阅读说明：本技术 用平面波导制造流动池 (Fabrication of flow cells with planar waveguides ) 是由 袁大军 M.S.博温 M.钟 于 2020-05-28 设计创作，主要内容包括：在一个实例中提供了制造流动池的方法,该方法包括：形成核心层,所述核心层被设置在基底和纳米孔层之间,所述纳米孔层具有纳米孔以接收样品,所述核心层具有比基底和纳米孔层高的折射率；和形成光栅以将光耦合到核心层。(In one example, a method of manufacturing a flow cell is provided, the method comprising: forming a core layer disposed between a substrate and a nanopore layer, the nanopore layer having a nanopore to receive a sample, the core layer having a higher refractive index than the substrate and the nanopore layer; and forming a grating to couple light into the core layer.)

1. A method of manufacturing a flow cell, the method comprising:

forming a core layer disposed between a substrate and a nanopore layer having a nanopore to receive a sample, the core layer having a higher refractive index than the substrate and the nanopore layer; and

a grating is formed using nanoimprinting to couple light to the core layer.

2. The method of claim 1, wherein the core layer is formed in a common process with forming the grating.

3. The method of claim 1, wherein the core layer is formed in a different process than forming the grating.

4. The method of claim 3, wherein the grating is formed onto the substrate by nanoimprinting, the core layer is formed onto the grating, and the nanopore layer is formed onto the core layer.

5. The method of claim 3, wherein the grating is formed onto the substrate, the core layer is formed onto the grating, an additional layer is formed onto the core layer, and the nanopore layer is formed onto the additional layer.

6. The method of claim 5, wherein the additional layer and the nanopore layer are initially free of the nanopore prior to forming the nanopore, the method further comprising patterning the nanopore layer to form the nanopore with the additional layer remaining intact, and subsequently transferring the pattern of the nanopore layer to the additional layer to expose the core layer in the nanopore.

7. The method of claim 6, wherein the pattern is transferred by etching.

8. The method of claim 1, wherein the core layer is formed onto the substrate, and wherein the grating and the nanopore layer are formed onto the core layer.

9. The method of claim 3, wherein the core layer is formed onto the substrate, the grating is formed onto the core layer, and the nanopore layer is formed onto the grating.

10. The method of claim 3, wherein the core layer is formed onto the substrate, a first layer is formed onto the core layer, a second layer is formed onto the first layer, and wherein the grating and the nanopore are formed into the first and second layers, respectively.

11. The method of claim 3, wherein the core layer is formed onto the substrate, a resin layer is formed onto the core layer, and the grating and the nanopore are formed into the resin layer.

12. The method of any of claims 1-3, 8, or 10, wherein the grating and the nanopore layer are formed in the same process.

13. The method of claim 12, wherein the grating and the nanopore layer are formed in the same layer of the flow cell.

14. The method of claim 12, wherein the grating and the nanopore layer are formed in separate layers of a flow cell.

15. A flow cell manufactured using the method of any preceding claim, comprising:

a substrate;

a nanopore layer having a nanopore to receive a sample;

a core layer disposed between the substrate and the nanopore layer, the core layer having a higher refractive index than the substrate and the nanopore layer; and

a grating to couple light to the core layer.

16. The flow cell of claim 15, wherein the grating is disposed over the substrate, the core layer is disposed over the grating, and the nanopore layer is disposed over the core layer.

17. The flow cell of claim 16, wherein a grating layer overlies the substrate, the grating layer including the grating.

18. The flow cell of claim 15, wherein a first resin layer comprising the grating is disposed over the substrate and a second resin layer comprising the nanopore is disposed over the first resin layer.

19. The flow cell of claim 15, wherein the grating is disposed over the substrate, a first polymer layer is disposed over the grating, and a second polymer layer is disposed over the first polymer layer, wherein the nanopores are disposed in the first and second polymer layers.

20. The flow cell of claim 15, wherein the core layer is disposed over the substrate, a resin layer is disposed over the core layer, and the grating and nanopore are disposed in the resin layer.

21. The flow cell of claim 15, wherein the core layer is disposed over the substrate, the grating is disposed over the core layer, and the nanopore layer is disposed over the grating.

22. The flow cell of claim 21 wherein a grating layer overlies a core layer, the grating layer comprising the grating.

23. The flow cell of claim 15, wherein the core layer is disposed over the substrate, a polymer layer is disposed over the core layer, and a resin layer is disposed over the polymer layer, wherein the grating is disposed in the polymer layer, and wherein the nanopore is disposed in the resin layer.

24. The flow cell of claim 15, wherein a polymer layer is disposed over the substrate and a resin layer is disposed over the polymer layer, wherein the grating is disposed in the polymer layer, and wherein the nanopore is disposed in the resin layer.

Background

Samples of different materials may be analyzed using one or more of a variety of analytical methods. For example, sequencing, such as high-throughput DNA sequencing, can be the basis for genomic analysis and other genetic studies. For example, modified deoxyribonucleotide triphosphates (dntps) are used by sequencing-by-synthesis (SBS) techniques, which include a terminator and a fluorescent dye having an emission spectrum. In this and other types of sequencing, characteristics of a sample of genetic material are determined by illuminating the sample and detecting emitted light (e.g., fluorescence) generated in response to the illumination. The quality of the illumination may determine the quality and efficiency of the detection of the emitted light. For example, if a large portion of the illumination light does not reach the relevant sample material, a low level of efficiency in the system may result. As another example, if the illumination light inadvertently strikes (imprge) other portions of the sample, it may cause the sample to degrade.

Summary of The Invention

In a first aspect, a method of making a flow cell includes: forming a core layer (core layer) disposed between a substrate and a nanopore layer having a nanopore to receive a sample, the core layer having a higher refractive index than the substrate and the nanopore layer; and forms a grating to couple light into the core layer.

Implementations may include any or all of the following features in any suitable combination. Forming the grating includes lithographic patterning, nanoimprinting, or both. The core layer is formed in the same process as the grating is formed. The core layer is formed in a different process than the formation of the grating. A grating is formed by photolithographic patterning on a substrate, a core layer is formed on the grating, and a nanopore layer is formed on the core layer. A grating is formed by nanoimprinting on a substrate, a core layer is formed on the grating, and a nanopore layer is formed on the core layer. A grating is formed on a substrate, a core layer is formed on the grating, an additional layer is formed on the core layer, and a nanopore layer is formed on the additional layer. The method further includes patterning the nanopore layer to form the nanopore while the additional layer remains intact, and then transferring the pattern of the nanopore layer to the additional layer to expose the core layer in the nanopore. The pattern is transferred by etching. A core layer is formed on a substrate, and wherein a grating and a nanopore layer are formed on the core layer. A core layer is formed on a substrate, a grating is formed on the core layer, and a nanopore layer is formed on the grating. The method includes forming a core layer on a substrate, forming a first layer on the core layer, forming a second layer on the first layer, and wherein a grating and a nanopore are formed on the first layer and the second layer, respectively. A core layer is formed on a substrate, a resin layer is formed on the core layer, and a grating and a nanopore are formed in the resin layer. The grating and the nanopore layer are formed in the same process. The grating and the nanopore layer are formed in the same layer of the flow cell. The grating and the nanopore layer are formed in different layers of the flow cell.

In a second aspect, a flow cell comprises: a substrate; a nanopore layer having a nanopore to receive a sample; a core layer disposed between the substrate and the nanopore layer, the core layer having a higher refractive index than the substrate and the nanopore layer; and a grating coupling light to the core layer.

Implementations may include any or all of the following features. A grating is disposed on a substrate, a core layer is disposed on the grating, and a nanopore layer is disposed on the core layer. A grating layer overlies the substrate, the grating layer including a grating. A first resin layer including a grating is disposed on a substrate, and a second resin layer including a nanopore is disposed on the first resin layer. A grating is disposed on a substrate, a first polymer layer is disposed on the grating, and a second polymer layer is disposed on the first polymer layer, wherein nanopores are disposed in the first and second polymer layers. A core layer is disposed on a substrate, a resin layer is disposed on the core layer, and a grating and a nanopore are disposed in the resin layer. A core layer is disposed on a substrate, a grating is disposed on the core layer, and a nanopore layer is disposed on the grating. A grating layer overlies the core layer, the grating layer including a grating. A core layer is disposed on a substrate, a polymer layer is disposed on the core layer, and a resin layer is disposed on the polymer layer, wherein a grating is disposed in the polymer layer, and wherein a nanopore is disposed in the resin layer. A polymer layer is disposed on a substrate, and a resin layer is disposed on the polymer layer, wherein a grating is disposed in the polymer layer, and a nanopore is disposed in the resin layer.

It should be understood that all combinations of the foregoing concepts and additional concepts discussed in greater detail below (provided that such concepts do not contradict each other) are considered to be part of the inventive subject matter disclosed herein and that the benefits described herein can be achieved. In particular, all combinations of claimed subject matter appearing at the end of this disclosure are considered part of the inventive subject matter disclosed herein and the benefits described herein can be realized.

Brief Description of Drawings

Fig. 1 shows an example of fabricating a planar waveguide.

Fig. 2 shows a method of manufacturing a planar waveguide in relation to the planar waveguide of fig. 1.

Fig. 3 shows an example of fabricating a planar waveguide.

