The present disclosure relates to optical detectors.
Fluorescence spectroscopy is one of the most sensitive detection techniques to quantify molecules. This is because the measurement is done in a dark background and the light source is incident with an off-axis angle. Furthermore, fluorescence intensity is not dependent on the path length of the sample, which is a limitation to absorbance spectroscopy. There are two methods of doing fluorescence spectroscopy, time resolved and phase modulation.
In a conventional phase modulation system, the sample is illuminated with a modulated light source where the frequency is chosen based on the lifetime of the fluorophores in the sample. The light from the sample is detected by a photomultiplier tube (PMT) and the phase and amplitude of the output signal are compared to those of the light modulation signal.
Fluorescence lifetime is one of the most robust fluorescence parameters and is used, for example, in applications where it is necessary to discriminate against the high background fluorescence from biological samples. The fluorescence lifetime is the average decay time of a fluorescent molecule from its excited state to ground state by emitting photons. As can be seen in
Where I0 is the original value of the excitation state and τ is the lifetime. The lifetime is defined as the time of the excited intensity to decay to 1/e or 36.79% of its original value.
The inventors have appreciated that at least some of the problems associated with known methods of spectroscopy can be overcome by using lock-in detection on an application specific integrated circuit (ASIC) chip. Lock-in detection is a method that is capable of extracting signal amplitudes and phases in extremely noisy environments. The working principle of a lock-in measurement is by extracting a signal at a defined frequency that is identical to the reference modulated frequency and eliminating all other frequency components. This approach utilises homodyne detection and band-pass filtering to measure the signal's amplitude and phase that is relative to the reference frequency. By doing so, the signal of interest can be measured accurately and a high SNR can be achieved.
Lock-in detection could increase SNR in spectroscopic measurements and be used to perform fluorescence lifetime measurements. A disadvantage of existing lock-in detection is the bulky electronics. Existing systems have bulky bench top equipment with discrete components that are expensive.
According to a first aspect of the present invention there is provided an optical detector on an application specific integrated circuit (ASIC) comprising at least one photodiode for receiving incident light and configured to provide respective diode signals; a modulator configured to provide an AC drive signal and to provide a reference signal associated with the AC drive signal; and a lock-in amplifier configured to receive the diode signals from the at least one photodiode and to receive the reference signal from the modulator, and to determine a phase and/or an amplitude of the diode signals using the reference signal. The modulator is typically a light source modulator configured to drive a light source with the AC drive signal. For some applications the modulator can be configured to drive a heating element (e.g. a heating coil) coupled to the sample or to apply a voltage directly across the sample in order to excite the sample and cause it to luminesce.
The optical detector can be included as a single integrated system to obtain fluorescence lifetime using lock-in detection and phase modulation fluorescence with improved SNR. For spectroscopic measurements, the SNR may be improved by a few orders of magnitude compared to a DC optical detector (i.e. an optical detector without frequency modulation or lock-in detection). Compared to existing lock-in detection systems, the optical detector has the advantage of fewer standalone or individual components (such as PMTs), which allows the optical detector to be made more compact, improve alignment robustness, and can especially reduce noise. For example, the product package of the ASIC chip comprising the optical detector may have dimensions in the following ranges: Width=2 mm to 5 mm; Length=2 mm to 5 mm; and Height=0.2 mm to 2 mm. The product package may comprise a light source such as an LED, or the light source can be provided separately.
The optical detector is typically a spectrometer. One or more of the at least one photodiodes typically comprise a colour filter, to be sensitive to a particular colour (i.e. frequency range). For example, dichroic filters having a FWHM of about 5 nm to 40 nm may be used. The at least one photodiode may comprise a mixture of filtered photodiodes and clear (un-filter) photodiodes. Two or more photodiodes may comprise the same colour filter. The ASIC chip may integrate filters into standard CMOS silicon via Nano-optic deposited interference filter technology. Using the optical detector with specific dichroic filters can allow the system to discriminate the measurement of the specific wavelength of the fluorescence emission, while rejecting any stray light coming from the excitation light source.
