Patent Application
20230258862
The present application is directed generally to devices, methods and techniques for coupling optical energy into a device and distributing optical energy to many regions of the device. The optical device may be used for performing parallel, quantitative analysis of biological and/or chemical samples.
Detection and analysis of biological samples may be performed using biological assays (“bioassays”). Bioassays conventionally involve large, expensive laboratory equipment requiring research scientists trained to operate the equipment and perform the bioassays. Moreover, bioassays are conventionally performed in bulk such that a large amount of a particular type of sample is necessary for detection and quantitation.
Some bioassays are performed by tagging samples with luminescent markers that emit light of a particular wavelength. The markers are illuminated with a light source to cause luminescence, and the luminescent light is detected with a photodetector to quantify the amount of luminescent light emitted by the markers. Bioassays using luminescent markers conventionally involve expensive laser light sources to illuminate samples and complicated luminescent detection optics and electronics to collect the luminescence from the illuminated samples.
Some embodiments are directed to an integrated device comprising a plurality of waveguides, a grating coupler having a grating region, a plurality of output waveguides having varying widths and configured to optically couple with the grating coupler, and a plurality of optical splitters. At least one of the optical splitters is positioned between one of the plurality of output waveguides and at least two of the plurality of waveguides.
In some embodiments, the grating region comprises a plurality of gratings oriented substantially in a direction planar to a surface of the integrated device. In some embodiments, individual output waveguides of the plurality of output waveguides are arranged on a side of the grating region. In some embodiments, the plurality of output waveguides includes a first output waveguide and a second output waveguide, and wherein the first output waveguide is more proximate to a center of a side of the grating region than the second output waveguide and has a smaller width than the second output waveguide. In some embodiments, the plurality of output waveguides includes a first output waveguide and a second output waveguide, and wherein the first output waveguide is more proximate to an edge of a side of the grating region than the second output waveguide and has a smaller width than the second output waveguide. In some embodiments, a number of optical splitters between the second output waveguide and one of the plurality of waveguides is greater than a number of optical splitters between the first output waveguide and another of the plurality of waveguides. In some embodiments, the plurality of output waveguides and the plurality of optical splitters radially distribute from the grating region. In some embodiments, individual waveguides of the plurality of waveguides are arranged substantially perpendicular to gratings in the grating region. In some embodiments, at least one of the plurality of optical splitters is positioned less than 1 mm from the grating coupler.
In some embodiments, individual waveguides of the plurality of waveguides have a tapered dimension in a direction perpendicular to the direction of light propagation along one of the plurality of waveguides such that the tapered dimension is smaller at a location proximate to the grating coupler than at a distal location. In some embodiments, individual waveguides of the plurality of waveguides are positioned to optically couple with a plurality of sample wells. In some embodiments, at least one of the plurality of waveguides has a first thickness at a location overlapping with at least one sample well of the plurality of sample wells and a second thickness at a location non-overlapping with the at least one sample well, the first thickness being larger than the second thickness. In some embodiments, a surface of at least one sample well of the plurality of sample wells is in contact with a surface of a first waveguide of the plurality of waveguides. In some embodiments, at least one of the plurality of waveguides is a multimode waveguide configured to support propagation of a plurality of optical modes along the multimode waveguide. In some embodiments, power distribution along the multimode waveguide is broader in a first region that overlaps with at least one of the plurality of sample wells than in a second region separate from the first region. In some embodiments, individual waveguides of the plurality of waveguides are configured to support propagation of excitation energy having an evanescent field extending from one of the plurality of waveguides that optically couples with at least one sample well of the plurality of sample wells. In some embodiments, at least one sample well of the plurality of sample wells comprises a sidewall spacer formed on at least a portion of a sidewall of the at least one sample well. In some embodiments, the integrated device further comprises at least one metal layer, and wherein a surface of at least one of the plurality of sample wells is recessed from the at least one metal layer. In some embodiments, the integrated device further comprises a sensor configured to receive light from one of the plurality of sample wells. In some embodiments, a distance between the one sample well and the sensor is less than 10 micrometers. In some embodiments, the integrated device further comprises a metal layer formed on a surface of the integrated device, the metal layer having an opening that overlaps with an aperture of one of the plurality of sample wells. In some embodiments, a first waveguide of the plurality of waveguides is configured to optically couple with a portion of a first set of the plurality of sample wells, a second waveguide of the of the plurality of waveguides is configured to optically couple with a portion of a second set of the plurality of sample wells, and wherein an optical splitter of the plurality of optical splitters is positioned between the first set of sample wells and the second set of sample wells and is configured to optically couple to at least one of the first and second waveguides.
In some embodiments, the integrated device further comprises one or more photodetectors positioned to receive excitation energy that passes through the grating coupler. In some embodiments, the integrated device further comprises one or more photodetectors positioned to receive excitation energy that passes through a region proximate to the grating coupler.
Some embodiments are directed to a method of forming an integrated device comprising forming a plurality of waveguides, forming a grating coupler having a grating region, forming a plurality of output waveguides having varying widths and configured to optically couple with the grating coupler, and forming a plurality of optical splitters, wherein at least one of the optical splitters is positioned between one of the plurality of output waveguides and at least two of the plurality of waveguides.
In some embodiments, forming the grating coupler further comprises forming a plurality of gratings in the grating region, the plurality of gratings being oriented substantially in a direction planar to a surface of the integrated device. In some embodiments, forming the plurality of output waveguides further comprises forming individual output waveguides of the plurality of output waveguides arranged on a side of the grating region. In some embodiments, forming the plurality of output waveguides further comprises forming a first output waveguide and a second output waveguide. The first output waveguide is more proximate to a center of a side of the grating region than the second output waveguide and has a smaller width than the second output waveguide. In some embodiments, forming the plurality of output waveguides further comprises forming a first output waveguide and a second output waveguide. The first output waveguide is more proximate to an edge of a side of the grating region than the second output waveguide and has a smaller width than the second output waveguide. In some embodiments, forming the plurality of optical splitters further comprises forming a number of optical splitters between the second output waveguide and one of the plurality of waveguides that is greater than a number of optical splitters between the first output waveguide and another of the plurality of waveguides. In some embodiments, forming the plurality of output waveguides further comprises forming the plurality of output waveguides to radially distribute from the grating region. In some embodiments, forming a plurality of waveguides further comprises forming individual waveguides of the plurality of waveguides arranged substantially perpendicular to gratings in the grating region. In some embodiments, forming the plurality of waveguides further comprises forming the plurality of waveguides to have a tapered dimension in a direction perpendicular to the direction of light propagation along one of the plurality of waveguides such that the tapered dimension is smaller at a location proximate to the grating coupler than at a distal location.
In some embodiments, the method further comprises forming a plurality of sample wells, wherein individual waveguides of the plurality of waveguides are positioned to optically couple with the plurality of sample wells. In some embodiments, forming the plurality of waveguides further comprises forming at least one of the plurality of waveguides with a first thickness at a location overlapping with at least one sample well of the plurality of sample wells and a second thickness at a location non-overlapping with the at least one sample well, the first thickness being larger than the second thickness. In some embodiments, forming the plurality of sample wells further comprises forming a surface of at least one sample well of the plurality of sample wells in contact with a surface of a first waveguide of the plurality of waveguides. In some embodiments, forming the plurality of waveguides further comprises forming a multimode waveguide configured to support propagation of a plurality of optical modes along the multimode waveguide. In some embodiments, individual waveguides of the plurality of waveguides are configured to support propagation of excitation energy having an evanescent field extending from one of the plurality of waveguides that optically couples with at least one sample well of the plurality of sample wells. In some embodiments, forming the plurality of sample wells further comprises forming a sidewall spacer on at least a portion of a sidewall of at least one sample well of the plurality of sample wells. In some embodiments, forming the plurality of sample wells further comprises forming at least one metal layer and forming a surface of at least one of the plurality of sample wells recessed from the at least one metal layer. In some embodiments, the method further comprises forming a sensor configured to receive light from one of the plurality of sample wells. In some embodiments, a distance between the one sample well and the sensor is less than 10 micrometers. In some embodiments, a first waveguide of the plurality of waveguides is configured to optically couple with a portion of a first set of the plurality of sample wells, a second waveguide of the of the plurality of waveguides is configured to optically couple with a portion of a second set of the plurality of sample wells, and wherein an optical splitter of the plurality of optical splitters is positioned between the first set of sample wells and the second set of sample wells and is configured to optically couple to at least one of the first and second waveguides.
Some embodiments are directed to an integrated device comprising a plurality of first waveguides, a grating coupler having a grating region, a plurality of output waveguides having varying widths and configured to optically couple with the grating coupler, and a plurality of optical splitters. At least one of the optical splitters is positioned between one of the plurality of output waveguides and at least two of the plurality of first waveguides.
Some embodiments are directed to an integrated device comprising a first waveguide configured to optically couple with a portion of a first set of sample wells, a second waveguide configured to optically couple with a portion of a second set of sample wells, and an optical splitter positioned between the first set of sample wells and the second set of sample wells and configured to optically couple to at least one of the first and second waveguides.
In some embodiments, the integrated device further comprises at least one input waveguide configured to optically couple with the optical splitter. In some embodiments, the integrated device further comprises a grating coupler configured to optically couple with the at least one input waveguide. In some embodiments, gratings of the grating coupler are substantially parallel to the at least one input waveguide.
Some embodiments are directed to an integrated device comprising at least one sample well, and a waveguide configured to couple excitation energy to the at least one sample well, wherein the waveguide has a first thickness at a location overlapping with the at least one sample well and a second thickness at a location non-overlapping with the at least one sample well, and the first thickness is larger than the second thickness.
In some embodiments, the waveguide is configured to support propagation of excitation energy having an evanescent field extending from the waveguide. In some embodiments, the waveguide has a tapered dimension in a direction perpendicular to the direction of light propagation along the waveguide such that the tapered dimension is smaller at a location proximate to the grating coupler than at a distal location. In some embodiments, a surface of the at least one sample well contacts a surface of the waveguide. In some embodiments, the at least one sample well includes a plurality of sample wells in an array. In some embodiments, the at least one sample well is recessed from a metal layer of the integrated device. In some embodiments, the waveguide is a multimode waveguide configured to support propagation of a plurality of optical modes along the waveguide. In some embodiments, power distribution along the multimode waveguide is broader in a first region that overlaps with the at least one sample well than in a second region separate from the first region. In some embodiments, the first thickness is between 200 nm and 400 nm. In some embodiments, the second thickness is between 100 nm and 250 nm. In some embodiments, the waveguide is formed, at least in part, from a layer of silicon nitride. In some embodiments, the integrated device further comprising a sensor configured to receive emission energy emitted by a sample located in the at least one sample well. In some embodiments, a distance between the at least one sample well and the sensor is less than 10 micrometers. In some embodiments, a distance between the at least one sample well and the sensor is less than 7 micrometers. In some embodiments, a distance between the at least one sample well and the sensor is less than 3 micrometers. In some embodiments, the integrated device further comprises a metal layer formed on a surface of the integrated device, the metal layer having an opening that overlaps with an aperture of the at least one sample well. In some embodiments, the metal layer includes a first layer having aluminum and a second layer having titanium nitride, and wherein the first layer is proximate to the waveguide.