Fig. 4 illustrates a method of manufacturing a planar waveguide in connection with the planar waveguide of fig. 3.

Fig. 5 shows an example of fabricating a planar waveguide.

Fig. 6 illustrates a method of manufacturing a planar waveguide in relation to the planar waveguide of fig. 5.

Fig. 7 shows an example of fabricating a planar waveguide.

Fig. 8 illustrates a method of manufacturing a planar waveguide in connection with the planar waveguide of fig. 7.

Fig. 9 shows an example of fabricating a planar waveguide.

Fig. 10 illustrates a method of manufacturing the planar waveguide related to the planar waveguide of fig. 9.

Fig. 11 shows an example of fabricating a planar waveguide.

Fig. 12 shows a method of manufacturing the planar waveguide related to the planar waveguide in fig. 11.

Fig. 13 shows an example of fabricating a planar waveguide.

Fig. 14 illustrates a method of manufacturing the planar waveguide related to the planar waveguide of fig. 13.

Fig. 15 shows an example of fabricating a planar waveguide.

Fig. 16 shows a method of manufacturing the planar waveguide related to the planar waveguide in fig. 15.

Fig. 17 shows an example of a flow cell.

Fig. 18 is a diagram of a system including an instrument, a cartridge, and a flow cell.

Fig. 19 is a diagram of an example lighting system.

Detailed Description

The present disclosure describes systems, techniques, articles of manufacture, and/or compositions of matter that facilitate improved sample analysis. Sample analysis may include, but is not limited to, genetic sequencing (e.g., determining the structure of genetic material), genotyping (e.g., determining differences in individual genetic make-up), gene expression (e.g., synthesizing gene products using genetic information), proteomics (e.g., large scale studies of proteins), or a combination thereof. As described herein, substrates for holding samples during analysis can be manufactured in a more efficient manner, and/or can have improved properties. In some implementations, flow cells may have improved structures that facilitate efficient use of illumination light to excite active elements (e.g., fluorophores) in a sample. For example, the structure may include a core layer disposed between the substrate and the nanopore layer, and a grating that couples light (i.e., illumination or excitation light) to the core layer. The materials of the flow cell structure may be chosen such that their respective refractive indices have a favorable ratio with respect to each other. The core layer may help to efficiently use the illumination light for exciting the sample. For example, the structure may be based on the principle of Total Internal Reflection (TIR) and may be designed such that evanescent light (evanescent light) reaches the sample in one or more dedicated regions (e.g. nanopores) in an efficient manner.

As scan speed increases in sample analysis, and the corresponding data density increases, systems can be designed with higher levels of illumination (e.g., laser) power. This results in higher instrument costs. However, as a result of this development, damage to optics and substrates (e.g., flow cells) can increase. Some implementations, such as those associated with planar waveguides, may be designed with a view to improving excitation efficiency and/or reducing background noise. For example, a planar waveguide flow cell may include a substrate (e.g., of glass), one or more optical coupling gratings, a high refractive index core layer, and an aqueous buffer or patterned polymer cladding (cladding layer). Improved fabrication techniques for such flow cells and/or structures thereof are described in this disclosure.

Some embodiments described herein relate to sequencing of genetic material. Sequencing can be performed on a sample to determine which building blocks (called nucleotides) constitute a particular genetic material in the sample. The genetic material may be purified and then repeated a number of times to prepare a sample of suitable size before sequencing.

Imaging may be performed as part of a process of analyzing sample material. This may involve fluorescence imaging, such as when a sample of genetic material is illuminated with light (e.g., a laser beam), triggering a fluorescence reaction by one or more markers on the genetic material. Some nucleotides of genetic material may have a fluorescent tag applied thereto which allows the presence of the nucleotide to be determined by shining light onto the sample and looking for a characteristic reaction from the sample. The fluorescent reaction can be detected during the course of the assay and used to establish a record of the nucleotides in the sample.

The examples described herein relate to flow cells. A flow cell is a substrate that can be used to prepare and contain or carry one or more samples during at least one stage of an analytical process. The flow cell is made of a material compatible with the genetic material, the lighting and the chemical reaction to be exposed. The substrate may have one or more channels in which sample material may be deposited. Material (e.g., liquid) may flow through the channel in which the sample genetic material is present to trigger one or more chemical reactions and/or remove unwanted material. The flow cell may enable imaging by facilitating that the sample in the flow cell channel can be subjected to illumination light and any fluorescence reaction from the sample can be detected. Some embodiments of the system may be designed for use with at least one flow cell, but may not include a flow cell during one or more stages (e.g., during shipping or when shipped to a customer). For example, the flow cell may be installed in a customer-preset implementation for performing the analysis.

Examples herein relate to coupling light (e.g., a laser beam) into and/or out of a waveguide through one or more gratings. The grating may couple light impinging on the grating by diffracting at least a portion of the light, thereby causing a portion of the light to propagate in one or more other directions. In some embodiments, coupling may involve one or more interactions, including but not limited to reflection, refraction, and/or transmission of a portion of light. Embodiments may be designed to meet one or more requirements, including but not limited to requirements related to mass production (mass production), cost control, and/or high optical coupling efficiency.

Providing a planar waveguide in a substrate (e.g., a flow cell) may provide one or more advantages. The use of TIR-based evanescent light excitation may provide higher illumination efficiency. In some previous methods, the entire laser beam is used to irradiate the substrate holding the sample, such as during scanning. This method may cause a large portion of the light waves to propagate through the substrate without effectively illuminating the sample. As a result, only a small portion of the light applied by such a system may actually be used to excite fluorophores in the sample. In contrast, evanescent light may penetrate only a certain depth (e.g., between about 150nm to about 200nm, such as between about 165nm to about 185nm in one example; in some examples, the depth may be about 155nm, about 170nm, about 180nm, about 195nm, etc.) through a material (e.g., a cladding layer adjacent to a core layer). For example, the flow cell may be designed to configure one or more nanopores so that the evanescent field is largely confined to the region of the pore. As a result, evanescent light may be a very efficient method of exciting fluorophores. For example, a system operating according to an earlier illumination method may require a laser with a certain power; in contrast, with evanescent light, a much lower laser power may be sufficient.

The examples described herein refer to one or more gratings that may provide coupling for light. The gratings may be identical or similar to each other or may be different types of gratings. The grating may comprise one or more forms of periodic structures. In some embodiments, the grating may be formed by removing or omitting material from the substrate (e.g., from waveguide material included in the flow cell) or other material. For example, the flow cell may be provided with a set of slits (slit) and/or grooves (groove) therein to form a grating. In some embodiments, the grating may be formed by adding a substance to the flow cell (e.g., to a waveguide material included in the flow cell) or other material. For example, the flow cell may be provided with a set of ridges (ridges), strips or other protruding longitudinal structures to form a grating. A combination of these approaches may be used.

Examples herein relate to photolithography. Photolithography-based methods may involve the use of a photoresist (photoresist) that is patterned by a stepper or mask aligner (stepper), the pattern present on the reticle/photomask (reticle/photomask) is transferred into the photoresist by radiation exposure, and then developed to create a structured film (photoresist) on top of the substrate. The structured resist may be the final substrate that may be used for subsequent coating of the core layer. As another example, the pattern in the resist may be transferred into the substrate or other material by additional processing. The subsequent process (follow on process) operation may include a reactive ion etch (plasma based dry etch) or a wet etch (chemical based) process. If the pattern is transferred into a substrate/material, the patterned photoresist is then removed to produce a patterned substrate (e.g., for subsequent core layer coating). In the photoresistA sacrificial film of an underlying material, such as chromium or titanium or another metal, first transfers the pattern in the photoresist to the metal film, which then serves as a hard mask through which the pattern is transferred to the substrate. After the pattern is transferred into the substrate, the film can be removed and is therefore considered sacrificial to the fabrication process. One or more of a variety of materials may be used in the lithographic process. In some embodiments, an oxide material is used. For example, SiO can be used₂(silica). The lift-off process may be similar to the patterned photoresist process: instead of removing material by dry or wet etching, a material (e.g. SiO) may be deposited₂) And then stripped, which involves removing the photoresist and deposited material from above. The grating structure may also or alternatively be formed.

Examples herein relate to sputtering (sputtering). Sputter deposition may refer to a Physical Vapor Deposition (PVD) process for depositing thin films or coatings. Such processes may involve causing material to be ejected from a source and deposited onto a substrate. In some embodiments, the sputter deposition forms a thin layer of waveguides on the surface of the substrate. One or more of a variety of materials may be used in sputtering. Waveguide materials used in sputter deposition may include metals and metal oxides having high refractive indices and low adsorption characteristics. For example, the waveguide material may include tantalum pentoxide (Ta)₂O₅) Or silicon nitride (e.g., one or more compounds represented by the formula SiNx, including but not limited to Si₃N₄). When patterning the lower surface, the waveguide layer/coating may adopt a pattern in the lower surface, thereby forming optical features on the surface of the substrate. This may enable manipulation of light on the substrate surface in downstream imaging processes based on the design of the pattern.

