ Source Drain p Induced Charge

10. pH 7 PhotoCurrent pH 5 pH 9 Bias Voltage Reference Electrode I Insulator Induced Charge p Conductive Gate Transparent Substrate Single Stage Transduction (Solid State I) • Variations on the Basic Electrochemical Cell: LAPs • Light, rather than the FET, structure is used to amplify the induced charge from electrochemical interactions at the sensor surface • Requires an external light source and optics for operation • Eliminates need for drain and source electrodes • A single “gate” is required for an entire array of sensors • Active sensing area is determined by illumination area. AC Light Source

11. Single Stage Transduction (Solid State I): pH ChemFETs The sensors are designed and fabricated in a standard CMOS process (1.5 micron AMI). Various combinations of Silicon Nitride and Aluminum oxide, native to the CMOS process are used for pH sensitivity. These designs are extensions of designs and results initially presented by: J. Bausells, J. Carrabina, A. Errachid, A. Merlos, “Ion-sensitive field-effect transistors fabricated in a commercial CMOS technology,” Sensors & Actuators B – Chemical, 7 Sept 1999, B57(1-3), 56-62. pH Sensor (8 Total)

12. Single Stage Transduction (Solid State I): pH ChemFETs M2FETThe M2FET uses the second layer of aluminum in a standard CMOS process for pH sensitivity. Aluminum oxide grows on top of the aluminum layer when exposed to an oxygen environment, providing a pH sensitive layer that is then measured via the underlying field effect (MOS) structure

13. Single Stage Transduction (Solid State I): pH ChemFETs M2FET:The results show sub-microamp differences between the drain currents of the FET when operated in the linear (ohmic) region.These changes in drain current reflect changes in threshold voltage of the FET induced by accumulation of charge (OH- ions) on top of the aluminum oxide layer.

14. Single Stage Transduction (Solid State I): pH ChemFETs These sensors use the overlying passivation layer (Silicon Nitride) as the pH sensitive layer (the bottom structure is an interdigitated version of the top structure)

15. Single Stage Transduction (Solid State I) • Conductivity-Based Sensors (Chemiresistors) • Composite Polymer • Combination of stable, conductive material (e.g. carbon black) and • Chemically sensitive, insulating material • Molecule in sensing environment adsorbs into polymer sensor • Carbon black molecules “stretch” • Conductivity decreases; resistance increases • Highly sensitive to humidity (caused polymer to swell) http://nsl.caltech.edu/resnose.html

16. - - - - - - - - - Single Stage Transduction (Solid State I) • Conductivity-Based Sensors (Chemiresistors) • Metal-Oxide Sensors • Composed of metal-oxide and a catalyst to enhance selectivity • Oxygen naturally binds to the sensor surface • Binding processes extract electrons from the bulk • Analytes of interest in the sensing environment bind with Oxygen • Electrons are re-injected into the bulk • Conductivity increases (resistance decreases) • Type of oxygen (and resulting sensitivity) is dependent on temperature • Sensors are highly sensitive to humidity (which blocks binding sites) O2- O2- O2- O2- O2- O2- O2- O2- Tin-Oxide with no Analyte Present Tin-Oxide with Analyte Present

17. Multiple Stage Transduction (Solid State II) • Class Definition: Biochemical Activity/Energy is converted to an intermediate form of energy before being converted to Electrical Energy • Example: Acoustic Wave Devices • Basic Operation: Input path • an AC voltage is used to piezoelectrically convert • Electrical energy to • Mechanical energy (a pressure wave) propagating through a material • Basic Operation: Output path • Loss (attenuation) and • Delay (phase changes) of mechanical energy (the pressure wave) are affected by: • Viscoelastic properties of the material and • Overall mass of the active sensing area which is • Responsive to Biochemical binding and events on the surface of the sensor • Altered mechanical energy is converted back to Electrical Energy

18. Multiple Stage Transduction (Solid State II) • Classes of Acoustic Wave Sensors • Bulk Acoustic Wave: basic operation • Wave moves parallel to the surface of the sensor and • Permeates through the entire material • Bulk Acoustic Wave: characteristics • Poor Sensitivity: only surface wave responds to biochemical activity • Stable: on a quartz substrate • Relatively Inexpensive • Strong cross-sensitivity to temperature Direction of Propagation Electrodes Direction of Motion