Some embodiments are directed to an integrated device comprising a metal layer disposed on a surface of the integrated device, the metal layer having a discontinuity, and at least one sample well having a top aperture corresponding with the discontinuity of the metal layer. A surface of the at least one sample well extends beyond the metal layer along a direction that is substantially perpendicular to the surface of the integrated device.
In some embodiments, the surface of the at least one sample well is positioned at a distance from the metal layer that is between 100 nm and 350 nm. In some embodiments, the at least one sample well comprises a sidewall spacer formed on at least a portion of a sidewall of the sample well. In some embodiments, the integrated device further comprises a waveguide distal the surface of the at least one sample well. In some embodiments, the waveguide comprises a slab and a raised region. In some embodiments, the waveguide is tapered. In some embodiments, the metal layer includes a first layer having aluminum and a second layer having titanium nitride, and the first layer is proximate to the waveguide. In some embodiments, a distance from the waveguide to the surface of the at least one sample well is less than 200 nm. In some embodiments, an opening in the metal layer corresponds to a grating coupler for the waveguide. In some embodiments, the waveguide is formed, at least in part, from a layer of silicon nitride. In some embodiments, the integrated device further comprises a sensor configured to receive emission energy emitted by a sample located in the at least one sample well. In some embodiments, a distance between the at least one sample well and the sensor is less than 10 micrometers. In some embodiments, a distance between the at least one sample well and the sensor is less than 7 micrometers. In some embodiments, a distance between the at least one sample well and the sensor is less than 3 micrometers.
Some embodiments are directed to a method of forming an integrated device comprising: providing semiconductor substrate having a dielectric film disposed on the semiconductor substrate; forming a waveguide having a slab and a raised region by partially etching a portion of the dielectric film; forming a top cladding such that the top cladding is in contact with the waveguide; forming a metal layer on a surface of the top cladding; and forming a sample well over the waveguide by etching the metal layer and a portion of the top cladding.
In some embodiments, forming the waveguide comprises a timed etch process. In some embodiments, forming the waveguide comprises an etch process using an etch stop layer. In some embodiments, forming the sample well comprises etching the top cladding until at least a portion of the waveguide is uncovered. In some embodiments, a distance between a bottom surface of the sample well and the waveguide is between 10 nm and 200 nm. In some embodiments, the method further comprises forming a spacer on at least a portion of a sidewall of the sample well. In some embodiments, the method further comprises forming the metal layer comprises forming a plurality of metal sub-layers. In some embodiments, the method further comprises etching a portion of the slab to form a ridge waveguide. In some embodiments, the method further comprises etching a portion of the slab to form a rib waveguide. In some embodiments, forming the waveguide further comprises forming a taper having a variable width.
Some embodiments are directed to an integrated device comprising a plurality of sample wells, a first optical waveguide configured to couple excitation energy to a first portion of the plurality of sample wells, a second optical waveguide configured to couple the excitation energy to a second portion of the plurality of sample wells, and a grating coupler configured to receive the excitation energy from an optical source positioned outside the integrated device, and to couple the excitation energy to the first optical waveguide and to the second optical waveguide.
In some embodiments, the integrated device further comprises one or more photodetectors positioned to receive excitation energy that passes through the grating coupler. In some embodiments, the integrated device further comprises one or more photodetectors positioned to receive excitation energy that passes in a region proximate to the grating coupler. In some embodiments, the grating coupler is a first optical grating coupler, and the integrated device further comprises a second optical coupler optically coupled to the first waveguide and configured to receive the excitation energy from the first waveguide and to couple the excitation energy to a photodetector positioned in the integrated device. In some embodiments, the first optical waveguide is configured to couple the excitation energy to the first portion of the plurality of sample wells via evanescent coupling. In some embodiments, the integrated device further comprises a metal layer disposed on a surface of the integrated device, where the plurality of sample wells is formed through the metal layer. In some embodiments, at least one of the plurality of sample wells comprises a bottom surface proximate to the first waveguide, the bottom surface being recessed through the metal layer. In some embodiments, the bottom surface is positioned at a distance from the metal layer that is between 100 nm and 350 nm. In some embodiments, the bottom surface is positioned at a distance from the first optical waveguide that is between 10 nm and 200 nm. In some embodiments, the metal layer includes an aluminum layer and a titanium nitride layer, and the aluminum layer is proximate to the first and second waveguides. In some embodiments, the optical grating comprises an etched region formed in a layer of silicon nitride. In some embodiments, at least one sample well of the plurality of sample wells comprises a sidewall spacer formed on at least a portion of a sidewall of the at least one sample well.
Various aspects and embodiments of the application will be described with reference to the following figures. It should be appreciated that the figures are not necessarily drawn to scale. Items appearing in multiple figures are indicated by the same reference number in all the figures in which they appear.
The inventors have recognized and appreciated that a compact, high-speed apparatus for performing detection and quantitation of single molecules or particles could reduce the cost of performing complex quantitative measurements of biological and/or chemical samples and rapidly advance the rate of biochemical technological discoveries. Moreover, a cost-effective device that is readily transportable could transform not only the way bioassays are performed in the developed world, but provide people in developing regions, for the first time, access to essential diagnostic tests that could dramatically improve their health and well-being.
The inventors have also recognized and appreciated that integrating a sample well and a sensor in a single integrated device capable of measuring luminescent light emitted from biological samples reduces the cost of producing such a device such that disposable bioanalytical integrated devices may be formed. Disposable, single-use integrated devices that interface with a base instrument may be used anywhere in the world, without the constraint of requiring high-cost biological laboratories for sample analyses. Thus, automated bioanalytics may be brought to regions of the world that previously could not perform quantitative analysis of biological samples.
A pixelated sensor device with a large number of pixels (e.g., hundreds, thousands, millions or more) allows for the detection of a plurality of individual molecules or particles in parallel. The molecules may be, by way of example and not limitation, proteins, DNA, and/or RNA. Moreover, a high-speed device that can acquire data at more than one hundred frames per second allows for the detection and analysis of dynamic processes or changes that occur over time within the sample being analyzed.
The inventors have also recognized and appreciated that, when a sample is tagged with a plurality of different types of luminescent markers, any suitable characteristic of luminescent markers may be used to identify the type of marker that is present in a particular pixel of the integrated device. For example, characteristics of the luminescence emitted by the markers and/or characteristics of the excitation absorption may be used to identify the markers. In some embodiments, the emission energy of the luminescence (which is directly related to the wavelength of the light) may be used to distinguish a first type of marker from a second type of marker. Additionally, or alternatively, luminescence lifetime measurements may also be used to identify the type of marker present at a particular pixel. In some embodiments, luminescence lifetime measurements may be made with a pulsed excitation source using a sensor capable of distinguishing a time when a photon is detected with sufficient resolution to obtain lifetime information. Additionally, or alternatively, the energy of the excitation light absorbed by the different types of markers may be used to identify the type of marker present at a particular pixel. For example, a first marker may absorb light of a first wavelength, but not equally absorb light of a second wavelength, while a second marker may absorb light of the second wavelength, but not equally absorb light of the first wavelength. In this way, when more than one excitation light source, each with a different excitation energy, may be used to illuminate the sample in an interleaved manner, the absorption energy of the markers can be used to identify which type of marker is present in a sample. Different markers may also have different luminescent intensities. Accordingly, the detected intensity of the luminescence may also be used to identify the type of marker present at a particular pixel.
One non-limiting example of an application of a device contemplated by the inventors is a device capable of performing sequencing of a biomolecule, such as a nucleic acid sequence (e.g., DNA, RNA) or a polypeptide (e.g. protein) having a plurality of amino acids. Diagnostic tests that may be performed using such a device include sequencing a nucleic acid molecule in a biological sample of a subject, such as sequencing of cell free deoxyribonucleic acid molecules or expression products in a biological sample of the subject.
The present application provides devices, systems and methods for detecting biomolecules or subunits thereof, such as nucleic acid molecules. Sequencing can include the determination of individual subunits of a template biomolecule (e.g., nucleic acid molecule) by synthesizing another biomolecule that is complementary or analogous to the template, such as by synthesizing a nucleic acid molecule that is complementary to a template nucleic acid molecule and identifying the incorporation of nucleotides with time (e.g., sequencing by synthesis). As an alternative, sequencing can include the direct identification of individual subunits of the biomolecule.
During sequencing, signals indicative of individual subunits of a biomolecule may be collected in memory and processed in real time or at a later point in time to determine a sequence of the biomolecule. Such processing can include a comparison of the signals to reference signals that enable the identification of the individual subunits, which in some cases yields reads. Reads may be sequences of sufficient length (e.g., at least about 30, 50, 100 base pairs (bp) or more) that can be used to identify a larger sequence or region, e.g., that can be aligned to a location on a chromosome or genomic region or gene.
Individual subunits of biomolecules may be identified using markers. In some examples, luminescent markers are used to identify individual subunits of biomolecules. Luminescent markers (also referred to herein as “markers”) may be exogenous or endogenous markers. Exogenous markers may be external luminescent markers used in a reporter and/or tag for luminescent labeling. Examples of exogenous markers may include, but are not limited to, fluorescent molecules, fluorophores, fluorescent dyes, fluorescent stains, organic dyes, fluorescent proteins, enzymes, species that participate in fluorescence resonance energy transfer (FRET), enzymes, and/or quantum dots. Such exogenous markers may be conjugated to a probe or functional group (e.g., molecule, ion, and/or ligand) that specifically binds to a particular target or component. Attaching an exogenous marker to a probe allows identification of the target through detection of the presence of the exogenous marker. Examples of probes may include proteins, nucleic acid (e.g. DNA, RNA) molecules, lipids and antibody probes. The combination of an exogenous marker and a functional group may form any suitable probes, tags, and/or labels used for detection, including molecular probes, labeled probes, hybridization probes, antibody probes, protein probes (e.g., biotin-binding probes), enzyme labels, fluorescent probes, fluorescent tags, and/or enzyme reporters.