Examples herein relate to chemical vapor deposition. Chemical Vapor Deposition (CVD) may include all techniques in which a volatile material (sometimes referred to as a precursor) is caused to react and/or decompose on the surface of a substrate on which a deposit is formed. CVD can be characterized by one or more aspects. For example, CVD can be characterized by the physical characteristics of a vapor (vapor) (e.g., whether CVD is aerosol-assisted or involves direct liquid injection). For example, CVD can be characterized by the type of substrate heating (e.g., whether the substrate is directly heated or indirectly heated, such as by a heating chamber). Examples of types of CVD that can be used include, but are not limited to, atmospheric pressure CVD, low pressure CVD, ultra high vacuum CVD, metal organic CVD, laser assisted CVD, and plasma enhanced CVD.

Examples herein relate to atomic layer deposition. Atomic layer deposition can be considered a form of CVD and includes all techniques in which a thin film is grown on a substrate by exposure to a gas. For example, gaseous precursors may be alternately introduced into the chamber. Molecules of one of the precursors may react with the surface until the layer is formed and the reaction is terminated, and then the next gaseous precursor may be introduced to begin forming a new layer and one or more cycles may be followed.

Examples herein relate to spraying (spray coating). Spraying may include any or all techniques by which a particular material is caused to be deposited onto a substrate. This may include, but is not limited to, thermal spraying, plasma spraying, cold spraying, warm spraying, and/or other procedures involving atomized or atomized materials.

Examples herein relate to spin coating. Spin coating may include applying an amount of coating to a substrate and dispensing or spreading the coating on the substrate by centrifugal force due to rotation or spinning of the substrate.

Examples herein relate to nanoimprinting (nanoimprinting). In nanoimprint lithography, a pre-fabricated nanoscale template can mechanically displace a fluid resin to mold (mold) the desired nanostructures. The resin may then be cured (cured) using a nanoscale template in place. After removal of the nanoscale template, a molded solid resin can be produced that adheres to the desired substrate. In some embodiments, the nanoimprinting process may begin by completely or partially covering a substrate or wafer with an imprint resin (e.g., the resins exemplified below) (wafer). The nano-scale template may be used to form one or more nano-structures in an imprint resin in a molding process. The imprint resin may be cured against the substrate or wafer, and a resin removal process may be applied to remove residue from the wafer or substrate. For example, resin removal can form a chamber channel adjacent to the nanostructure. The substrate or wafer so formed may have another substrate or gasket (gasket) applied thereto to form a flow cell with the nanostructures and a flow cell chamber formed by closing the chamber channel. In some embodiments, the process of applying the imprint resin may be configured to produce little or no resin residue, and in such embodiments, the resin removal process may be omitted. In some applications, the cured resin may also be functionalized by chemical treatment or attachment of biomolecules, depending on the end use. In nanoimprint lithography, the imprinted photoresist may be a sacrificial material and similarly used as an intermediate tool for transferring the patterned resist into the substrate, or variations of the resist may be used, such that the imprinted resist is used as an input (input) for the subsequent coating process. One example of a resist that will remain after patterning is a material formed by a process that converts monomers into a colloidal solution as a precursor to particles and/or polymer gels, sometimes referred to as a sol-gel based material.

The examples described herein mention that one or more resins may be used. Any suitable resin may be used for nanoimprinting in the methods described herein. In some embodiments, organic resins may be used, including, but not limited to, acrylic (acrylic) resins, polyimide (polyimide) resins, melamine (melamine) resins, polyester resins, polycarbonate (polycarbonate) resins, phenol resins, epoxy (epoxy) resins, polyacetal (polyacetal) resins, polyether resins, polyurethane (polyurethane) resins, polyamide resins (and/or nylons), furan resins, diallyl phthalate (diallyl phthalate) resins, or combinations thereof. In some embodiments, the resin may include an inorganic siloxane polymer including Si-O-Si bonds between compounds (including silicon, oxygen, and hydrogen), and is formed by using a siloxane polymer-based material represented by silica glass as a starting material. The resin used may also or alternatively be an organosiloxane polymer in which the hydrogen bonded to the silicon is substituted with an organic group, such as a methyl or phenyl group, and is represented by an alkylsiloxane (alkylsiloxane) polymer, an alkylsilsequioxane (alkylsilsequioxane) polymer, a silsesquioxane hydride (silsesquioxane hydride) polymer or an alkylsilsequioxane hydride (alkylsilsequioxane) polymer. Non-limiting examples of siloxane polymers include polyhedral oligomeric silsesquioxane (POSS), Polydimethylsiloxane (PDMS), tetraethyl orthosilicate (TEOS), poly (organo) siloxane (silicone), and perfluoropolyether (PFPE). An example of a POSS may be that described in Kehagias et al, microelectronic Engineering 86(2009), page 776-778, which is incorporated herein by reference in its entirety. The resin may be doped with a metal oxide. In some embodiments, the resin may be a sol-gel material including, but not limited to, titanium oxide, hafnium oxide (hafnium oxide), zirconium oxide (zirconia oxide), tin oxide (tin oxide), zinc oxide, or germanium oxide (germanium oxide), and using a suitable solvent. Any of a variety of other resins may be used as appropriate depending on the application.

Examples herein relate to substrates. A substrate may refer to any material that provides a substantially rigid structure, or a structure that retains its shape, rather than adopting the shape of a container in which it is placed in contact. The material may have a surface to which another material may be attached, including, for example, smooth supports (e.g., metal, glass, plastic, silicon, and ceramic surfaces), and textured and/or porous materials. Possible substrates include, but are not limited to, glass and modified or functionalized glass, plastics (including acrylic, polystyrene and copolymers of styrene with other materials), polypropylene, polyethylene, polybutylene, polyurethane, Teflon^TMEtc.), polysaccharides, nylon or nitrocellulose, resins, silica or silica-based materials including silicon and modified silicon, carbon, metals, inorganic glass, plastics, fiber optic strands, and a variety of other polymers. In general, the substrate allows optical detection and does not itself emit significant emissionsFluorescence of (2).

Examples herein refer to polymers. The polymer layer may comprise a film of a polymer material. Exemplary film forming polymers include, but are not limited to, acrylamide or copolymers with C1-C12; aromatic and hydroxy derivatives; an acrylate copolymer; copolymers of vinylpyrrolidone (vinylpyrrolidine) and vinylpyrrolidone (vinylpyrrolidine); sugar-based polymers, such as starch or polydextrose; or other polymers such as polyacrylic acid, polyethylene glycol, polylactic acid, silicone, siloxanes, polyvinylamine, guar gum (guar gum), carrageenan (carrageenan), alginate (alginate), lotus bean gum, methacrylate copolymers, polyimide, cyclic olefin (olefin) copolymers, or combinations thereof. In some embodiments, the polymer layer comprises at least one photocurable polymer. For example, the photocurable polymer may include urethane (urethane), acrylate, silicone, epoxy, polyacrylic, polyacrylate, epoxysilicone (epoxysilicone), epoxy, Polydimethylsiloxane (PDMS), silsesquioxane (silsequioxane), acyloxysilane, maleate polyester, vinyl ether, monomers or copolymers having vinyl or ethynyl groups, or combinations thereof. In some embodiments, the layer may comprise a covalently attached polymer coating. For example, it may include a polymer coating that forms a chemical bond with the functionalized surface of the substrate, as opposed to being attached to the surface in other ways, such as adhesion or electrostatic interaction. In some embodiments, the polymer contained in the functionalizable layer is poly (N- (5-azidoacetamidylpentyl) acrylamide-co-acrylamide), sometimes referred to as PAZAM.

Fig. 1 shows an example of fabricating a planar waveguide 100. The planar waveguide 100 may be used in one or more of the examples described herein. For example, the planar waveguide 100 may house one or more samples to facilitate illumination and excitation during sample analysis. Fig. 2 illustrates a method 200 of fabricating a planar waveguide in relation to the planar waveguide of fig. 1. Method 200 may be combined with one or more of the other methods described herein. More or fewer operations may be performed, and/or two or more operations may be performed in a different order, unless otherwise indicated.

At 210, lithographic (PL) patterning of the grating may be performed. In some embodiments, the substrate 102 is provided with one or more gratings 104. For example, grating 104 may be a laser coupling grating. In some embodiments, the lithographic pattern may include a deposition or etching process. For example, it may be on oxide (e.g., SiO)₂) And carrying out patterning. Here the gratings 104 are shown longitudinally such that the respective ridges 104' extend into the plane of the drawing. The grating layer 104 may include a plurality of ridges 104', for example, organized in respective groups 106A and 106B. For example, the groups 106A-106B may be positioned such that the region 108 of the substrate 102 is substantially free of the grating 104. The ridges 104' may have a pitch (pitch) of about 200nm to about 300 nm. Such as between about 220nm and about 280nm, to name just one example. In some embodiments, the pitch may be about 205nm, about 215nm, about 235nm, about 265nm, about 285nm, and the like.

At 220, a core layer may be formed. In some embodiments, the core layer 110 is formed on the substrate 102. In some embodiments, the core layer 110 is formed on the grating 104. In some embodiments, the substrate 102 may function as a cladding layer for the core layer 110. The core layer 110 may have a higher refractive index than the substrate 102. For example, the substrate 102 may have a refractive index of about 1.5, while the core layer may have a refractive index of about 2.2, or in a range of greater than about 1.5 to 2.2, such as about 1.6 to about 2.1. In some embodiments, the refractive index may be about 1.65, about 1.85, about 2.05, and the like. The core layer 110 may cover substantially the entire facing surface of the substrate 102. The core layer 110 may have a higher refractive index than the grating layer 104. In some embodiments, the core layer 110 includes Ta₂O₅And/or SiN_x. For example, the core layer 110 may be formed by sputtering, chemical vapor deposition, atomic layer deposition, spin coating, and/or spray coating.