The optical detector typically comprises an array of photodiodes. That is, the at least one photodiode is typically a plurality of photodiodes arranged in an array. The amplifier of the optical detector may comprise a multiplexer configured to multiplex the diode signals from the plurality photodiodes. For example, the plurality of photodiodes may be an 8×8 array, providing 64 individual signals, while the ASIC may only comprise, for example, 16 physical channels to process the signals. The multiplexer can then multiplex the 64 signals to 16 signals, which can then be processed in parallel on the 16 channels. Each photodiode can be individually lock-in detected to determine its signal strength (amplitude) and phase. Alternatively, signals from groups of similar photodiodes (e.g. having the same colour filter) may be processed as one signal, wherein it is assumed that the phase of signals from photodiodes within the group is substantially the same.
The optical detector may use analogue mixing (i.e. mixing of analogue signals) of the pixel diode signal with the driver reference, thereby determining via normal lock-in detection the amplitude and phase of the (each) pixel diode signal. The lock-in amplifier may comprise: a mixer configured to mix the reference signal with an output from the multiplexer to provide demodulated signals; a second multiplexer coupled to the first multiplexer and configured to multiplex the demodulated signals; and one or more analogue to digital converters (ADCs) configured to convert the demodulated signals to digital signals. The amplifier provides analogue mixing and lock-in detection, by demodulating the analogue signals from the diodes before they are digitizing by the ADCs.
Alternatively, the optical detector may be configured to use digital mixing (i.e. mixing of digital signals) of the photodiode signal with the driver reference to determine, via digital lock-in detection, the amplitude and phase of the (each) pixel diode signal. In this case, the amplifier may comprise one or more analogue to digital converters (ADCs) configured to convert an output from the multiplexer into digital signals; a mixer configured to mix the digital signals with the reference signal to provide demodulated signals; and a second multiplexer coupled to the first multiplexer and configured to multiplex the demodulated signal. The lock-in amplifier thereby provides digital demodulation and digital lock-in detection.
The first and a second multiplexers can be coupled to select each photodiode signal (or each group/set of photodiode signals) for demodulation by the mixer and then to bring the demodulated signal to the data buffer or MCU. The optical detector can comprise one or more further lock-in amplifiers connected in parallel and configured to determine the phase and/or amplitude of signals using the reference signal. The set of MUX, MIX, MUX, and ADC can be made on the ASIC in multitudes (duplo, triple, . . . multiple) in parallel to increase the speed of measurement and data analysis.
The light source may comprise at least one of a light emitting diode (LED), a lamp (e.g. a light bulb), and/or a vertical cavity surface emitting laser (VCSEL). The light source modulator may comprise a programmable maximum duty cycle and frequency oscillator, or may comprise an analogue current/amplitude modulator. The light source modulator may be configured to perform pulse width modulation (PWM). The light source modulator can provide an AC drive signal to the light source in order to provide modulated light. The AC may optionally have a DC-offset. The AC drive signal can be one of a sine wave, a square wave, and a triangular wave. In principle having AC-output from zero-max and may have an offset. Other waves, including stochastic, pseudo-random and quasi random drive signals may also be used, since the lock-in amplifier is provided with an associated reference signal, which enables demodulation. The AC drive signal may have a frequency in the range of typically 2 Hz to 10 MHz, and the reference signal has the same frequency as the drive signal. The large frequency range provided by the light source modulator may be beneficial for spectroscopic analysis of a large variety of samples (e.g. different fluorophores having different fluorescence lifetimes). In absorbance and reflectance mode, the optical detector can provide broad spectral ranges for identifying compounds.
The components of the optical detector, i.e, the light source modulator, the photodiodes and lock-in amplifier, are integrated on a single ASIC chip, such as an integrated CMOS chip. The ASIC can be configured to be powered by a supply voltage (VDD) in the range of 1.6 V to 2.0 V, for example 1.8 V. The low voltage can reduce power consumption. The small form factor of the ASIC may therefore be especially suited for Point-of-Care settings, wearables and battery driven, low power devices. It can have improved cost, low noise through minimized parasitics, and a small form factor.