While exogenous markers may be added to a sample, endogenous markers may be already part of the sample. Endogenous markers may include any luminescent marker present that may luminesce or “autofluoresce” in the presence of excitation energy. Autofluorescence of endogenous fluorophores may provide for label-free and noninvasive labeling without requiring the introduction of exogenous fluorophores. Examples of such endogenous fluorophores may include hemoglobin, oxyhemoglobin, lipids, collagen and elastin crosslinks, reduced nicotinamide adenine dinucleotide (NADH), oxidized flavins (FAD and FMN), lipofuscin, keratin, and/or prophyrins, by way of example and not limitation.
While some embodiments may be directed to diagnostic testing by detecting single molecules in a specimen, the inventors have also recognized that some embodiments may use the single molecule detection capabilities to perform nucleic acid (e.g. DNA, RNA) sequencing of one or more nucleic acid segments such as, for example, genes, or polypeptides. Nucleic acid sequencing allows for the determination of the order and position of nucleotides in a target nucleic acid molecule. Nucleic acid sequencing technologies may vary in the methods used to determine the nucleic acid sequence as well as in the rate, read length, and incidence of errors in the sequencing process. For example, some nucleic acid sequencing methods are based on sequencing by synthesis, in which the identity of a nucleotide is determined as the nucleotide is incorporated into a newly synthesized strand of nucleic acid that is complementary to the target nucleic acid molecule. Some sequencing by synthesis methods require the presence of a population of target nucleic acid molecules (e.g., copies of a target nucleic acid) or a step of amplification of the target nucleic acid to achieve a population of target nucleic acids.
Having recognized the need for simple, less complex apparatuses for performing single molecule detection and/or nucleic acid sequencing, the inventors have conceived of a technique for detecting single molecules using sets of markers, such as optical (e.g., luminescent) markers, to label different molecules. A tag may include a nucleotide or amino acid and a suitable marker. Markers may be detected while bound to single molecules, upon release from the single molecules, or while bound to and upon release from the single molecules. In some examples, markers are luminescent tags. Each luminescent marker in a selected set is associated with a respective molecule. For example, a set of four markers may be used to “label” the nucleobases present in DNA—each marker of the set being associated with a different nucleobase to form a tag, e.g., a first marker being associated with adenine (A), a second marker being associated with cytosine (C), a third marker being associated with guanine (G), and a fourth marker being associated with thymine (T). Moreover, each of the luminescent markers in the set of markers has different properties that may be used to distinguish a first marker of the set from the other markers in the set. In this way, each marker is uniquely identifiable using one or more of these distinguishing characteristics. By way of example and not limitation, the characteristics of the markers that may be used to distinguish one marker from another may include an emission wavelength or band of emission wavelengths of light emitted by the marker in response to excitation, a wavelength or band of wavelengths of the excitation energy that excites a particular marker, the temporal characteristics of the light emitted by the marker (e.g., emission decay time periods), and/or temporal characteristics of a marker's response to emission energy (e.g., probability of absorbing an excitation photon). Accordingly, luminescent markers may be identified or discriminated from other luminescent markers based on detecting these properties. Such identification or discrimination techniques may be used alone or in any suitable combination. In the context of nucleic acid sequencing, distinguishing a marker from among a set of four markers based on one or more the marker's emission characteristics may uniquely identify a nucleobase associated with the marker.
The system may include an integrated device and an instrument configured to interface with the integrated device. The integrated device may include an array of pixels, where a pixel includes a sample well and at least one sensor. A surface of the integrated device may have a plurality of sample wells, where a sample well is configured to receive a sample from a specimen placed on the surface of the integrated device. A specimen may contain multiple samples, and in some embodiments, different types of samples. The plurality of sample wells may have a suitable size and shape such that at least a portion of the sample wells receive one sample from a specimen. In some embodiments, the number of samples within a sample well may be distributed among the sample wells such that some sample wells contain one sample with others contain zero, two or more samples.
In some embodiments, a specimen may contain multiple single-stranded DNA templates, and individual sample wells on a surface of an integrated device may be sized and shaped to receive a single-stranded DNA template. Single-stranded DNA templates may be distributed among the sample wells of the integrated device such that at least a portion of the sample wells of the integrated device contain a single-stranded DNA template. The specimen may also contain tagged dNTPs which then enter in the sample well and may allow for identification of a nucleotide as it is incorporated into a strand of DNA complementary to the single-stranded DNA template in the sample well. In such an example, the “sample” may refer to both the single-stranded DNA and the tagged dNTP currently being incorporated by a polymerase. In some embodiments, the specimen may contain single-stranded DNA templates and tagged dNTPS may be subsequently introduced to a sample well as nucleotides are incorporated into a complementary strand of DNA within the sample well. In this manner, timing of incorporation of nucleotides may be controlled by when tagged dNTPs are introduced to the sample wells of an integrated device.
Excitation energy is provided from an excitation source located separate from the pixel array of the integrated device. The excitation energy is directed at least in part by elements of the integrated device towards one or more pixels to illuminate an illumination region within the sample well. A marker or tag may then emit emission energy when located within the illumination region and in response to being illuminated by excitation energy. In some embodiments, one or more excitation sources are part of the instrument of the system where components of the instrument and the integrated device are configured to direct the excitation energy towards one or more pixels.
Emission energy emitted by a sample may then be detected by one or more sensors within a pixel of the integrated device. Characteristics of the detected emission energy may provide an indication for identifying the marked associated with the emission energy. Such characteristics may include any suitable type of characteristic, including an arrival time of photons detected by a sensor, an amount of photons accumulated over time by a sensor, and/or a distribution of photons across two or more sensors. In some embodiments, a sensor may have a configuration that allows for the detection of one or more timing characteristics associated with a sample's emission energy (e.g., fluorescence lifetime). The sensor may detect a distribution of photon arrival times after a pulse of excitation energy propagates through the integrated device, and the distribution of arrival times may provide an indication of a timing characteristic of the sample's emission energy (e.g., a proxy for fluorescence lifetime). In some embodiments, the one or more sensors provide an indication of the probability of emission energy emitted by the marker or tag (e.g., fluorescence intensity). In some embodiments, a plurality of sensors may be sized and arranged to capture a spatial distribution of the emission energy. Output signals from the one or more sensors may then be used to distinguish a marker from among a plurality of markers, where the plurality of markers may be used to identify a sample within the specimen. In some embodiments, the In some embodiments, a sample may be excited by multiple excitation energies, and emission energy and/or timing characteristics of the emission energy emitted by the sample in response to the multiple excitation energies may distinguish a marker from a plurality of markers.
A schematic overview of the system 1-100 is illustrated in
A pixel 1-112 has a sample well 1-108 configured to receive a sample and a sensor 1-110 for detecting emission energy emitted by the sample in response to illuminating the sample with excitation energy provided by the excitation source 1-106. In some embodiments, sample well 1-108 may retain the sample in proximity to a surface of integrated device 1-102, which may ease delivery of excitation energy to the sample and detection of emission energy from the sample.
Optical elements for coupling excitation energy from excitation energy source 1-106 to integrated device 1-102 and guiding excitation energy to the sample well 1-108 are located both on integrated device 1-102 and the instrument 1-104. Source-to-well optical elements may comprise one or more grating couplers located on integrated device 1-102 to couple excitation energy to the integrated device and waveguides to deliver excitation energy from instrument 1-104 to sample wells in pixels 1-112. One or more optical splitter elements may be positioned between a grating coupler and the waveguides. The optical splitter may couple excitation energy from the grating coupler and deliver excitation energy to at least one of the waveguides. In some embodiments, the optical splitter may have a configuration that allows for delivery of excitation energy to be substantially uniform across all the waveguides such that each of the waveguides receives a substantially similar amount of excitation energy. Such embodiments may improve performance of the integrated device by improving the uniformity of excitation energy received by sample wells of the integrated device.
Sample well 1-108, a portion of the excitation source-to-well optics, and the sample well-to-sensor optics are located on integrated device 1-102. Excitation source 1-106 and a portion of the source-to-well components are located in instrument 1-104. In some embodiments, a single component may play a role in both coupling excitation energy to sample well 1-108 and delivering emission energy from sample well 1-108 to sensor 1-110. Examples of suitable components, for coupling excitation energy to a sample well and/or directing emission energy to a sensor, to include in an integrated device are described in U.S. patent application Ser. No. 14/821,688 titled “INTEGRATED DEVICE FOR PROBING, DETECTING AND ANALYZING MOLECULES,” and U.S. patent application Ser. No. 14/543,865 titled “INTEGRATED DEVICE WITH EXTERNAL LIGHT SOURCE FOR PROBING, DETECTING, AND ANALYZING MOLECULES,” both of which are incorporated by reference in their entirety.
Pixel 1-112 is associated with its own individual sample well 1-108 and at least one sensor 1-110. The plurality of pixels of integrated device 1-102 may be arranged to have any suitable shape, size, and/or dimensions. Integrated device 1-102 may have any suitable number of pixels. The number of pixels in integrated device 2-102 may be in the range of approximately 10,000 pixels to 1,000,000 pixels or any value or range of values within that range. In some embodiments, the pixels may be arranged in an array of 512 pixels by 512 pixels. Integrated device 1-102 may interface with instrument 1-104 in any suitable manner. In some embodiments, instrument 1-104 may have an interface that detachably couples to integrated device 1-104 such that a user may attach integrated device 1-102 to instrument 1-104 for use of integrated device 1-102 to analyze a sample and remove integrated device 1-102 from instrument 1-104 to allow for another integrated device to be attached. The interface of instrument 1-104 may position integrated device 1-102 to couple with circuitry of instrument 1-104 to allow for readout signals from one or more sensors to be transmitted to instrument 1-104. Integrated device 1-102 and instrument 1-104 may include multi-channel, high-speed communication links for handling data associated with large pixel arrays (e.g., more than 10,000 pixels).
Instrument 1-104 may include a user interface for controlling operation of instrument 1-104 and/or integrated device 1-102. The user interface may be configured to allow a user to input information into the instrument, such as commands and/or settings used to control the functioning of the instrument. In some embodiments, the user interface may include buttons, switches, dials, and a microphone for voice commands. The user interface may allow a user to receive feedback on the performance of the instrument and/or integrated device, such as proper alignment and/or information obtained by readout signals from the sensors on the integrated device. In some embodiments, the user interface may provide feedback using a speaker to provide audible feedback. In some embodiments, the user interface may include indicator lights and/or a display screen for providing visual feedback to a user.