At 230, nanopore layer patterning is performed. In some embodiments, the nanopore layer 112 is formed at the core layer 110. The nanopore layer 112 may facilitate a patterned flow cell. The nanopore layer 112 may be included in two or moreOne or more nanopores 114 defined between the walls 116. In some embodiments, the nanopore 114 may have a size such that one or more dimensions thereof range in a range of about one or more nanometers. For example, the nanopore 114 may be configured to receive and hold a sample during an analysis process, such as in a cluster. The end (e.g., bottom) of the nanopore 114 may have a thickness that accommodates propagation of evanescent light. For example, the thickness may be from about 0 to about 500nm, such as between about 100nm to about 400 nm. In some embodiments, the thickness may be about 10nm, about 50nm, about 100nm, about 200nm, about 300nm, about 450nm, and the like. The nanopore layer 112 may be formed by a nanoimprint process or a lift-off process. In some embodiments, the nanopore layer 112 may comprise one or more resins. The nanopore layer may cover substantially the entire facing surface of the core layer 110. For example, the resin may have a refractive index of about 1.5. In some embodiments, the nanopore layer 112 may have an average pitch between the nanopores 114 of at least about 10nm, about 0.1 μm, about 0.5 μm, about 1 μm, about 5 μm, about 10 μm, about 100 μm, or more, and/or may have an average pitch of at most about 100 μm, about 10 μm, about 5 μm, about 1 μm, about 0.5 μm, about 0.1 μm, or less. For example, the nanopore layer 112 may have a pitch between the nanopores 114 of about 600 to about 650nm, for example, the pitch may be about 610nm to about 640 nm. In some embodiments, the pitch may be about 605nm, about 615nm, about 635nm, about 655nm, and the like. The depth of each nanopore 114 may be at least about 0.1 μm, about 1 μm, about 10 μm, about 100 μm, or more. Alternatively or additionally, the depth may be up to about 1 × 10³μ m, about 100 μm, about 10 μm, about 1 μm, about 0.1 μm, or less.

Fig. 3 shows an example of making a planar waveguide 300. The planar waveguide 300 may be used in one or more of the examples described herein. For example, the planar waveguide 300 may house one or more samples to facilitate illumination and excitation during sample analysis. Fig. 4 illustrates a method 400 of fabricating a planar waveguide in relation to the planar waveguide of fig. 3. Method 400 may be combined with one or more of the other methods described herein. More or fewer operations may be performed, and/or two or more operations may be performed in a different order, unless otherwise indicated.

At 410, nanoimprinting of the grating may be performed. In some embodiments, substrate 302 is provided with one or more grating layers 304. For example, grating layer 304 may include a laser coupling grating. For example, the resin may be nanoimprinted. The grating layer 304 is here shown longitudinally, so that the respective ridges 304' extend into the plane of the drawing. Grating layer 304 may include a plurality of ridges 304', for example, organized in respective groups 306A and 306B. For example, groups 306A-306B may be separated by regions 308 of grating layer 304 that also cover the surface of substrate 302. The region 308 may form a thin residual layer on the surface of the substrate 302. The ridges 304 may have a pitch of about 200nm to about 300nm, such as between about 220nm to about 280nm, to name a few. In some embodiments, the pitch may be about 205nm, about 215nm, about 235nm, about 265nm, about 285nm, and the like. Applying grating layer 304 via a nanoimprinting process may provide one or more advantages. In some embodiments, nanoimprinting can be compatible with existing flow cell fabrication processes. This may reduce the manufacturing cost of the substrate, for example.

At 420, a core layer may be formed. In some embodiments, the core layer 310 is formed on the substrate 302. In some embodiments, core layer 310 is formed on grating layer 304. In some embodiments, grating layer 304 may function as a cladding layer for core layer 310. The core layer 310 may have a higher refractive index than the substrate 302. For example, the substrate 302 may have a refractive index of about 1.5, while the core layer may have a refractive index of about 2.2, or in a range of greater than about 1.5 to about 2.2, such as about 1.6 to about 2.1. In some embodiments, the refractive index may be about 1.65, about 1.85, about 2.05, and the like. Core layer 310 may cover substantially the entire facing surface of grating layer 304. Core layer 310 may have a higher refractive index than grating layer 304. In some embodiments, the core layer 310 comprises Ta₂O₅And/or SiN_x. For example, the core layer 310 may be formed by sputtering, chemical vapor deposition, atomic layer deposition, spin coating, and/or spray coating.

At 430, a nanopore layer map is performedAnd (6) patterning. In some embodiments, the nanopore layer 312 is formed at the core layer 310. The nanopore layer 312 may facilitate a patterned flow cell. The nanopore layer 312 may include one or more nanopores 314 defined between two or more walls 316. In some embodiments, the nanopore 314 may have a size such that one or more dimensions thereof are in a range of about one or more nanometers in magnitude. For example, the nanopore 314 may be configured to receive and hold a sample during a sample analysis process, such as in a cluster. The end (e.g., bottom) of the nanopore 314 may have a thickness that accommodates propagation of evanescent light. For example, the thickness may be from about 0 to about 500nm, such as between about 100nm to about 400 nm. In some embodiments, the thickness may be about 10nm, about 50nm, about 100nm, about 200nm, about 300nm, about 450nm, and the like. The nanopore layer 312 may be formed by a nanoimprint process or a lift-off process. In some embodiments, the nanoporous layer 312 may comprise one or more resins. The nanopore layer may cover substantially the entire facing surface of the core layer 310. For example, the resin may have a refractive index of about 1.5. In some embodiments, the nanopore layer 312 may have an average pitch between the nanopores 314 of at least about 10nm, about 0.1 μm, about 0.5 μm, about 1 μm, about 5 μm, about 10 μm, about 100 μm, or more, and/or may have an average pitch of at most about 100 μm, about 10 μm, about 5 μm, about 1 μm, about 0.5 μm, about 0.1 μm, or less. For example, the nanopore layer 312 may have a pitch between the nanopores 314 of about 600 to about 650nm, for example, the pitch may be about 610nm to about 640 nm. In some embodiments, the pitch may be about 605nm, about 615nm, about 635nm, about 655nm, and the like. The depth of each nanopore 314 may be at least about 0.1 μm, about 1 μm, about 10 μm, about 100 μm, or more. Alternatively or additionally, the depth may be up to about 1 × 10³μ m, about 100 μm, about 10 μm, about 1 μm, about 0.1 μm, or less.

Fig. 5 shows an example of fabricating a planar waveguide 500. The planar waveguide 500 may be used in one or more of the examples described herein. For example, the planar waveguide 500 may house one or more samples to facilitate illumination and excitation during sample analysis. Fig. 6 illustrates a method 600 of fabricating a planar waveguide in relation to the planar waveguide of fig. 5. Method 600 may be combined with one or more of the other methods described herein. More or fewer operations may be performed, and/or two or more operations may be performed in a different order, unless otherwise indicated.

At 610, nano-imprinting of the grating may be performed in the core layer. In some embodiments, substrate 502 is provided with one or more core layers 504. For example, core layer 504 may include a laser coupling grating. For example, the resin may be nanoimprinted. The core layers 504 are here shown longitudinally, such that the respective ridges 504' extend into the plane of the drawing. The core layer 504 may include a plurality of ridges 504' organized in respective groups 506A and 506B, for example. For example, groups 506A-506B may be separated by regions 508 of core layer 504 that also cover the surface of substrate 502. The ridges 504' may have a pitch of about 200nm to about 300nm, for example, between about 220 and about 280nm, to name a few. In some embodiments, the pitch may be about 205nm, about 215nm, about 235nm, about 265nm, about 285nm, and the like. For example, substrate 502 may have a refractive index of about 1.5, while core layer 504 may have a refractive index higher than about 1.5. For example, core layer 504 may be made of a high refractive index polymer material. Core layer 504 may cover substantially the entire facing surface of substrate 502.