In addition to increasing SNR by eliminating noise, the optical detector can allow the differentiation of different fluorescence lifetimes. This may be particularly useful in medical devices, as general human biofluid samples have different auto-fluorescence emission when excited in the UV to visible range. Using a phase modulation technique with the optical detector, the different phase shifts and modulation shifts provide the fluorescence lifetime. Therefore, the auto-fluorescence can be determined and only the corresponding phase/modulation shift to the target fluorophore picked out in the measurements. Furthermore, the optical detector can be used in a multiplexing technique, when more fluorophores with the same emission wavelength are used. The phase modulation technique allows the differentiation of the different fluorophores.
The invention integrates lock-in detection with photodiodes into a single ASIC chip being a spectral sensor chip. Compared to dc-spectral sensors, the invention can add sensitivity and higher dynamic range. The chip can provide the modulation in the driver currents for illumination devices such as LEDs, (miniaturized) lamps and VCSELs, whereas classically choppers where applied. Note that each pixel of the diode detector array is demodulated for response in amplitude and phase. Each pixel diode may contain an optical filter or not.
According to a second aspect of the invention there is provided a system for performing spectroscopic measurements of a sample comprising: means for exciting the sample; and an optical detector according to the first aspect of the invention arranged such that the at least one photodiode receives light from the sample when in use. The means for exciting the sample may comprise one of a light source, a heating element (e.g. current carrying coil) and electrodes for applying a voltage across the sample.
The system may further comprise a sample holder for holding the sample. The sample holder may comprise a lateral flow test strip comprising a test line, wherein the light source is configured to illuminate the test line. The optical detector can then be arranged such that the at least one photodiode receives light reflected from the test line or emitted by the test line.
The system can be configured to be used in applications measuring at least one of reflectance, transmission/absorbance, and fluorescence or luminescence.
The optical detector can be housed in a product package having dimensions of about 2 mm×3 mm×1 mm (width×length×height), whereby ‘about’ indicates a variance of 10%. The miniaturised nature of the chip allows for a very small product package compared to existing solutions. The means for exciting the sample (e.g. light source, heater, voltage source) may be located outside the product package and is driven by the ASIC. For example, the light source may be provided as a separate module that is connected to the ASIC inside the product package.
According to another aspect of the invention there is provided a method of performing spectroscopic measurements using an optical detector according to the first aspect. The step of using the optical detector may comprise driving a light source with the AC drive signal from the light source modulator; illuminating a sample with the light source; receiving with the at least one photodiode light reflected by or emitted from or transmitted through the sample; and using the lock-in amplifier to determine the phase and/or amplitude of the light received by the at least one photodiode. The step of using the lock-in amplifier may comprise mixing the at least one diode signal from the at least one diode with the reference signal from the light source modulator. One or more of the at least one photodiode may have an optical filter for a specific wavelength and with a specific bandwidth.
According to a further aspect of the invention there is provided a method of determining the amplitude and/or phase of light using an optical detector on an application specific integrated circuit (ASIC), comprising driving a light source with an AC drive signal from a light source modulator; illuminating a sample with the light source; receiving with at least one photodiode light reflected or emitted from the sample; receiving at a lock in amplifier at least one diode signal from the at least one diode and a reference signal associated with the AC drive signal from the light source modulator; and using lock-in detection to determine the phase and/or amplitude of the at least one diode signal from the at least one diode signal and the reference signal.
There are two methods of performing time-resolved fluorescence measurements, time domain and frequency domain. In the time domain, the sample with fluorophores is excited with a short pulse of light and the bandwidth of the pulse is shorter than T. Then, the time dependent intensity is measured over a period until 1/e of the original value at t=0 to obtain the lifetime or taking the slope of a plot log I(t) vs t.