In some embodiments, instrument 2-104 may include a computer interface configured to connect with a computing device. Computer interface may be a USB interface, a FireWire interface, or any other suitable computer interface. Computing device may be any general purpose computer, such as a laptop or desktop computer. In some embodiments, computing device may be a server (e.g., cloud-based server) accessible over a wireless network via a suitable computer interface. The computer interface may facilitate communication of information between instrument 1-104 and the computing device. Input information for controlling and/or configuring the instrument 1-104 may be provided to the computing device and transmitted to instrument 1-104 via the computer interface. Output information generated by instrument 1-104 may be received by the computing device via the computer interface. Output information may include feedback about performance of instrument 1-104, performance of integrated device 2-112, and/or data generated from the readout signals of sensor 1-110.
In some embodiments, instrument 1-104 may include a processing device configured to analyze data received from one or more sensors of integrated device 1-102 and/or transmit control signals to excitation source(s) 2-106. In some embodiments, the processing device may comprise a general purpose processor, a specially-adapted processor (e.g., a central processing unit (CPU) such as one or more microprocessor or microcontroller cores, a field-programmable gate array (FPGA), an application-specific integrated circuit (ASIC), a custom integrated circuit, a digital signal processor (DSP), or a combination thereof.) In some embodiments, the processing of data from one or more sensors may be performed by both a processing device of instrument 1-104 and an external computing device. In other embodiments, an external computing device may be omitted and processing of data from one or more sensors may be performed solely by a processing device of integrated device 1-104.
A cross-sectional schematic of integrated device 1-102 illustrating a row of pixels 1-112 is shown in
Coupling region 1-201 may include one or more optical components configured to couple excitation energy from external excitation source 1-106. Coupling region 1-201 may include grating coupler 1-216 positioned to receive some or all of a beam of excitation energy from excitation source 1-106. The beam of excitation energy may have any suitable shape and/or size. In some embodiments, a cross-section of the excitation energy beam may have an elliptical shape. In other embodiments, a cross-section of the excitation energy beam may have a circular shape.
Grating coupler 1-216 may be positioned to receive excitation energy from excitation source 1-106. Grating coupler 1-216 may be formed from one or more materials. In some embodiments, grating coupler 1-216 may include alternating regions of different materials along a direction parallel to propagation of light in the waveguide. Grating coupler 1-216 may include structures formed from one material surrounded by a material having a larger index of refraction. As an example, a grating coupler may include structures formed of silicon nitride and surrounded by silicon dioxide. Any suitable dimensions and/or inter-grating spacing may be used to form grating coupler 1-216. Spacing between structures of grating coupler 1-216 along a direction parallel to light propagation in waveguide 1-220, such as along the z-direction as shown in
Grating coupler 1-216 may couple excitation energy received from excitation source 1-214 to waveguide 1-220. Waveguide 1-220 is configured to propagate excitation energy to the proximity of one or more sample wells 1-108. In some embodiments, grating coupler 1-216 and waveguide 1-220 are formed in substantially the same plane of integrated device 1-102. In some embodiments, grating coupler 1-216 and waveguide 1-220 are formed from the same layer of integrated device 4-200 and may include the same material. In some embodiments, a mirror positioned over grating coupler 1-216 may direct excitation energy from an excitation source towards grating coupler 1-216. The mirror may be integrated into part of a housing positioned over the surface of the integrated device having the sample wells, where the housing may provide fluid containment for a sample. One or more sensors 1-230 may be positioned to receive excitation energy that passes through grating coupler 1-216 and/or passes through a region proximate to grating coupler 1-216, such as a region in the plane of grating coupler 1-216 outside of grating coupler 1-216.
In some embodiments, one or more filters may be positioned between waveguide 1-220 and sensors 1-110. The one or more filters may be configured to reduce or prevent excitation energy from passing towards sensors 1-110, which may contribute to signal noise of the sensors 1-110.
Coupling region may include reflective layer 1-226 positioned to receive excitation energy that may pass through grating coupler 1-216 (as shown by dashed lines in
Components located off of the integrated device may be used to position and align the excitation source 1-106 to the integrated device. Such components may include optical components including lenses, mirrors, prisms, apertures, attenuators, and/or optical fibers. Additional mechanical components may be included in the instrument to allow for control of one or more alignment components. Such mechanical components may include actuators, stepper motors, and/or knobs. Examples of suitable excitation sources and alignment mechanisms are described in U.S. Pat. Application 62/310,398 titled “PULSED LASER AND SYSTEM,” which is incorporated by reference in its entirety. Another example of a beam-steering module is described in U.S. Pat. Application 62/435,679 titled “COMPACT BEAM SHAPING AND STEERING ASSEMBLY,” which is incorporated herein by reference in its entirety.
Optical dump region 1-204 of integrated device 1-102 may include one or more components 1-240 at an end of waveguide 1-220 opposite to coupling region 1-201. Component(s) 1-240 may act to direct remaining excitation energy propagating through waveguide 1-220 after coupling with sample wells 1-110 out of waveguide 1-220. Component(s) 1-240 may improve performance of the integrated device by directing the remaining excitation energy away from the pixel region 1-203 of integrated device 1-102. Component(s) 1-240 may include grating coupler(s), optical coupler(s), taper(s), hairpin(s), undulator(s), or any other suitable optical components. In some embodiments, optical dump region 1-204 includes one or more sensors 1-242 positioned to receive excitation energy coupled out of waveguide 1-220. Signals from the one or more sensors 1-242 may provide an indication of optical power of the excitation energy propagating through waveguide 1-220, and in some embodiments, may be used to control optical power of an excitation energy beam generated by excitation source 1-106. In this manner, one or more sensors 1-242 may act as monitoring sensor(s). In some embodiments, optical bump region 1-204 may include component 1-240 and sensor 1-242 for each waveguide of integrated device 1-102.
A sample to be analyzed may be introduced into sample well 1-108 of pixel 1-112. The sample may be a biological sample or any other suitable sample, such as a chemical sample. The sample may include multiple molecules and the sample well may be configured to isolate a single molecule. In some instances, the dimensions of the sample well may act to confine a single molecule within the sample well, allowing measurements to be performed on the single molecule. An excitation source 1-106 may be configured to deliver excitation energy into the sample well 1-108, so as to excite the sample or at least one luminescent marker attached to the sample or otherwise associated with the sample while it is within an illumination area within the sample well 1-108.
When an excitation source delivers excitation energy to a sample well, at least one sample within the well may luminesce, and the resulting emission may be detected by a sensor. As used herein, the phrases “a sample may luminesce” or “a sample may emit radiation” or “emission from a sample” mean that a luminescent tag, marker, or reporter, the sample itself, or a reaction product associated with the sample may produce the emitted radiation.
One or more components of an integrated device may direct emission energy towards a sensor. The emission energy or energies may be detected by the sensor and converted to at least one electrical signal. The electrical signals may be transmitted along conducting lines in the circuitry of the integrated device connected to the instrument through the integrated device interface. The electrical signals may be subsequently processed and/or analyzed. Processing or analyzing of electrical signals may occur on a suitable computing device either located on or off the instrument.
In operation, parallel analyses of samples within the sample wells are carried out by exciting some or all of the samples within the wells using the excitation source and detecting signals from sample emission with the sensors. Emission energy from a sample may be detected by a corresponding sensor and converted to at least one electrical signal. The resulting signal, or signals, may be processed on the integrated device in some embodiments, or transmitted to the instrument for processing by the processing device and/or computing device. Signals from a sample well may be received and processed independently from signals associated with the other pixels.
In some embodiments, a sample may be labeled with one or more markers, and emission associated with the markers is discernable by the instrument. For example the sensor may be configured to convert photons from the emission energy into electrons to form an electrical signal that may be used to discern a lifetime that is dependent on the emission energy from a specific marker. By using markers with different lifetimes to label samples, specific samples may be identified based on the resulting electrical signal detected by the sensor.
A sample may contain multiple types of molecules and different luminescent markers may uniquely associate with a molecule type. During or after excitation, the luminescent marker may emit emission energy. One or more properties of the emission energy may be used to identify one or more types of molecules in the sample. Properties of the emission energy used to distinguish among types of molecules may include a fluorescence lifetime value, intensity, and/or emission wavelength. A sensor may detect photons, including photons of emission energy, and provide electrical signals indicative of one or more of these properties. In some embodiments, electrical signals from a sensor may provide information about a distribution of photon arrival times across one or more time intervals. The distribution of photon arrival times may correspond to when a photon is detected after a pulse of excitation energy is emitted by an excitation source. A value for a time interval may correspond to a number of photons detected during the time interval. Relative values across multiple time intervals may provide an indication of a temporal characteristic of the emission energy (e.g., lifetime). Analyzing a sample may include distinguishing among markers by comparing values for two or more different time intervals within a distribution. In some embodiments, an indication of the intensity may be provided by determining a number of photons across all time bins in a distribution.
An exemplary instrument 1-104 may comprise one or more mode-locked laser modules 1-258 mounted as a replaceable module within, or otherwise coupled to, the instrument, as depicted in
The output pulses 1-252 may be separated by regular intervals T. For example, T may be determined by a round-trip travel time between an output coupler and a cavity end mirror of laser module 1-258. According to some embodiments, the pulse-separation interval T may be in the range of approximately 1 ns to approximately 30 ns, or any value or range of values within that range. In some cases, the pulse-separation interval T may be in the range of approximately 5 ns to approximately 20 ns, corresponding to a laser-cavity length (an approximate length of an optical axis within a laser cavity of laser module 1-258) between about 0.7 meter and about 3 meters.
According to some embodiments, a desired pulse-separation interval T and laser-cavity length may be determined by a combination of the number of sample wells on integrated device 1-102, fluorescent emission characteristics, and the speed of data-handling circuitry for reading data from integrated device 1-102. The inventors have recognized and appreciated that different fluorophores may be distinguished by their different fluorescent decay rates or characteristic lifetimes. Accordingly, there needs to be a sufficient pulse-separation interval T to collect adequate statistics for the selected fluorophores to distinguish between their different decay rates. Additionally, if the pulse-separation interval T is too short, the data handling circuitry cannot keep up with the large amount of data being collected by the large number of sample wells. The inventors have recognized and appreciated that a pulse-separation interval T between about 5 ns and about 20 ns is suitable for fluorophores that have decay rates up to about 2 ns and for handling data from between about 60,000 and 600,000 sample wells.