At 620, nanopore layer patterning is performed. In some embodiments, a nanopore layer 510 is formed at core layer 504. The nanopore layer 510 may facilitate a patterned flow cell. In some embodiments, nanopore layer 510 and substrate 502 may function as cladding layers of core layer 504. The nanopore layer 510 may include one or more nanopores 512 defined between two or more walls 514. In some embodiments, the nanopore 512 may have a size such that one or more dimensions thereof are in a range of about one or more nanometers in magnitude. For example, the nanopore 512 may be configured to receive and hold a sample during an analysis process, such as in a cluster. The end (e.g., bottom) of the nanopore 512 may have a thickness that accommodates propagation of evanescent light. For example, the thickness may be from about 0 to about 500nm, such as between about 100nm to about 400 nm. In some embodiments, the thickness may be about 10nm, about 50nm, about 100nm, about 200nm, about 300nm, about 450nm, and the like. The nanopore layer 510 may be formed by a nanoimprint process or a lift-off process. In some embodiments, the nanopore layer 510 may comprise one or more resins. For example, the resin may have a refractive index of about 1.5. In some embodiments, the nanopore layer 510 may have an average pitch between the nanopores 512 of at least about 10nm, about 0.1 μm, about 0.5 μm, about 1 μm, about 5 μm, about 10 μm, about 100 μm, or more, and/or may have an average pitch of at most about 100 μm, about 10 μm, about 5 μm, about 1 μm, about 0.5 μm, about 0.1 μm, or less. For example, the nanopore layer may cover substantially the entire facing surface of core layer 504. The nanopore layer 510 may have a pitch between the nanopores 512 of about 600 to about 650nm, for example, the pitch may be about 610nm to about 640 nm. In some embodiments, the pitch may be about 605nm, about 615nm, about 635nm, about 655nm, and the like. The depth of each nanopore 512 may be at least about 0.1 μm, about 1 μm, about 10 μm, about 100 μm, or more. Alternatively or additionally, the depth may be up to about 1 × 10³μ m, about 100 μm, about 10 μm, about 1 μm, about 0.1 μm, or less.

Fig. 7 shows an example of fabricating a planar waveguide 700. The planar waveguide 700 may be used in one or more of the examples described herein. For example, the planar waveguide 700 may house one or more samples to facilitate illumination and excitation during sample analysis. Fig. 8 illustrates a method 800 of fabricating a planar waveguide in relation to the planar waveguide of fig. 7. Method 800 may be combined with one or more other methods described herein. More or fewer operations may be performed, and/or two or more operations may be performed in a different order, unless otherwise indicated.

At 810, nanoimprinting of the gratings may be performed. In some embodiments, substrate 702 is provided with one or more grating layers 704. For example, grating layer 704 may include a laser coupling grating. For example, the resin may be nanoimprinted. The grating layer 704 is here shown longitudinally, such that the respective ridges 704' extend into the plane of the illustration. The grating layer 704 may include a plurality of ridges 704', e.g., organized in respective groups 706A and 706B. For example, the groups 706A-706B may be separated by regions 708 of the grating layer 704 that also cover the surface of the substrate 702. This region 708 may form a thin residual layer on the surface of the substrate 702. The ridges 704' may have a pitch of about 200nm to about 300nm, such as between about 220nm to about 280nm, to name a few. In some embodiments, the pitch may be about 205nm, about 215nm, about 235nm, about 265nm, about 285nm, and the like. Applying grating layer 704 via a nanoimprinting process may provide one or more advantages. In some embodiments, nanoimprinting can be compatible with existing flow cell fabrication processes. This may reduce the manufacturing cost of the substrate, for example.

At 820, a core layer may be formed. In some embodiments, core layer 710 is formed at grating layer 704. In some embodiments, grating layer 704 may function as a cladding layer for core layer 710. The core layer 710 may have a higher refractive index than the substrate 702. For example, the substrate 702 may have a refractive index of about 1.5, while the core layer may have a refractive index of about 2.2, or in a range of greater than about 1.5 to about 2.2, such as from about 1.6 to about 2.1. In some embodiments, the refractive index may be about 1.65, about 1.85, about 2.05, and the like. Core layer 710 may cover substantially the entire facing surface of grating layer 704. Core layer 710 may have a higher refractive index than grating layer 704. In some embodiments, the core layer 710 includes Ta₂O₅And/or SiN_xOr a polymeric material. For example, the core layer 710 may be formed by sputtering, chemical vapor deposition, atomic layer deposition, spin coating, and/or spray coating.

At 830, additional layers may be formed. In some embodiments, layer 712 is formed at core layer 710. Layer 712 may be formed by sputtering, chemical vapor deposition, atomic layer deposition, spin coating, and/or spray coating, to name a few examples. Layer 712 has a lower index of refraction than core layer 710. In some embodiments, layer 712 is formed from a polymeric material having a refractive index similar to that of the water-based agent (e.g., about 1.35). For example, layer 712 may include, but is not limited to, a fluoropolymer that is transparent, electrically insulating, water and oil repellent, and/or chemically resistant. Layer 712 may have a thickness of about 100 to about 200nm, for example, about 120nm to about 180nm, to name just one example. In some embodiments, the thickness may be about 105nm, about 115nm, about 135nm, about 165nm, about 195nm, and the like.

At 840, nanopore layer patterning is performed. In some embodiments, a nanopore layer 714 is formed at layer 712. The nanopore layer 714 may facilitate a patterned flow cell. The nanopore layer 714 may include one or more nanopores 716 defined between two or more walls 718. In some embodiments, the nanopores 716 can have sizes such that one or more dimensions thereof are in a range of about one or more nanometer scales. For example, the nanopore 716 may be configured to receive and hold a sample during an analysis process, such as in a cluster. The end (e.g., bottom) of the nanopore 716 may have a thickness that accommodates propagation of evanescent light. For example, the thickness may be from about 0 to about 500nm, such as between about 100nm to about 400 nm. In some embodiments, the thickness may be about 10nm, about 50nm, about 100nm, about 200nm, about 300nm, about 450nm, and the like. The nanopore layer 714 may be formed by a nanoimprint process or a lift-off process. In some embodiments, the nanopore layer 714 may comprise one or more resins. The nanopore layer may cover substantially the entire facing surface of the layer 712. For example, the resin may have a refractive index of about 1.5. In some embodiments, the nanopore layer 714 may have an average pitch between the nanopores 716 of at least about 10nm, about 0.1 μm, about 0.5 μm, about 1 μm, about 5 μm, about 10 μm, about 100 μm, or more, and/or may have an average pitch of at most about 100 μm, about 10 μm, about 5 μm, about 1 μm, about 0.5 μm, about 0.1 μm, or less. For example, the nanopore layer 714 may have a pitch between nanopores 716 of about 600 to about 650nm, for example, the pitch may be about 610nm to about 640 nm. In some embodiments, the pitch may be about 605nm, about 615nm, about 635nm, about 655nm, and the like. The depth of each nanopore 716 can be at least about 0.1 μm, about 1 μm, about 10 μm, about 100 μm, or more. Alternatively or additionally, the depth may be up to about 1 × 10³μ m, about 100 μm, about 10 μm, about 1 μm, about 0.1 μm, or less.

Having layer 712 between core layer 710 and nanopore layer 714 may provide one or more advantages. In some embodiments, during sample analysis, the planar waveguide 700 is immersed in a water-based reagent, which may have a relatively low refractive index, such as about 1.35. The nanopore layer 714 may have a refractive index of about 1.5, which may be less than the ideal index to match with the water-based agent if the layer 712 is not present. For example, scattering or other photonic crystal effects may occur in the event of a refractive index mismatch. However, layer 712 may provide a relatively lower index of refraction than other materials. The evanescent light may have only a particular depth of penetration, such as from about 150 to about 200nm, for example, between about 165nm to about 185 nm; in some embodiments, the depth may be about 155nm, about 170nm, about 180nm, about 195nm, and the like. Thus, the thickness of layer 712 may be controlled such that evanescent light encounters only the water-based reagent in layer 712 and nanopore 716. That is, the evanescent light cannot (substantially) reach the nanopore layer 714. This may avoid or reduce unwanted effects, such as scattering.

The pattern of the nanopore layer 714 may be transferred into the layer 712. In some embodiments, layer 712 may be formed to cover substantially the entire surface of core layer 710. A nanopore layer 714 may be formed at layer 712. Thereafter, the nanopore layer 714 may be imprinted with the nanopore layer 716. The nanoimprint process may not penetrate or perforate layer 712; rather, layer 712 may remain substantially intact after nanoimprinting. Thereafter, an etch (e.g., a reactive ion etch) may be performed to remove portions of layer 712 at the ends (e.g., bottom) of nanopore 716. Thus, the pattern of nanopores 716 may be transferred to layer 712.

Fig. 9 shows an example of making a planar waveguide 900. Planar waveguide 900 may be used in one or more of the examples described herein. For example, the planar waveguide 900 may house one or more samples to facilitate illumination and excitation during sample analysis. Fig. 10 illustrates a method 1000 of fabricating a planar waveguide in relation to the planar waveguide of fig. 9. Method 1000 may be combined with one or more other methods described herein. More or fewer operations may be performed, and/or two or more operations may be performed in a different order, unless otherwise indicated.

At 1010, canA core layer is formed. In some embodiments, a core layer 902 is formed at a substrate 904. In some embodiments, substrate 904 may function as a cladding layer of core layer 902. The core layer 902 may have a higher refractive index than the substrate 904. For example, the substrate 904 may have a refractive index of about 1.5, while the core layer may have a refractive index higher than about 1.5. The core layer 902 may cover substantially the entire facing surface of the substrate 904. In some embodiments, the core layer 902 comprises Ta₂O₅And/or SiN_xOr a polymeric material. For example, the core layer 902 may be formed by sputtering, chemical vapor deposition, atomic layer deposition, spin coating, and/or spray coating. In some embodiments, the core layer 902 includes a high refractive index polymeric material, such as a resin. For example, a polymer may be spin coated to form the core layer 902.