19. Multiple Stage Transduction (Solid State II) • Classes of Acoustic Wave Sensors • Surface Acoustic Wave: basic operation • Wave moves perpendicular to the surface of the sensor and • Propagates only on the surface of the material • Surface Acoustic Wave: characteristics • Frequencies of operation on the order of 100’s of MHz and GHz • High Sensitivity: surface wave is large part of overall activity • Poorly suited to measurement in liquid (due to perpendicular propagation) • Relatively Expensive (high measurement overhead) Direction of Motion Electrodes Direction of Propagation

20. Multiple Stage Transduction (Solid State II) • Classes of Acoustic Wave Sensors • Shear Horizontal Surface Acoustic Wave • Wave moves parallel to the surface of the sensor and • Propagates only on the surface of the material • Difficult to fabricate: requires careful control of cutting piezoelectric material • Can be used in liquids • Shear Horizontal Acoustic Plate Mode • Electrodes on one side of a plate generate a • Pressure wave that is guided through the bulk of the material • To the other side of the plate where the sensing environment is located • Across the plate and back down to the electrodes on the opposite side of the plate • Wave moves parallel to the surface of the sensor • Electrodes and electronics are protected from the sensing environment • Moderate sensitivity: active area is not “optimal” percentage of overall sensor volume • Sample Application: detect mercury levels in drinking water

21. Single Stage Transduction (Optical I) • Class Definition: Optical energy (which is directly related to biochemical behavior) is directly converted to Electrical Energy • Luminescence Spectroscopy: light emitted by a system is a function of biochemical activity/energy • Chemiluminescence: emission light represents biochemical reactions • Fluorescence: emission light quickly (on the order of ns) indicates presence of particular molecules • Electrons are excited into singlet state (paired with opposite spin electron) by incoming light; radiate energy in the form of light upon returning to ground state • Phosphorescence: emission light slowly (on the order of ms) indicates presence of particular molecules • Electrons are excited into triplet states (paired with same spin electron) by incoming light; radiate energy in the form of light on returning to ground state but take longer because the triplet state is forbidden. • Fluorescence/Phosphorescence Spectroscopy • Input Energy: optical • Output Energy: optical converted to electrical through a photodetection system

22. Single Stage Transduction (Optical I) • Fluorescence Analysis Systems • Regardless of the “color” of light input, the output (emission) spectrum is the same. • The spectrum (“color”) of the input light influences only efficiency of the emission

23. White Light Source Conventional Optics Conventional Optics Single Stage Transduction (Optical I) Traditional Fluorescence Analysis Systems Sample Signal Processing Photomultiplier Tube Optical Dispersion

24. LED Array Fiber Optic Coupling Fiber Optic Coupling Single Stage Transduction (Optical I) Portable Fluorescence Analysis Systems Sample Signal Processing Photodiode Array Optical Dispersion

25. Single Stage Transduction (Optical I): Fluorescence Analysis Optimization Schemes for LED-based (miniaturized) systems In this case, input light is transmitted (inadvertently) to the output path; thus optimization involves reducing the overlap between excitation (input) light and the emission spectrum. Optimization criteria depend on system configuration and experimental constraints. (a) “Best” Configuration (b) “Worst” Configuration

26. Multiple Stage Transduction (Optical II) • Class Definition: Biochemical Activity/Energy is converted to Optical Energy before being converted to Electrical Energy • Surface Plasmon Resonance • Biochemical events/analytes of interest influence refractive index of the sensing environment very close (within 100nm) of the SPR surface (in the evanescent field). • Refractive index influences how input light is reflected back to the output path (as a function of incidence angle or wavelength, depending on the system configuration. • Transduction Path: • Light Energy is converted to • Surface Charge Oscillations (surface plasmons) • The unconverted Light Energy is then converted to • Electrical Energy (via Photodetection) in the output path

27. Sample Surface Plasma Wave Evanescent Wave Metal θinc Substrate Incident Light Reflected Light Optical Fiber w/ Cladding Gold Coating Exposed Core Multiple Stage Transduction (Optical II): SPR When the wave vector from a white light source closely matches that of the surface plasma wave at the metal-sample interface on the probe, reflected light is significantly attenuated (compared to the attenuation in the reference media where no analyte is present)