According to some implementations, a beam-steering module may receive output pulses from the mode-locked laser module 1-125 and be configured to adjust at least the position and incident angles of the optical pulses onto an optical coupler of the integrated device 1-102. In some cases, the output pulses from the mode-locked laser module may be operated on by a beam-steering module to additionally or alternatively change a beam shape and/or beam rotation at an optical coupler on the integrated device 1-102. In some implementations, the beam-steering module may further provide focusing and/or polarization adjustments of the beam of output pulses onto the optical coupler. One example of a beam-steering module is described in U.S. patent application Ser. No. 15/161,088 titled “PULSED LASER AND BIOANALYTIC SYSTEM,” filed May 20, 2016, which is incorporated herein by reference. Another example of a beam-steering module is described in U.S. Pat. Application 62/435,679 titled “COMPACT BEAM SHAPING AND STEERING ASSEMBLY,” which is incorporated herein by reference in its entirety.
Referring to
Each waveguide 1-312 may include a tapered portion 1-315 below the sample wells 1-330 to equalize optical power coupled to the sample wells along the waveguide. The reducing taper may force more optical energy outside the waveguide's core, increasing coupling to the sample wells and compensating for optical losses along the waveguide, including losses for light coupling into the sample wells. A second grating coupler 1-317 may be located at an end of each waveguide to direct optical energy to an integrated photodiode 1-324. The integrated photodiode may detect an amount of power coupled down a waveguide and provide a detected signal to feedback circuitry that controls a beam-steering module.
The sample wells 1-330 may be aligned with the tapered portion 1-315 of the waveguide and recessed in a tub 1-340. There may be time-binning photodetectors 1-322 located on the semiconductor substrate 1-305 for each sample well 1-330. A metal coating and/or multilayer coating 1-350 may be formed around the sample wells and above the waveguide to prevent optical excitation of fluorophores that are not in the sample well (e.g., dispersed in a solution above the sample wells). The metal coating and/or multilayer coating 1-350 may be raised beyond edges of the tub 1-340 to reduce absorptive losses of the optical energy in the waveguide 1-312 at the input and output ends of each waveguide.
There may be a plurality of rows of waveguides, sample wells, and time-binning photodetectors on the integrated device. For example, there may be 128 rows, each having 512 sample wells, for a total of 65,536 sample wells in some implementations. Other implementations may include fewer or more sample wells, and may include other layout configurations. Optical power from a mode-locked laser may be distributed to the multiple waveguides via one or more star couplers and/or multi-mode interference couplers, or by any other means, located between an optical coupler of the integrated device and the plurality of waveguides.
A non-limiting example of a biological reaction taking place in a sample well 1-330 is depicted in
When a labeled nucleotide and/or nucleotide analog 1-610 is incorporated into a growing strand of complementary nucleic acid, as depicted in
According to some embodiments, an instrument 1-104 that is configured to analyze samples based on fluorescent emission characteristics may detect differences in fluorescent lifetimes and/or intensities between different fluorescent molecules, and/or differences between lifetimes and/or intensities of the same fluorescent molecules in different environments. By way of explanation,
A second fluorescent molecule may have a decay profile that is exponential, but has a measurably different lifetime TB, as depicted for curve B in
The inventors have recognized and appreciated that differences in fluorescent emission lifetimes can be used to discern between the presence or absence of different fluorescent molecules and/or to discern between different environments or conditions to which a fluorescent molecule is subjected. In some cases, discerning fluorescent molecules based on lifetime (rather than emission wavelength, for example) can simplify aspects of an 1 instrument 1-104. As an example, wavelength-discriminating optics (such as wavelength filters, dedicated detectors for each wavelength, dedicated pulsed optical sources at different wavelengths, and/or diffractive optics) may be reduced in number or eliminated when discerning fluorescent molecules based on lifetime. In some cases, a single pulsed optical source operating at a single characteristic wavelength may be used to excite different fluorescent molecules that emit within a same wavelength region of the optical spectrum but have measurably different lifetimes. An analytic system that uses a single pulsed optical source, rather than multiple sources operating at different wavelengths, to excite and discern different fluorescent molecules emitting in a same wavelength region can be less complex to operate and maintain, more compact, and may be manufactured at lower cost.
Although analytic systems based on fluorescent lifetime analysis may have certain benefits, the amount of information obtained by an analytic system and/or detection accuracy may be increased by allowing for additional detection techniques. For example, some analytic systems 2-160 may additionally be configured to discern one or more properties of a sample based on fluorescent wavelength and/or fluorescent intensity.
Referring again to
For a single molecule or a small number of molecules, however, the emission of fluorescent photons occurs according to the statistics of curve B in
Examples of a time-binning photodetector 1-322 are described in U.S. patent application Ser. No. 14/821,656, filed Aug. 7, 2015, titled “INTEGRATED DEVICE FOR TEMPORAL BINNING OF RECEIVED PHOTONS,” which is incorporated herein by reference. For explanation purposes, a non-limiting embodiment of a time-binning photodetector is depicted in
In operation, fluorescent photons may be received at the photon-absorption/carrier-generation region 1-902 at different times and generate carriers. For example, at approximately time ti three fluorescent photons may generate three carrier electrons in a depletion region of the photon-absorption/carrier-generation region 1-902. An electric field in the device (due to doping and/or an externally applied bias to electrodes 1-920 and 1-922, and optionally or alternatively to 1-932, 1-934, 1-936) may move the carriers to the carrier-travel region 1-906. In the carrier-travel region, distance of travel translates to a time after excitation of the fluorescent molecules. At a later time t5, another fluorescent photon may be received in the photon-absorption/carrier-generation region 1-902 and generate an additional carrier. At this time, the first three carriers have traveled to a position in the carrier-travel region 1-906 adjacent to the second storage bin 1-908b. At a later time t7, an electrical bias may be applied between electrodes 1-932, 1-934, 1-936 and electrode 1-940 to laterally transport carriers from the carrier-travel region 1-906 to the storage bins. The first three carriers may then be transported to and retained in the first bin 1-908a and the later-generated carrier may be transported to and retained in the third bin 1-908c. In some implementations, the time intervals corresponding to each storage bin are at the sub-nanosecond time scale, though longer time scales may be used in some embodiments (e.g., in embodiments where fluorophores have longer decay times).
The process of generating and time-binning carriers after an excitation event (e.g., excitation pulse from a pulsed optical source) may occur once after a single excitation pulse or be repeated multiple times after multiple excitation pulses during a single charge-accumulation cycle for the photodetector 1-900. After charge accumulation is complete, carriers may be read out of the storage bins via the read-out channel 1-910. For example, an appropriate biasing sequence may be applied to at least electrode 1-940 and a downstream electrode (not shown) to remove carriers from the storage bins 1-908a, 1-908b, 1-908c.
After a number of excitation events, the accumulated signal in each electron-storage bin may be read out to provide a histogram having corresponding bins that represent the fluorescent emission decay rate, for example. Such a process is illustrated in
In some implementations, only a single photon on average may be emitted from a fluorophore following an excitation event, as depicted in
After a large number of excitation events and signal accumulations, the electron-storage bins of the time-binning photodetector 1-322 may be read out to provide a multi-valued signal (e.g., a histogram of two or more values, an N-dimensional vector, etc.) for a sample well. The signal values for each bin may depend upon the decay rate of the fluorophore. For example and referring again to
To further aid in understanding the signal analysis, the accumulated, multi-bin values may be plotted as a histogram, as depicted in
In some implementations, fluorescent intensity may be used additionally or alternatively to distinguish between different fluorophores. For example, some fluorophores may emit at significantly different intensities or have a significant difference in their probabilities of excitation (e.g., at least a difference of about 35%) even though their decay rates may be similar. By referencing binned signals (bins 1-3) to measured excitation energy bin 0, it may be possible to distinguish different fluorophores based on intensity levels.
In some embodiments, different numbers of fluorophores of the same type may be linked to different nucleotides or nucleotide analogs, so that the nucleotides may be identified based on fluorophore intensity. For example, two fluorophores may be linked to a first nucleotide (e.g., “C”) or nucleotide analog and four or more fluorophores may be linked to a second nucleotide (e.g., “T”) or nucleotide analog. Because of the different numbers of fluorophores, there may be different excitation and fluorophore emission probabilities associated with the different nucleotides. For example, there may be more emission events for the “T” nucleotide or nucleotide analog during a signal accumulation interval, so that the apparent intensity of the bins is significantly higher than for the “C” nucleotide or nucleotide analog.
The inventors have recognized and appreciated that distinguishing nucleotides or any other biological or chemical specimens based on fluorophore decay rates and/or fluorophore intensities enables a simplification of the optical excitation and detection systems in an instrument 1-104. For example, optical excitation may be performed with a single-wavelength source (e.g., a source producing one characteristic wavelength rather than multiple sources or a source operating at multiple different characteristic wavelengths). Additionally, wavelength discriminating optics and filters may not be needed in the detection system. Also, a single photodetector may be used for each sample well to detect emission from different fluorophores.
The phrase “characteristic wavelength” or “wavelength” is used to refer to a central or predominant wavelength within a limited bandwidth of radiation (e.g., a central or peak wavelength within a 20 nm bandwidth output by a pulsed optical source). In some cases, “characteristic wavelength” or “wavelength” may be used to refer to a peak wavelength within a total bandwidth of radiation output by a source.
The inventors have recognized and appreciated that fluorophores having emission wavelengths in a range between about 560 nm and about 900 nm can provide adequate amounts of fluorescence to be detected by a time-binning photodetector (which may be fabricated on a silicon wafer using CMOS processes). These fluorophores can be linked to biological molecules of interest such as nucleotides or nucleotide analogs. Fluorescent emission in this wavelength range may be detected with higher responsivity in a silicon-based photodetector than fluorescence at longer wavelengths. Additionally, fluorophores and associated linkers in this wavelength range may not interfere with incorporation of the nucleotides or nucleotide analogs into growing strands of DNA. The inventors have also recognized and appreciated that fluorophores having emission wavelengths in a range between about 560 nm and about 660 nm may be optically excited with a single-wavelength source. An example fluorophore in this range is Alexa Fluor 647, available from Thermo Fisher Scientific Inc. of Waltham, Mass. The inventors have also recognized and appreciated that excitation energy at shorter wavelengths (e.g., between about 500 nm and about 650 nm) may be required to excite fluorophores that emit at wavelengths between about 560 nm and about 900 nm. In some embodiments, the time-binning photodetectors may efficiently detect longer-wavelength emission from the samples, e.g., by incorporating other materials, such as Ge, into the photodetectors active region.