At 1020, nanoimprinting of the gratings and nanopores is performed. In some embodiments, layer 906 is formed at core layer 902. Layer 906 may cover substantially the entire facing surface of core layer 902. Layer 906 can facilitate patterned flow cell and laser coupling through the grating. For example, the resin may be nanoimprinted. Here the layers 906 are shown longitudinally such that the respective ridges 908 extend into the plane of the drawing. Layer 906 may include, for example, a plurality of ridges 908 organized in groups 910A and 910B. For example, groups 910A-910B may be respective input and output gratings for a laser. The ridges 908 can have a pitch of about 200 to about 300nm, such as between about 220 to about 280nm, to name a few. In some embodiments, the pitch may be about 205nm, about 215nm, about 235nm, about 265nm, about 285nm, and the like.

Layer 906 may include one or more nanopores 912 defined between two or more walls 914. In some embodiments, the nanopore 912 may have a size such that one or more dimensions thereof are in a range of about one or more nanometers in magnitude. For example, the nanopore 912 can be configured to receive and hold a sample during an analysis process, such as in a cluster. The end (e.g., bottom) of the nanopore 912 may have a thickness that accommodates propagation of evanescent light. For example, the thickness may be from about 0 to about 500nm, such as between about 100nm to about 400 nm. In some casesIn embodiments, the thickness may be about 10nm, about 50nm, about 100nm, about 200nm, about 300nm, about 450nm, and the like. In some embodiments, the layer 906 can have an average pitch between the nanopores 912 of at least about 10nm, about 0.1 μm, about 0.5 μm, about 1 μm, about 5 μm, about 10 μm, about 100 μm or more, and/or can have an average pitch of at most about 100 μm, about 10 μm, about 5 μm, about 1 μm, about 0.5 μm, about 0.1 μm or less. For example, the nanopores 912 can have a pitch of about 600 to about 650nm, e.g., the pitch can be about 610nm to about 640 nm. In some embodiments, the pitch may be about 605nm, about 615nm, about 635nm, about 655nm, and the like. The depth of each nanopore 912 can be at least about 0.1 μm, about 1 μm, about 10 μm, about 100 μm, or greater. Alternatively or additionally, the depth may be up to about 1 × 10³μ m, about 100 μm, about 10 μm, about 1 μm, about 0.1 μm, or less. For example, layer 906 may be formed of a resin having a refractive index of about 1.5.

Forming the ridges 908 and nanopores 912 of the grating in layer 906 may involve nanoimprinting to more than one depth in layer 906. In some embodiments, a dual depth stamp may be used in a nanoimprinting process. For example, ridges 908 may be formed in layer 906 at a relatively greater depth than nanopores 912.

Fig. 11 shows an example of fabricating a planar waveguide 1100. Planar waveguide 1100 may be used in one or more of the examples described herein. For example, the planar waveguide 1100 may house one or more samples to facilitate illumination and excitation during sample analysis. Fig. 12 illustrates a method 1200 of fabricating a planar waveguide in relation to the planar waveguide of fig. 11. Method 1200 may be combined with one or more other methods described herein. More or fewer operations may be performed, and/or two or more operations may be performed in a different order, unless otherwise indicated.

At 1210, a core layer may be formed. In some embodiments, a core layer 1102 is formed at the substrate 1104. In some embodiments, substrate 1104 may function as a cladding layer for core layer 1102. The core layer 1102 may have a higher refractive index than the substrate 1104. For example, the substrate 1104 may have a refractive index of about 1.5, while the core layer may beHaving a refractive index higher than about 1.5. The core layer 1102 may cover substantially the entire facing surface of the substrate 1104. In some embodiments, the core layer 1102 comprises Ta₂O₅、SiN_xA polymeric material, or a combination thereof. For example, the core layer 1102 may be formed by sputtering, chemical vapor deposition, atomic layer deposition, spin coating, and/or spray coating. In some embodiments, the core layer 1102 comprises a high index polymeric material, such as a resin. For example, a polymer may be spin coated to form the core layer 1102.

At 1220, nanoimprinting of the grating may be performed. In some embodiments, the core layer 1102 is provided with one or more grating layers 1106. For example, grating layer 1106 may include a laser coupling grating. For example, the resin may be nanoimprinted. The grating layer 1106 is shown here longitudinally such that the respective ridges 1108 extend into the plane of the drawing. Grating layer 1106 can include a plurality of ridges 1108, organized in respective groups 1110A and 1110B, for example. For example, groups 1110A-1110B may be separated by regions 1112 of grating layer 1106 that also cover the surface of substrate 1104. The regions 1112 may form a thin residual layer on the surface of the substrate 1104. The ridges 1108 may have a pitch of about 200 to about 300nm, such as between about 220 to about 280nm, to name a few. In some embodiments, the pitch may be about 205nm, about 215nm, about 235nm, about 265nm, about 285nm, and the like. Applying the grating layer 1106 by a nanoimprinting process may provide one or more advantages. In some embodiments, nanoimprinting can be compatible with existing flow cell fabrication processes. This may reduce the manufacturing cost of the substrate, for example.

At 1230, nanoimprinting of the nanopore layer is performed. In some implementations, the nanopore layer 1114 is formed at the grating layer 1106. The nanopore layer 1114 may facilitate a patterned flow cell. The nanopore layer 1114 may include one or more nanopores 1116 defined between two or more walls 1118. In some embodiments, the nanopores 1116 may be of a size such that one or more dimensions thereof are in a range of about one or more nanometers in magnitude. For example, the nanopores 1116 may be configured to receive and hold a sample during an analytical process, such as in the form of a cluster. The end (e.g., bottom) of the nanopore 1116 may have a thickness that accommodates propagation of evanescent light. For example, the thickness may be from about 0 to about 500nm, such as between about 100nm to about 400 nm. In some embodiments, the thickness may be about 10nm, about 50nm, about 100nm, about 200nm, about 300nm, about 450nm, and the like. The nanopore layer 1114 may be formed by a nanoimprint process or a lift-off process. In some embodiments, the nanopore layer 1114 may comprise one or more resins. The nanopore layer may cover substantially the entire facing surface of the grating layer 1106. In some embodiments, the nanopore layer 1114 may have an average pitch between the nanopores 1116 of at least about 10nm, about 0.1 μm, about 0.5 μm, about 1 μm, about 5 μm, about 10 μm, about 100 μm, or more, and/or may have an average pitch of at most about 100 μm, about 10 μm, about 5 μm, about 1 μm, about 0.5 μm, about 0.1 μm, or less. For example, the nanopore layer 1114 may have a pitch between the nanopores 1116 of about 600 to about 650nm, for example, the pitch may be about 610nm to about 640nm, to name one example only. In some embodiments, the pitch may be about 605nm, about 615nm, about 635nm, about 655nm, and the like. The depth of each nanopore 1116 may be at least about 0.1 μm, about 1 μm, about 10 μm, about 100 μm, or greater. Alternatively or additionally, the depth may be up to about 1 × 10³μ m, about 100 μm, about 10 μm, about 1 μm, about 0.1 μm, or less.

Forming the grating layer 1106 at the core layer 1102 may provide one or more advantages. In some embodiments, the core layer 1102 may be preformed (e.g., by pre-sputtering) on the substrate 1104 prior to delivery to a facility (e.g., a fabrication facility of a flow cell manufacturer), and then the remainder of the planar waveguide 1100 may be formed (e.g., by nanoimprinting) without further transfer of the planar waveguide 1100 from the facility.

Grating layer 1106 may have a higher refractive index than one or more other materials. In some embodiments, grating layer 1106 has a refractive index that is higher than the refractive index of core layer 1102. In some embodiments, the core layer 1102 has a refractive index higher than the refractive index of the nanopore layer 1114. For example, the grating layer 1106 may have a higher refractive index than the nanopore layer 1114.

Fig. 13 shows an example of making a planar waveguide 1300. The planar waveguide 1300 may be used in one or more of the examples described herein. For example, the planar waveguide 1300 may house one or more samples to facilitate illumination and excitation during sample analysis. Fig. 14 illustrates a method 1400 of fabricating a planar waveguide in relation to the planar waveguide of fig. 13. Method 1400 may be combined with one or more other methods described herein. More or fewer operations may be performed, and/or two or more operations may be performed in a different order, unless otherwise indicated.

At 1410, a core layer may be formed. In some embodiments, a core layer 1302 is formed at a substrate 1304. In some embodiments, substrate 1304 may function as a cladding layer for core layer 1302. The core layer 1302 may have a higher index of refraction than the substrate 1304. For example, the substrate 1304 may have a refractive index of about 1.5, while the core layer may have a refractive index higher than about 1.5. The core layer 1302 may cover substantially the entire facing surface of the substrate 1304. In some embodiments, the core layer 1302 includes Ta₂O₅And/or SiN_xOr a polymeric material. For example, the core layer 1302 may be formed by sputtering, chemical vapor deposition, atomic layer deposition, spin coating, and/or spray coating. In some embodiments, the core layer 1302 includes a high index of refraction polymer material, such as a resin. For example, a polymer may be spin coated to form the core layer 1302.