The other method of measurement is frequency domain or phase modulation technique. In this technique, the sample with fluorophores is excited with an intensity modulated light source and normally in the form of a sine wave to avoid harmonic frequencies that could generate noise. The intensity of the light source has to be modulated at a frequency that is comparable to the reciprocal of the lifetime r. By doing this, the emission of the fluorescence is forced to respond at the same modulation frequency. However, due to the lifetime of the fluorescence, there is a delay in time relative the modulated excitation. This delay can be seen in
In addition, another effect due to fluorescence lifetime is the peak-to-peak height of the emission relative to modulated excitation,
The decrease in modulation is because some of the fluorophores that are excited are still emitting photons when the excitation is at a minimum, which is due to the quantum yield of the common fluorophores is less than 100%. This effect is named demodulation and can also be used to calculate the fluorescence lifetime:
In addition, in medical devices that normally measure biological samples that have auto-fluorescence in the visible range, using a phase modulation technique, each separate lifetime components can be separated to obtain the correct signal from the designated fluorophores that are used for detection.
In one embodiment, an optical detector has 11 channels for spectral identification and colour matching applications used in mobile devices. The optical detector comprises a light source modulator for driving a light source and a lock-in amplifier connected to the photodiodes and connected to the light source modulator for demodulating the diode signals. The optical detector may be configured to measure the spectral response defined in the wavelengths from approximately 350 nm to 1000 nm. Six channels can be processed in parallel by independent ADCs while the other channels are accessible via a multiplexer. Eight optical channels associated with 16 photodiodes (4×4 photodiode array) cover the visible spectrum (VIS). One channel can be used to measure near infra-red (NIR) light, and another channel is associated with a photodiode without filter (“clear”). The optical detector may also integrate a dedicated channel to detect 50 Hz or 60 Hz ambient light flicker. The flicker detection engine can also buffer data for calculating other flicker frequencies externally. The NIR channel in combination with the other VIS channel may provide information of surrounding ambient light conditions (light source detection). The optical detector can be synchronized to external signals via a general purpose input/output (GPIO) pin.
In one embodiment, the ASIC chip integrates filters into standard CMOS silicon via Nano-optic deposited interference filter technology. A built in aperture is provided to control the light entering the photodiode array. Control and spectral data access is implemented through a serial I2C interface. The device can have an ultra-low profile package with dimensions of 3.1 mm×2 mm×1 mm.
Embodiments of the optical detector can be implemented in a lateral flow test. A typical lateral flow test will have two measureable lines, test and control lines. The test line gives information on the different concentrations of analytes as a function of fluorescence intensity. Normally, to measure this in a lateral flow test, a reflection mode is used.
In general, embodiments of the optical detector can be advantageously used for bio-diagnostics in lateral flow tests. Embodiments can improve the sensitivity, especially when configured to operate in fluorescence mode. The small package size and improved robustness can enable implementation of the optical detector in hand held systems, which has previously not been possible. The detection can be done in the frequency domain as well as in the time domain.
Embodiments of the optical detector can be used for miniaturized reflectance applications. For example, such a spectrometer can be used for color measurements, for example, to measure skin tone and/or to measure moisture of samples e.g. grain, beans, etc. The spectrometer can provide faster results and shorter integration time. The spectrometer may also be used for measuring smaller areas, which can be particularly useful for samples that are not homogeneous.
Embodiments of the optical detector can be used for miniaturized scatter measurements, and can be used in a particle sensor and/or smoke sensor. The optical detector can provide an increased dynamic range and greater sensitivity in order to detect smaller concentrations of particles as well as smaller particles. The optical detector can be integrated in a small sensor module (e.g. due to the small form factor of the ASIC chip), which can make it particularly suitable for household appliances.
Other embodiments of the optical detector can be used for miniaturized Raman spectroscopy, for example to measure hydration.
An embodiment of the optical detector can be integrated in a vital sensor, configured to optically measure blood pressure with reduced noise compared to existing methods.
Although the invention has been described in terms of preferred embodiments as set out above, these embodiments are illustrative only and the claims are not limited to those embodiments. Those skilled in the art will be able to make modifications and alternatives in view of the disclosure which fall within the scope of the claims. Each feature disclosed or illustrated in the present specification may be incorporated in the invention, whether alone or in any appropriate combination with any other feature disclosed or illustrated herein.
Number | Date | Country | Kind |
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2007683.2 | May 2020 | GB | national |
Filing Document | Filing Date | Country | Kind |
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PCT/EP2021/063118 | 5/18/2021 | WO |