Although the prospect of sequencing DNA using an excitation source that emits a single characteristic wavelength can simplify some of the optical system, it can place technically challenging demands on the excitation source, as noted above. For example, the inventors have recognized and appreciated that optical pulses from the excitation source should extinguish quickly for the detection schemes described above, so that the excitation energy does not overwhelm or interfere with the subsequently detected fluorescent signal. In some embodiments and referring again to
Performance of an integrated device in analyzing samples can depend on various factors related to optics of the integrated device, including coupling efficiency of an optical coupler (e.g., grating coupler) of the integrated device, optical loss in splitting excitation energy into individual waveguides, and coupling efficiency of individual waveguides into sample wells. These factors may become exaggerated as more sample wells and optical components are included on the integrated device to deliver excitation energy to the sample wells. Aspects of the present application relate to optical couplers, optical splitters, waveguides, and techniques for arranging these optical components in an integrated device to reduce optical loss and/or improve coupling efficiency. In addition, the techniques described herein may improve uniformity in the delivery of excitation energy to the sample wells of an integrated device.
Performance of an integrated device in analyzing samples can also depend on the amount of excitation energy (e.g., optical power) delivered to individual sample wells. As excitation energy propagates from an excitation source to a sample well, optical loss may occur which can reduce the amount of excitation energy that couples to the sample well and may impact the performance of the pixel associated with the sample well in detecting a sample. For an array of sample wells, such optical loss may limit the number of pixels capable of sample detection. Such optical loss may also reduce the uniformity in delivering excitation energy to individual sample wells in the array, which may also impact the performance of the integrated device. A waveguide of the integrated device may couple excitation energy to a number of sample wells (e.g., 512 sample wells) positioned proximate to the waveguide. As excitation energy propagates along the waveguide, the amount of total optical loss may increase, reducing the amount of excitation energy that couples to sample wells positioned further along the waveguide. In this manner, optical loss along the waveguide may impact the uniformity in the amount of excitation energy coupled to individual sample wells positioned along the waveguide. Aspects of the present application relate to integrated devices, and methods of forming integrated devices, that improve uniformity of excitation energy within the array of sample wells by reducing optical loss as excitation energy propagates along a waveguide.
In some cases, problems can arise when trying to couple power from an optical source efficiently to a large plurality of integrated optical waveguides. To provide sufficient power to each waveguide and sample well for a large number of sample wells, the average power in the input beam rises proportionally with the increase in the number of sample wells. For some integrated optical waveguides (e.g., a silicon-nitride waveguide core/silicon-dioxide cladding), high powers can cause temporal changes in the transmission loss of the waveguide and therefore cause appreciable power instabilities in the sample wells over time. Time-dependent transmission loss in integrated optical waveguides at high powers has been measured by the inventors, and example results are plotted in
Insertion loss was measured as a function of time for three identical lengths of single-mode waveguides having a silicon-nitride core. The initial average power levels coupled into the three waveguides was 0.5 mW, 1 mW, and 2 mW. The plot of
In cases where emission intensities from the sample wells are low or where characterization of a sample depends upon intensity values from the sample wells, it is beneficial that the power delivered to the sample wells remains stable over time. For example, if the power delivered to the sample wells decreases by 3 dB (see
One approach to reducing the effects of time-dependent waveguide transmission loss is to reduce the length of integrated waveguides used in an integrated device. In some cases, appreciable lengths of waveguides may be needed to route excitation energy to the sample wells. Alternatively or additionally, the intensity of radiation coupled into the waveguides may be reduced and/or the optical loss along a waveguide that arises from a metal layer may be increased. The inventors have recognized and appreciated that the time-dependent waveguide transmission loss may be most problematic where a beam from an excitation source is coupled first into a single waveguide of an integrated optical circuit and then redistributed among many waveguides (e.g., by using a binary tree of multimode interference splitters having one input and two outputs). At the coupling region, in such instances, the intensity may be very high and cause rapid changes in waveguide transmission loss.
Some embodiments of the present application relate to waveguide structures, and methods of forming waveguide structures, that provide an optical mode having a desired evanescent field extending from the waveguide. An evanescent field extending perpendicular to the direction of propagation along the waveguide may have a distribution of optical power that decreases from the waveguide. The evanescent field may have a characteristic decay at which the optical power decreases from the waveguide. A waveguide configured to support propagation of an optical mode may be considered to be a “confined” optical mode when the evanescent field decays quickly from the waveguide.
One or more dimensions of a waveguide may impact a characteristic of the evanescent field, including the decay rate, distance of the evanescent field from an interface between a waveguide material and a surrounding material (e.g., cladding), and optical power profile of the evanescent field in a direction perpendicular from the waveguide propagation direction. A dimension of a waveguide perpendicular to the direction of propagation along the waveguide may impact one or more characteristics of the evanescent field. In some embodiments, a thickness of a waveguide may impact one or more characteristics of an evanescent field. The thickness of the waveguide may impact the decay of the evanescent field of excitation energy propagating along the waveguide. In some embodiments, increasing the thickness of the waveguide may increase the decay of the evanescent field.
Some embodiments relate to waveguide structures that have a variable thickness to provide a desired evanescent field for coupling to one or more sample wells of an integrated device. In some embodiments, the thickness of the waveguide may be larger in a region that overlaps with one or more sample wells than in a region that is non-overlapping with the one or more sample wells. In such embodiments, the waveguide may provide an optical mode having an evanescent field that provides a desired amount of coupling of excitation energy into a sample well while reducing optical loss from the presence of a metal layer.
Another technique for reducing optical loss and improving optical performance of an integrated device may include varying the power distribution of excitation energy along the length of a waveguide of the integrated device. The power distribution may increase and/or broaden at locations along the waveguide that overlap with a sample well and decrease and/or narrow at locations along the waveguide that do not overlap with a sample well. In some embodiments, a waveguide of an integrated device may propagate a plurality of optical modes. Such a waveguide may be considered a “multimode waveguide.” The plurality of optical modes may interfere to vary the power distribution of excitation energy in a direction perpendicular to the direction of light propagation along the waveguide. The power distribution of the multimode waveguide may vary such that the power distribution broadens at one or more positions along the waveguide that overlap with a sample well.
A. Grating Coupler
To reduce time-dependent waveguide loss at coupling region 1-201, a sliced grating coupler 2-100, of which a simplified illustration is shown in
In some embodiments, a beam from the excitation source 1-106 may be expanded (or produced by the excitation source) so that it extends in the Y direction to essentially match the length L of the grating. For example, the extended beam 2-112 may have a shape as depicted by the dashed ellipse in
In some instances, it is desirable to provide for adjustable uniformity of coupling of power into the plurality of waveguides 2-120 with the sliced grating coupler 2-100 and beam arrangement depicted in
During operation, the angle α and the beam displacement in the X and Y directions may be adjusted to obtain and maintain uniform coupling of power across the plurality of waveguides 2-120. If a beam 2-122 has an asymmetric intensity profile in the Y direction, then the position of the beam may be adjusted in the X direction to compensate for the asymmetry. For example, if the intensity of the beam in the +Y direction is greater than the intensity of the beam in the −Y direction, then the beam may be moved in the −X direction (for the angle shown) so that a portion of the beam in the +Y direction moves off the grating 2-110 and reduces the amount of power coupled to the tapered ends 2-122 in the +Y direction. A portion of the beam in the −Y direction may move onto the grating 2-110 and increase the amount of power coupled to the tapered ends 2-122 in the −Y direction. If a beam 2-122 has a symmetric intensity profile in the Y direction, then adjustments in the ±Y directions and/or ±α directions can be made to improve uniformity of power coupled into the waveguides. An example of a beam-steering module used to align an elliptical beam to a sliced grating coupler is described in U.S. Pat. Application 62/435,679 titled “COMPACT BEAM SHAPING AND STEERING ASSEMBLY,” which is incorporated herein by reference in its entirety.
One or more dimensions of the tapered ends of a sliced grating coupler may vary to compensate for variation in optical intensity coupled to the sliced grating coupler. In some embodiments, the width (along the y-axis shown in
In some embodiments, one or more dimensions of the tapered ends of a sliced grating coupler may vary to account for optical components (e.g., optical splitters) within an optical system of an integrated device. To distribute the excitation energy among many waveguides within the integrated device, output waveguides from a sliced grating coupler may couple with an optical splitter to increase the number of waveguides propagating excitation energy. Some of the output waveguides may couple excitation energy along an optical path that has only one optical splitter, while output waveguides may couple excitation energy along an optical path that has two or more optical splitters. A dimension of the tapered ends may vary depending on the number of optical splitters in an optical path that each output waveguide couples to, in addition to accounting for intensity distribution within the grating. In some embodiments, a sliced grating coupler may have one tapered end with a larger dimension than both a tapered end proximate to an edge of the grating and a tapered end proximate to the center of a side of the grating.
B. Optical Splitter(s)
One or more optical splitters (e.g., multimode interference splitter) may be positioned between grating coupler 1-216 and waveguide 1-220, and may be included as part of routing region 1-202 in some embodiments. An optical splitter may couple to an output waveguide of the grating coupler as an input to the optical splitter and have two or more waveguides as outputs of the optical splitter. In some embodiments, multiple optical splitters may be used to divide the optical power received by the grating coupler 1-216 into waveguides 1-220 that propagate excitation energy to sample wells 1-108 in the pixel region 1-203 of the integrated device. In some embodiments, the number of optical splitters between the grating coupler and a waveguide that couples excitation energy to a sample well may vary depending on how output waveguides from the grating coupler are positioned and/or sized.
As shown in
C. Array Layout
Some embodiments of the present application relate to techniques for routing of waveguides and optical components in an integrated device in order to improve device performance and/or reduce time-dependent waveguide loss, as discussed above, such as by decreasing waveguide lengths. Another consideration in routing of waveguides and optical components may include reducing the footprint of the integrated device devoted to optical routing to allow for more surface area available for additional sample wells.
In some embodiments, waveguides may be routed in a radial distribution from the grating coupler. As shown in
In some embodiments, waveguides in a pixel region of an integrated device may be positioned substantially parallel to gratings of a grating coupler. An optical propagation region may optically couple the grating coupler to the waveguides. Such a waveguide layout may allow for shorter waveguides, which can reduce optical loss, including time-dependent waveguide loss.