At 1420, a polymer layer may be formed. In some embodiments, a polymer layer 1306 is formed at the core layer 1302. The polymer layer 1306 can be formed by sputtering, chemical vapor deposition, atomic layer deposition, spin coating, and/or spray coating, to name a few examples. The polymer layer 1306 has a lower index of refraction than the core layer 1302. The polymer layer 1306 may have a thickness of about 100 to about 200nm, for example, about 120nm to about 180nm, to name a few. In some embodiments, the thickness may be about 105nm, about 115nm, about 135nm, about 165nm, about 195nm, and the like.

At 1430, a resin layer may be formed. In some embodiments, a resin layer 1308 is formed at the polymer layer 1306. The resin layer 1308 may cover substantially the entire facing surface of the polymer layer 1306. The resin layer 1308 may be formed by spin coating and/or spray coating, to name a few. The resin layer 1308 has a lower refractive index than the polymer layer 1306.

At 1440, nanoimprinting of the gratings and nanopores is performed. The resin layer 1308 and the polymer layer 1306 may facilitate patterned flow cell and laser coupling through the grating. The polymer layer 1306 is shown here longitudinally such that the respective ridges 1310 extend into the plane of the drawing. The polymer layer 1306 can include a plurality of ridges 1310, for example, organized in respective groups 1312A and 1312B. For example, groups 1312A-1312B may be respective input and output gratings for the laser. The ridges 1310 may have a pitch of about 200 to about 300nm, for example, between about 220nm to about 280nm, to name a few. In some embodiments, the pitch may be about 205nm, about 215nm, about 235nm, about 265nm, about 285nm, and the like.

The resin layer 1308 may include one or more nanopores 1314 defined between two or more walls 1316. In some embodiments, the nanopores 1314 may have a size such that one or more dimensions thereof are in a range of about one or more nanometer-scale. For example, the nanopore 1314 may be configured to receive and hold a sample during an analysis process, such as in a cluster. The end (e.g., bottom) of the nanopore 1314 may have a thickness that accommodates propagation of evanescent light. For example, the thickness may be from about 0 to about 500nm, such as between about 100nm to about 400 nm. In some embodiments, the thickness may be about 10nm, about 50nm, about 100nm, about 200nm, about 300nm, about 450nm, and the like. In some embodiments, the resin layer 1308 can have an average pitch between nanopores 1314 of at least about 10nm, about 0.1 μm, about 0.5 μm, about 1 μm, about 5 μm, about 10 μm, about 100 μm or greater, and/or can have an average pitch of at most about 100 μm, about 10 μm, about 5 μm, about 1 μm, about 0.5 μm, about 0.1 μm or less. For example, the nanopores 1314 may have a pitch of about 600 to about 650nm, e.g., the pitch may be about 610nm to about 640 nm. In some embodiments, the pitch may be about 605nm, about 615nm, about 635nm, about 655nm, and the like. The depth of each nanopore 1314 may be at least about 0.1 μm,About 1 μm, about 10 μm, about 100 μm, or more. Alternatively or additionally, the depth may be up to about 1 × 10³μ m, about 100 μm, about 10 μm, about 1 μm, about 0.1 μm, or less.

Forming the ridges 1310 of the polymer layer 1306 and the nanopores 1314 of the resin layer 1308 may involve nanoimprinting to more than one depth. In some embodiments, a dual depth stamp (stamp) may be used in the nanoimprinting process. For example, the ridge 1310 may be formed at a relatively deeper depth than the nanopore 1314.

Fig. 15 shows an example of making a planar waveguide 1500. The planar waveguide 1500 may be used in one or more embodiments described herein. For example, the planar waveguide 1500 may house one or more samples to facilitate illumination and excitation during sample analysis. Fig. 16 illustrates a method 1600 of fabricating a planar waveguide in relation to the planar waveguide of fig. 15. Method 1600 may be combined with one or more other methods described herein. More or fewer operations may be performed, and/or two or more operations may be performed in a different order, unless otherwise indicated.

At 1610, a polymer layer may be formed. In some embodiments, a polymer layer 1502 is formed at the substrate 1504. The polymer layer 1502 may comprise a UV-curable or thermally-curable polymer material. The polymer layer 1502 may be formed by spin coating and/or spray coating, to name a few. The polymer layer 1502 may function as a core layer in the planar waveguide 1500. The polymer layer 1502 has a higher refractive index than the substrate 1504.

At 1620, a resin layer may be formed. In some embodiments, resin layer 1506 is formed at polymer layer 1502. The resin layer 1506 may cover substantially the entire facing surface of the polymer layer 1502. The resin layer 1506 may be formed by spin coating and/or spray coating, to name a few. The resin layer 1506 has a lower refractive index than the polymer layer 1502.

At 1630, nanoimprinting of the gratings and nanopores is performed. The resin layer 1506 and the polymer layer 1502 may facilitate patterned flow cell and laser coupling through the grating. The polymer layers 1502 are here shown longitudinally, such that the respective ridges 1508 extend into the plane of the drawing. The polymer layer 1502 may include a plurality of ridges 1508, for example, organized in respective groups 1510A and 1510B. For example, groups 1510A-1510B may be respective input and output gratings for a laser. The ridges 1508 can have a pitch of about 200 to about 300nm, for example, between about 220 to about 280nm, to name a few. In some embodiments, the pitch may be about 205nm, about 215nm, about 235nm, about 265nm, about 285nm, and the like.

The resin layer 1506 may include one or more nanopores 1512 defined between two or more walls 1514. In some embodiments, the nanopores 1512 may have a size such that one or more dimensions thereof are in a range of about one or more nanometer-scale. For example, the nanopores 1512 may be configured to receive and hold a sample during an analysis process, such as in a cluster. The end (e.g., bottom) of the nanopore 1512 may have a thickness that accommodates propagation of evanescent light. For example, the thickness may be from about 0 to about 500nm, e.g., between about 100nm to about 400 nm. In some embodiments, the thickness may be about 10nm, about 50nm, about 100nm, about 200nm, about 300nm, about 450nm, and the like. In some embodiments, the resin layer 1506 may have an average pitch between nanopores 1512 of at least about 10nm, about 0.1 μm, about 0.5 μm, about 1 μm, about 5 μm, about 10 μm, about 100 μm, or more, and/or may have an average pitch of at most about 100 μm, about 10 μm, about 5 μm, about 1 μm, about 0.5 μm, about 0.1 μm, or less. For example, the nanopores 1512 may have a pitch of about 600 to about 650nm, e.g., the pitch may be about 610nm to about 640 nm. In some embodiments, the pitch may be about 605nm, about 615nm, about 635nm, about 655nm, and the like. The depth of each nanopore 1512 may be at least about 0.1 μm, about 1 μm, about 10 μm, about 100 μm, or greater. Alternatively or additionally, the depth may be up to about 1 × 10³μ m, about 100 μm, about 10 μm, about 1 μm, about 0.1 μm, or less.

Forming the ridges 1508 of the polymer layer 1502 and the nanopores 1512 of the resin layer 1506 may involve nanoimprinting to more than one depth. In some embodiments, a dual depth stamp may be used in a nanoimprinting process. For example, the ridges 1508 may be formed at a relatively greater depth than the nanopores 1512.

The above examples illustrate a method of making a flow cell that includes forming a core layer (e.g., core layer 110, 310, 504, 710, 902, 1102, or 1302). The core layer is disposed between a substrate (e.g., substrate 102, 302, 502, 702, 904, 1104, 1304, or 1504) and a nanopore layer (e.g., nanopore layer 112, 312, 510, 714, or 1114). The nanopore layer has a nanopore to receive a sample. The core layer has a higher refractive index than the substrate and the nanopore layer.

The above embodiments illustrate methods of fabricating flow cells, wherein forming a grating comprises lithographic patterning (e.g., in fig. 1) or nanoimprinting (e.g., in fig. 3, 5, 7, 9, 11, 13, or 15).

The above example illustrates a method of manufacturing a flow cell in which a core layer is formed in the same process as the grating is formed (e.g., in fig. 5).

The above examples illustrate methods of fabricating flow cells in which the core layer is formed in a different process than the grating formation (e.g., in fig. 1, 3, 7, 9, 11, 13, or 15).

The above embodiments illustrate methods of fabricating flow cells, in which a grating is formed by photolithographic patterning onto a substrate, a core layer is formed on the grating, and a nanopore layer is formed on the core layer (e.g., in fig. 1).

The above embodiments illustrate methods of fabricating flow cells in which a grating is formed by nanoimprinting onto a substrate, a core layer is formed on the grating, and a nanopore layer is formed on the core layer (e.g., in fig. 3).

The above embodiments illustrate methods of fabricating a flow cell in which a grating is formed on a substrate, a core layer is formed on the grating, an additional layer is formed on the core layer, and a nanopore layer is formed on the additional layer (e.g., fig. 7). The additional layer and the nanopore layer may initially be free of nanopores, and the method may further include patterning the nanopore layer while the additional layer remains intact, and then transferring the pattern of the nanopore layer to the additional layer to expose the core layer in the nanopores. The pattern may be transferred by etching.

The above embodiments illustrate methods of fabricating a flow cell in which a core layer is formed on a substrate, and in which a grating and a nanopore layer are formed on the core layer (e.g., fig. 5, 9, 11, 13, or 15).

The above embodiments illustrate methods of fabricating a flow cell in which a core layer is formed on a substrate, a grating is formed on the core layer, and a nanopore layer is formed on the grating (e.g., fig. 5, 11, 13, or 15).