28. Multiple Stage Transduction (Optical II): SPR • The Big Picture • Why SPR? • Highly sensitive (10-4 to 10-6 RI units) • Very local (10-100nm from sensing surface) • Directly indicative (of interactions between sensor and environment) • Relatively unencumbered by sampling overhead (e.g. tagging, mixing, etc) • Readily referenced to compensate for background fluctuations (e.g. drift) • How is it used (SPR = transduction mechanism)? • Non-functionalized = bulk refractive index • Functionalized = specific analytes • The Full Spectrum of SPR-based instruments • User-Intensive, Single Measurements: Biacore • User-Intensive, Single Field Measurements: TI Spreeta (Chinowsky/Yee) • Distributed and Autonomous, Multiple Measurements: • Insertion-based probes • Compact signal processing • Streamlined, robust optical path

29. Multiple Stage Transduction (Optical II): SPR • Point of resonance can be detected at a • Particular angle (constant wavelength interrogation) • Particular wavelength (constant angle interrogation) • Constant Angle • Polychromatic light source at constant angle of incidence • Constant Wavelength • Monochromatic light source at different angles of incidence Constant Angle is chosen here for: inexpensive light source, easy alignment, and simpler, more compact configuration (= less overhead)

30. Multiple Stage Transduction (Optical II): SPR • Sampling Options: • In-line • “Dip” insertion-based probe • The probe configuration is : • easily replaced, easy to use • Less prone to sensor layer blocking, • but can be • more sensitive to ambient fluctuations • more susceptible to fouling

31. Air Multiple Stage Transduction (Optical II): SPR Increasing RI Raw Data (background overwhelms resonance) Referenced Data (Resonance is evident)

32. Multivariate Calibration Communication/ADC Overhead Measurement to Reference Ratio High Resolution Regression Low Resolution Regression Multivariate Calibration Multiple Stage Transduction (Optical II): SPR Approach #1 (Traditional) Software High Resolution Photodetection Approach #2 (Voltage-Mode, Partially Integrated) Software Integration Time Programming Low Resolution Photodetection “Flatlining” Reference Ratio

33. Approach #4 (Current-Mode, Fully Integrated) Software Multivariate Calibration Multivariate Calibration Dark Current Compensation Low Resolution Regression Low Resolution Photodetection “Flatlining” Current Scaling Approach #3 (Pulse-Mode, Fully Integrated) Software “Flatlining” Current Scaling Low Resolution Regression Low Resolution Photodetection Conversion to Pulse Mode Multiple Stage Transduction (Optical II): SPR

34. Multiple Stage Transduction (Optical II): SPR Approach #2 All Designs are mixed signal, fabricated in standard CMOS Approach #4 Approach #3

35. Pixel Analog Sampling Phototransistor Digital Control Multiple Stage Transduction (Optical II): SPR 15 pixel array fabricated on a 1cm2 die in the 1.5 micron AMI process through MOSIS 2mm

36. Multiple Stage Transduction (Optical II): SPR

37. Performance MetricsChemical and Biological Sensors • Steady State, Short Term • Sensitivity: Change in output/Change in Input • Chemiresistor; DR/DC • SPR: Dl/DRI • Resolution: • Input: smallest detectable change in input parameter • Output: smallest detectable change in output parameter • Offset: output in the presence of no input • Detection limit: minimum detectable change • Dynamic range: maximum - minimum detectable (input) signal • Signal to Noise Ratio: ratio of useful information to non-useful information at any given signal level • Selectivity: practical ratio of sensor response to (a) analyte of interest divided by response to (b) most significant interferent.

38. Performance MetricsChemical and Biological Sensors • Steady State, Long Term • Stability: Change in output for a constant input over a period of time several orders of magnitude greater than the response time of the sensor. • Drift: Loose term often used in place of stability -- means the non-monotonic change in sensor output over time in response to the same input conditions. • Transient • Hysteresis: maximum difference in output obtained when sensor input is increased as compared to sensor input decreased. • Response time: • Propagation delay: how long before a change in the sensing environment is reflected in a change in the sensor output (delay between a 50% change in the input and a 50% change in the output of the sensor is typically chosen) • Rise/Fall time: time required for the sensor output to rise (or fall) from 10% of its final output to 90% of its final output.