In some embodiments, one or more optical splitters (e.g., MMI splitters) may be positioned in a pixel region of an integrated device and configured to couple with two or more waveguides configured to optically couple with a row or column of sample wells. The one or more optical splitters may be positioned between two sets of sample wells. One or more input waveguides to an optical splitter may be positioned between the two sets of sample wells. An input waveguides may be a waveguide that couples to a propagation region, such as waveguides 3-130a and 3-130b that stem from propagation region 3-120 shown in
D. Sample Wells
An integrated device of the type described herein may comprise one or more sample wells configured to receive samples therein. The integrated device may comprise pixels disposed in rows of sample wells (e.g., 512 sample wells). Each sample well may receive a sample, which may be disposed on a surface of the sample well, such as a bottom surface. The surface on which the sample is to be disposed may have a distance from the waveguide that is configured to excite the sample with a desired level of excitation energy. In some embodiments, the sample well may be positioned, with respect to the waveguide, such that an evanescent field of an optical mode propagating along the waveguide overlaps with the sample.
A sample well may have a top aperture through which one or more samples may access the sample well. The size of the top aperture may depend on different factors. One such factor relates to the fact that one or more samples may be positioned in the sample well. Accordingly, the top aperture may be large enough to allow for placement of the sample in the sample well. Another factor relates to background signals, such as stray light. When one or more samples are disposed in the sample well and are excited with excitation energy, background signals may cause undesired fluctuations in the emission energy, thus making the measurement noisy. To limit such fluctuations, the size of the top aperture may be configured to block at least a portion of the background signals. Similarly, the top aperture blocks the exposure of the sample such that only the portion of the sample under the aperture receives substantial excitation energy. Another factor relates to the directivity of emission energy emitted by the sample(s) in response to receiving excitation energy. In some embodiments, the size of the top aperture may be configured to provide a desired level of directivity.
Some embodiments of the integrated device include sample wells formed within a metal layer on the surface of the integrated device. The metal layer may provide benefits in detecting emission energy from a sample well by one or more sensors. The metal layer may act to reduce background signals and to improve the amount of emission energy detected by the one or more sensors. Such metal layers may improve the signal-to-noise ratio of the sensors by reducing noise artifacts that can arise from background signals (e.g., stray light, background light or direct excitation energy). In some embodiments, the integrated device may include metal layers configured to act as wiring to transmit and/or receive electrical signals. Such wiring may couple to a sensor and transmit signals to control the sensor and/or receive signals indicative of the emission energy detected by the sensor.
The depth of a sample well may be configured to maintain a desired separation between the location of the sample(s) and the metal layers. Such separation may ensure that the sample well is provided with a desired level of excitation energy while limiting optical loss caused by the metal layers. In some embodiments, the depth of a sample well may be configured such that the evanescent field of an optical mode propagating along a waveguide overlaps with the sample while limiting the extent to which it interacts with the metal layers. In some embodiments, the depth of a sample well may impact the timing of photon emission events of a marker (e.g., lifetime) associated with the sample. Accordingly, the depth may allow for distinguishing among different markers in the sample well based on timing characteristics associated with the individual lifetimes of the different markers.
The shape and size of the sample well and/or the composition of metal layers may act to direct emission energy towards a sensor. In some embodiments, a portion of the energy emitted by a sample in the form of emission energy may propagate downward through the layers of the integrated device. A portion of the emission energy may be received by one or more sensors disposed on the integrated device in a pixel associated with the sample well.
Sample 4-191 may be excited with excitation energy provided through waveguide 4-102, such as by waveguide 4-102 optically coupling with sample well 4-108. While waveguide 4-102 is illustrated as having a rectangular cross section in
Sample well 4-108 may have a depth dW between a surface 4-112 of sample well 4-108 and interface 4-127 between cladding 4-118 and metal layer(s) 4-122. Depth dW may provide a suitable distance between a sample positioned at the surface 4-112 from metal layer(s) 4-122. Depth dW may impact the timing of photon emission events of a marker (e.g., fluorescence lifetime of a fluorophore) associated with sample 4-191. Accordingly, depth dW may allow for distinguishing among different markers in sample well 4-108 based on timing characteristics associated with the individual photon emission timing characteristics (e.g., fluorescence lifetimes) of the different markers. In some embodiments, depth dW of sample well 4-108 may impact the amount of excitation energy received. Depth dW of sample well 4-108 may be configured to improve the directivity of emission energy from sample 4-191. Depth dW may be in the range of 50 nm to 400 nm, or any value or range of values within that range. In some embodiments, depth dW is between 95 nm and 150 nm. In some embodiments, depth dW is between 250 nm and 350 nm.
An integrated device may include metal layer(s) 4-122 over top cladding 4-118. Metal layer(s) 4-122 may act as a reflector for emission energy emitted by a sample in a sample well and may improve detection of emission energy by reflecting emission energy towards a sensor of the integrated device. Metal layer(s) 4-122 may act to reduce the background signal due to photons that do not originate within the sample well. Metal layer(s) 4-122 may comprise one or more sub-layers. Examples of suitable materials to be used as layers of metal layer(s) may include aluminum, copper, aluminum-copper alloys, titanium, titanium nitride, tantalum, and tantalum nitride. As shown in
Metal layer(s) 4-122 may further include a second sub-layer 4-126 disposed over the first sub-layer 4-124. In some embodiments, the second sub-layer 4-126 may include titanium. Titanium may reduce the amount of corrosion that occurs within metal layer(s) 4-122. The thickness of the second sub-layer 4-126 may be in the range of 1 nm to 100 nm, or any value or range of values within that range. In some embodiments, the thickness of the second sub-layer may be approximately 10 nm.
Metal layer(s) 4-122 may further include a third sub-layer 4-128 disposed over the second sub-layer 4-126 and/or over the first sub-layer 4-124. The third sub-layer 4-128 may include titanium nitride and/or tanatalum nitride. The third sub-layer 4-128 may have a thickness in the range of 5 nm to 100 nm, or any value or range of values within that range. In some embodiments, the third sub-layer 4-128 may have a thickness of approximately 50 nm.
Sample well 4-108 may have one or more sidewalls covered, at least partially, with a sidewall spacer 4-190. The composition of sidewall spacer 4-190 may be configured to enable a particular type of interaction with sample 4-191. In some embodiments, sidewall spacer 4-190 may have a composition configured to passivate the sidewalls of sample well 4-108 to reduce the amount of sample that adheres to the sidewall of sample well 4-108. By providing a sample well with only the sidewalls coated with a spacer material, a different type of interaction with sample 4-191 may occur at sidewalls 4-190 than at surface 4-112. In some embodiments, the surface 4-112 of sample well 4-108 may be coated with a functionalized silane to improve adherence of sample 4-191 to the surface. By coating the sidewalls with spacer 4-190, the surface 4-112 of the sample well 4-108 may be selectively coated with the functionalized silane. The composition of sidewall spacer 4-190 may be selected to provide selective coatings of sidewall spacer 4-190 relative to surface 4-112 of sample well 4-108 that is substantially parallel to the waveguide, which may be considered as a “bottom surface” of the sample well. Sidewall spacer 4-190 may have a thickness in the range of 3 nm to 30 nm, or any value or range of values within that range. In some embodiments, sidewall spacer 4-190 may have a thickness of approximately 10 nm. Examples of suitable materials used to form sidewall spacer 4-190 include Al2O3, TiO2, TiN, TiON, TaN, Ta2O5, Zr2O5, Nb2O5, and HfO2. In some embodiments, sidewall spacer 4-190 includes TiN, which may provide a desired level of directionality of emission energy towards a sensor due to the refractive index of TiN. In some embodiments, sidewall spacer 4-190 may be configured to block scattered light, thus reducing the amount of scattered light that may illuminate sample 4-191.
In some embodiments, the sample well structure may have a portion proximate to waveguide 4-102 that lacks spacer material on the sidewalls. The distance between the bottom surface, such as surface 4-112 shown in
E. Waveguides
An excitation source may be used to generate excitation energy at a desired wavelength (e.g., 532 nm). The excitation energy may be provided to individual samples using one or more waveguides. The waveguide(s) may be configured to couple a portion of the excitation energy to individual samples, for example via evanescent coupling. In some embodiments, the sample wells may be arranged in rows and columns, and individual waveguides may be configured to deliver excitation energy to sample wells of a corresponding row or column. In some embodiments, the waveguide may be configured to substantially uniformly provide (e.g., with a variation in intensity that is less than 10%) excitation energy among the sample wells in a row or column. To provide such uniform illumination of the sample wells, the waveguide may be configured to have a coupling coefficient, with respect to the sample well, that varies along the length of the row or column. Accordingly, individual sample wells positioned relative to the waveguide may receive a fraction of the excitation energy propagating along the waveguide. As the excitation energy propagating along the waveguide is depleted by successive coupling with sample wells, the coupling coefficient may be progressively increased to provide a substantially uniform amount of excitation energy among the sample wells coupling with the waveguide. To provide such space-dependent coupling coefficients, tapering of the waveguide may be used. A “taper” may refer to a waveguide having a dimension (e.g., width) that varies along its length. The taper may be configured to progressively expand the supported optical mode farther into the surrounding region (e.g., cladding). Through such tapering of the waveguide, the coupling coefficient may increase along a propagation axis of the waveguide.
The waveguide(s) may be further configured to effectively couple excitation energy to the sample wells while reducing optical loss. Because the sample wells may be disposed in proximity to a metal layer, the excitation energy guided in the waveguide may experience optical loss due to metal scattering and/or metal absorption. To reduce optical loss caused by the metal layer(s), the waveguide may be configured to provide a mode confinement such that the spatial overlap of the mode with respect to the metal layer is reduced. The mode confinement may be selected to provide a desired overlap with the sample wells while reducing the interaction with the metal layer(s).
The waveguides may be fabricated from a material that is transparent (e.g., having a propagation loss that is less than 2 dB/cm) at the wavelength of the excitation energy. For example, silicon nitride may be used as a material for guiding excitation energy.
In some embodiments, channel waveguides may be used to provide excitation energy to the sample wells of an integrated device. An example of a suitable channel waveguide is shown in
In some embodiments, rib waveguides and/or ridge waveguides may be used to provide excitation energy to the sample wells. Rib waveguides, or ridge waveguides, may comprise a first layer, referred to as the “slab”, and a second layer, referred to as the “raised region”. The position of the raised region with respect to the slab may determine the location of the optical mode. The thickness of the slab and the raised region may be configured to provide a desired optical profile. For example, it may be desirable to have an optical mode profile such that the evanescent field overlaps with the sample while reducing the interaction with the metal layer(s).
Alternatively, individual waveguides may comprise separate slabs.
Waveguides 4-200 and 4-250 may comprise a bottom cladding 4-208 and a top cladding 4-206. The bottom and top cladding may be formed from materials having a refractive index that is lower than the refractive index of the raised regions 4-204 and 4-254. In some embodiments, the bottom and top cladding may comprise silicon oxide. The ratio TRR/TS may be selected to obtain a desired level of optical confinement. For example, such ratio may be selected so that the optical mode of the waveguide experiences reduced optical loss from the metal layer(s) while also providing a desired level of coupling to the sample wells. Requiring less fabrication steps compared to waveguide 4-250, waveguide 4-200 may be preferable in some embodiments. In other embodiments, waveguide 4-250 may be preferable because it provides a lower degree of coupling to other waveguides in comparison to waveguide 4-200.
In some embodiments, the slab of the waveguide and/or the raised region of the waveguide may comprise more than one layer.
A waveguide of the type described herein may be disposed in correspondence with a sample well as illustrated in
According to coupling configuration 4-300B, the bottom surface of sample well 4-312 may be disposed within waveguide 4-301. Compared to the configuration illustrated in
According to coupling configuration 4-300C, the bottom surface of sample well 4-312 may be disposed in contact with a surface of waveguide 4-301. The configuration may be obtained, for example, by using a surface of waveguide 4-301 as an etch stop to form sample well 4-312. Compared to the configuration illustrated in
As described above, a waveguide of the type describe herein may be configured to support at least one optical mode. As defined herein, the “optical mode”, or simply the “mode”, refers to the profile of the electromagnetic field associated with a particular waveguide. The optical mode may propagate excitation energy along a waveguide. The optical mode may be configured to evanescently couple to a sample well, thus exciting a sample disposed therein. In response the sample may emit emission energy. At the same time, the optical mode may be configured to limit optical loss associated with metal layer(s) formed at a surface of the device.
In some embodiments, the integrated device may be configured to excite individual samples with substantially uniform intensities (e.g., with a variation that is less than 10%). Having a substantially uniform excitation across the samples may improve the likelihood of the emission energy emitted by the samples being within the dynamic range of the sensors. An optical waveguide, including one according to the techniques described herein may be configured to provide an optical coupling to the sample wells that varies along its length so as to provide a substantially uniform excitation across samples located within the sample wells. According to some non-limiting embodiments, the width of the waveguide may vary along the length of the waveguide, thus providing a position-dependent mode profile. In some embodiments, a waveguide having one or more dimensions that vary along the length of the waveguide may be implemented. For example, a device according to some embodiments may include a waveguide having a tapered width that varies along the length of the waveguide.
For channel waveguides, the coupling coefficient may be increased for decreased waveguide width. Thus, a tapered channel waveguide will decrease in width along the direction of propagation to increase the coupling coefficient and provide compensation for optical loss. In some embodiments, a channel waveguide may have a taper with a dimension in the range of 600 nm to 1500 nm, or any value or range of values in that range, at the start of the taper and a dimension in the range of 200 nm to 500 nm, or any value or range of values in that range, at the end of the taper.
For rib waveguides and ridge waveguides, the coupling coefficient may increase with increased width of the raised region, WRR. Thus, a tapered rib/ridge waveguide may increase in WRR along the direction of propagation to increase the coupling coefficient and provide compensation for optical loss. In some embodiments, WRRIN may be in a range between 150 nm and 500 nm, or any value or range of values in that range. In some embodiments, WRROUT may be between 100 nm and 200 nm, or any value or range of values in that range.
A waveguide of the type described herein may be configured to limit optical loss associated with proximity to metal layer(s). In some embodiments, a waveguide may have a configuration, for example, to enhance the decay rate of the evanescent field. Compared to channel waveguides having rectangular cross sections, ridge or rib waveguides may exhibit a greater decay rate of the evanescent field.
Some embodiments relate to an integrated device having one or more waveguides configured to support multiple modes. The two or more modes of such a multimode waveguide may combine through interference of the modes in a manner where the power distribution of excitation energy varies in a direction perpendicular to the direction of light propagation along the multimode waveguide. The variation in power distribution may include regions along the direction of light propagation where the power distribution is broader in one or more directions perpendicular to the direction of light propagation than in other regions. In some embodiments, the power distribution may broaden in a direction towards a sample well in a region of the multimode waveguide proximate to the sample well. The broadening of the power distribution of excitation energy may improve coupling of excitation energy to the sample well. In some embodiments, the power distribution may decrease along the direction in a region of the multimode waveguide that is non-overlapping with the sample well. The decrease in the power distribution may reduce optical loss of excitation energy by reducing an amount of excitation energy that extends outside the waveguide. In some embodiments, two or more modes may interfere to beat with a characteristic beat length. The characteristic beat length may depend on the type of modes being combined by the multimode waveguide. In some embodiments, the characteristic beat length may be substantially similar to a distance between neighboring sample wells of the integrated device. The multi-mode waveguide may be configured to support any suitable number of modes (e.g., 2, 3, 4), type of modes (e.g., TE, TM) and/or order of the modes (e.g., 1st, 3rd). In some embodiments, the multimode waveguide combines first and third order TE modes of the excitation energy.
Formation of an integrated device of the type described herein may use various fabrication techniques, some of which may be performed within a standard semiconductor foundry. In some embodiments, conventional complementary metal-oxide-semiconductor (CMOS) fabrication techniques may be used. For example, at least some of the following fabrication techniques may be used: photolithography, wet etching, dry etching, planarization, metal deposition, chemical vapor deposition, atomic layer deposition, oxidation, annealing, epitaxial growth, ion implantation, diffusion, wire bonding, flip-chip bonding, etc.
Formation of an integrated device may include a plurality of photolithographic process steps. Each photolithographic process step may comprise an exposure to ultra-violet (UV) light through a photomask, a development process to form a relief image in the photoresist, and an etch process to transfer the photoresist relief image into at least one underlying layer. The photomask may be positive or negative, and may be patterned according to a desired configuration. For example, one or more photolithographic process steps may be used to form waveguides of the type described here. Additionally, one or more photolithographic process steps may be used to form sample wells of the type described here.
Fabrication of a rib waveguide, such as waveguide 4-200, may be performed using a variety of different processes. Regardless of the particular process utilized, the fabrication may comprise a photolithographic process step to form a raised region. Accordingly, following an exposure to UV light and subsequent development of the relief image, a partial etch process may be performed to form the raised region while retaining at least a portion of the slab.
In some embodiments, formation of an integrated device may include a timed etch process. The timed etch process may be used to form a rib waveguide. The duration of the etch process may be selected so as to remove a desired amount of dielectric material from the slab. Accordingly, based on the duration of the timed etch process, a desired ratio TRR/Ts may be defined. Formation of a rib waveguide based on a timed etch process may utilize a photolithographic fabrication step.
In the fabrication step illustrated in
In the fabrication step illustrated in
In the fabrication step illustrated in
Some embodiments relate to another technique to fabricate a rib waveguide of the type described herein. Unlike the fabrication process illustrated in
In the fabrication step illustrated in
In the fabrication step illustrated in
In the fabrication step illustrated in
Some embodiments relate to yet another technique to fabricate a rib waveguide of the type described herein. Such fabrication technique may utilize an endpoint layer. According to one such technique, light may be shined toward a surface of the substrate throughout the duration of the etch process. Reflected light may be sensed during the etch process. When the endpoint layer is at least partially uncovered, the reflected light may exhibit a recognizable pattern, such as a polarization pattern and/or an interference pattern and/or an optical intensity that is above or below a predetermined threshold. When the recognizable pattern is sensed, the etch process may be arrested. In this way, the thickness of the etched region may be finely controlled. Similarly to the fabrication technique illustrated in
In the fabrication step illustrated in
In the fabrication step illustrated in
In the fabrication step illustrated in
Some embodiments of the present application relate to techniques for forming a ridge waveguide, such as waveguide 4-250 of
In the fabrication step illustrated in
In the fabrication step illustrated in
In the fabrication step illustrated in
Having thus described several aspects and embodiments of the technology of this application, it is to be appreciated that various alterations, modifications, and improvements will readily occur to those of ordinary skill in the art. Such alterations, modifications, and improvements are intended to be within the spirit and scope of the technology described in the application. It is, therefore, to be understood that the foregoing embodiments are presented by way of example only and that, within the scope of the appended claims and equivalents thereto, inventive embodiments may be practiced otherwise than as specifically described. In addition, any combination of two or more features, systems, articles, materials, kits, and/or methods described herein, if such features, systems, articles, materials, kits, and/or methods are not mutually inconsistent, is included within the scope of the present disclosure.
Also, as described, some aspects may be embodied as one or more methods. The acts performed as part of the method may be ordered in any suitable way. Accordingly, embodiments may be constructed in which acts are performed in an order different than illustrated, which may include performing some acts simultaneously, even though shown as sequential acts in illustrative embodiments.
All definitions, as defined and used herein, should be understood to control over dictionary definitions, definitions in documents incorporated by reference, and/or ordinary meanings of the defined terms.
The indefinite articles “a” and “an,” as used herein in the specification and in the claims, unless clearly indicated to the contrary, should be understood to mean “at least one.”
The phrase “and/or,” as used herein in the specification and in the claims, should be understood to mean “either or both” of the elements so conjoined, i.e., elements that are conjunctively present in some cases and disjunctively present in other cases.
As used herein in the specification and in the claims, the phrase “at least one,” in reference to a list of one or more elements, should be understood to mean at least one element selected from any one or more of the elements in the list of elements, but not necessarily including at least one of each and every element specifically listed within the list of elements and not excluding any combinations of elements in the list of elements. This definition also allows that elements may optionally be present other than the elements specifically identified within the list of elements to which the phrase “at least one” refers, whether related or unrelated to those elements specifically identified.
In the claims, as well as in the specification above, all transitional phrases such as “comprising,” “including,” “carrying,” “having,” “containing,” “involving,” “holding,” “composed of,” and the like are to be understood to be open-ended, i.e., to mean including but not limited to. The transitional phrases “consisting of” and “consisting essentially of” shall be closed or semi-closed transitional phrases, respectively.
This application is a Continuation of U.S. application Ser. No. 15/844,403, filed Dec. 15, 2017, entitled “OPTICAL COUPLER AND WAVEGUIDE SYSTEM”, which is a Non-Prov of Prov (35 USC 119(e)) of U.S. Application Ser. No. 62/435,693, filed Dec. 16, 2016, entitled “OPTICAL COUPLER AND WAVEGUIDE SYSTEM”. The entire contents of these applications are incorporated herein by reference in their entirety.
| Number | Date | Country | |
|---|---|---|---|
| 62435693 | Dec 2016 | US |
| Number | Date | Country | |
|---|---|---|---|
| Parent | 15844403 | Dec 2017 | US |
| Child | 18138700 | US |