The above embodiments illustrate a method of manufacturing a flow cell, wherein a core layer is formed on a substrate, a first layer is formed on the core layer, a second layer is formed on the first layer, and wherein a grating and a nanopore are formed on the first layer and the second layer, respectively (e.g., fig. 11 or 13).

The above embodiment illustrates a method of manufacturing a flow cell, in which a core layer is formed on a substrate, a resin layer is formed on the core layer, and a grating and a nanopore are formed in the resin layer (e.g., fig. 9).

The above embodiments illustrate methods of fabricating flow cells in which the grating and nanopore layers are formed in the same process (e.g., fig. 9, 13, or 15).

The above example illustrates a method of manufacturing a flow cell in which a grating and a nanopore layer are formed in the same layer of the flow cell (e.g., fig. 9).

The above embodiments illustrate methods of fabricating flow cells in which the grating and nanopore layers are formed in separate layers of the flow cell (e.g., fig. 13 or 15).

Fig. 17 shows an example of a flow cell 1700. Flow cell 1700 can be used in one or more other embodiments described elsewhere herein. For example, one or more of the articles described above can be incorporated, and/or one or more of the techniques described above can be used to fabricate a flow cell. Flow cell 1700 can be produced according to one or more of the disclosed techniques. In flow cell 1700, a set of sealed chambers may be created due to negative spaces (negative spaces) in the gasket layer. The chamber may be sealed at the top and bottom by a base layer.

The flow cell 1700 here comprises a base layer (e.g., of borosilicate glass) 1710, a channel layer 1720 (e.g., of etched silicon or the like) overlying the base layer, and an overlying or top layer 1730. When the layers are assembled together, a closed channel is formed having an inlet/outlet at either end through the cover. Some flow cells may contain openings for channels at the bottom of the flow cell.

Fig. 18 is a diagram of a system 1800 including an instrument 1812, a cartridge 1814, and a flow cell 1816. System 1800 can be used for biological and/or chemical analysis. System 1800 may be used with or in an implementation of one or more other examples described elsewhere herein.

The cartridge 1814 may be used as a carrier for one or more samples, such as through a flow cell 1816. Cartridge 1814 can be configured to hold flow cell 1816 and transport flow cell 1816 into direct interaction with instrument 1812 or out of direct interaction with instrument 1812. For example, the instrument 1812 includes a receptacle 1818 (e.g., an opening in its housing) to receive and house the cartridge 1814 at least during collection of information from a sample. The cassette 1814 may be made of any suitable material. In some embodiments, the cassette 1814 comprises molded plastic or other durable material. For example, the cassette 1814 may form a frame for supporting or holding the flow cell 1816.

The examples herein refer to the sample being analyzed. Such samples may include genetic material. In some embodiments, the sample comprises one or more template strands of genetic material. For example, SBS can be performed on one or more template DNA strands using the techniques and/or systems described herein.

The flow cell 1816 may include one or more substrates configured to hold a sample to be analyzed by the instrument 1812. Any suitable material may be used for the substrate, including but not limited to glass, acrylic, and/or other plastic materials. The flow cell 1816 may allow a liquid or other fluid to selectively flow relative to the sample. In some embodiments, flow cell 1816 includes one or more flow structures that can hold a sample. In some embodiments, the flow cell 1816 can include at least one flow channel. For example, the flow channel may include one or more fluid ports to facilitate the flow of fluid.

The instrument 1812 may be operated to obtain any information or data related to at least one biological and/or chemical substance. The operations may be controlled by a central unit or by one or more distributed controllers. Here, an instrument controller 1820 is shown. For example, controller 1820 may be implemented using at least one processor, at least one storage medium (e.g., memory and/or drive) that retains instructions for operation of instrument 1812, and one or more other components, as described below. In some embodiments, the instrument 1812 may perform optical operations including, but not limited to, illumination and/or imaging of a sample. For example, the instrument 1812 may include one or more optical subsystems (e.g., an illumination subsystem and/or an imaging subsystem). In some embodiments, the instrument 1812 can perform a thermal treatment, including but not limited to a thermal treatment of a sample. For example, the instrument 1812 may include one or more thermal subsystems (e.g., heaters and/or coolers). In some embodiments, the instrument 1812 can perform fluid management including, but not limited to, adding and/or removing fluids in contact with the sample. For example, the instrument 1812 may include one or more fluidic subsystems (e.g., a pump and/or reservoir).

Fig. 19 is a diagram of an example lighting system 1900. Illumination system 1900 includes a light source assembly 1910, a mirror 1928, an objective lens 1934, a flow cell 1936, an emissive dichroic filter 1938, a first optical detection subsystem 1956, and a second optical detection subsystem 1958. Illumination system 1900 can image two color channels simultaneously. In some embodiments, another illumination system may be configured to enable imaging of more than two color channels (e.g., three color channels, four color channels, or more) simultaneously. It should be noted that there may be other optical configurations that can produce similar simultaneous imaging of multiple color channels.

Light source assembly 1910 generates excitation illumination incident on flow cell 1936. This excitation illumination will in turn produce emitted illumination or fluorescent illumination from one or more fluorescent dyes that will be collected using lenses 1942 and 1948. The light source module 1910 includes a first excitation illumination source 1912 and corresponding converging lens 1914, a second excitation illumination source 1916 and corresponding converging lens 1918, and a dichroic filter 1920.

First and second excitation illumination sources 1912, 1916 illustrate illumination systems (e.g., corresponding to respective color channels) that may simultaneously provide respective excitation illumination to the sample. In some embodiments, each of the first and second excitation illumination sources 1912, 1916 includes a Light Emitting Diode (LED). In some implementations, at least one of the first excitation illumination source 1912 and the second excitation illumination source 1916 includes a laser. Converging lenses 1914 and 1918 are each positioned a distance from a respective excitation illumination source 1912 and 1916 such that illumination emerging from each converging lens 1914/1918 is focused on a field aperture 1922. The dichroic filter 1920 reflects illumination from the first excitation illumination source 1912 and transmits illumination from the second excitation illumination source 1916.

In some embodiments, the mixed excitation illumination output from the dichroic filter 1920 may propagate directly to the objective lens 1934. In other embodiments, the mixed excitation illumination may be further modified and/or controlled by additional intermediate optical components prior to emission from objective lens 1934. The mixed excitation illumination may pass through a focal point in a viewing aperture 1922 to an optical filter 1924 and then through a color corrected collimating lens 1926. The collimated excitation illumination from lens 1926 is incident on mirror 1928, which it reflects on, and is incident on excitation/emission dichroic filter 1930. The excitation/emission dichroic filter 1930 reflects excitation illumination emitted from a light source assembly (assembly)1910 while allowing emission illumination to pass through the excitation/emission dichroic filter 1930 to be received by one or more optical subsystems 1956, 1958, as will be described further below. Optical subsystems 1956 and 1958 illustrate light collection systems that can collect multiple fluorescent lights simultaneously. The excitation illumination reflected from the excitation/emission dichroic filter 1930 is then incident on a mirror 1932, from which mirror 1932 is incident on an objective lens 1934 toward a flow cell 1936.

The objective lens 1934 focuses the collimated excitation light from the mirror 1932 onto the flow cell 1936. In some embodiments, objective lens 1934 is a microscope objective lens with a specified magnification of, for example, 1X, 2X, 4X, 5X, 6X, 8X, 10X, or higher. The objective lens 1934 focuses excitation illumination incident from the mirror 1932 on the flow cell 1936 at a cone of angle (cone of angle) or a numerical aperture (numerical aperture) determined by magnification. In some embodiments, objective lens 1934 can move in an axis perpendicular (normal) to the flow cell ("z-axis"). In some embodiments, the illumination system 1900 independently adjusts the z-position of the tube lens 1948 and the tube lens 1942.

Flow cell 1936 contains a sample to be analyzed, such as a nucleotide sequence or any other material. Flow cell 1936 may include one or more channels 1960 (schematically illustrated here in an enlarged cross-sectional view) configured to hold sample material and facilitate actions taken against the sample material, including but not limited to initiating a chemical reaction or adding or removing material. An object plane 1962 (shown schematically using dashed lines herein) of objective lens 1934 extends through flow cell 1936. For example, an object plane 1962 may be defined adjacent to the channel 1960.

The objective lens 1934 may define a field of view. The field of view may define an area on flow cell 1936 from which the image detector captures emitted light using objective lens 1934. One or more image detectors, such as detectors 1946 and 1954, may be used. The illumination system 1900 may include separate image detectors 1946 and 1954 for each wavelength (or range of wavelengths) of emitted light. At least one of the image detectors 1946 and 1954 may include a Charge Coupled Device (CCD), such as a time delay integration CCD camera, or a sensor fabricated based on Complementary Metal Oxide Semiconductor (CMOS) technology, such as a chemically sensitive field effect transistor (chemFET), an Ion Sensitive Field Effect Transistor (ISFET), and/or a Metal Oxide Semiconductor Field Effect Transistor (MOSFET).

In some implementations, the illumination system 1900 can include a Structured Illumination Microscope (SIM). SIM imaging is based on spatially structured illumination light and reconstruction to obtain a higher resolution image than would be produced using only the magnification of objective lens 1934. For example, the structure may be composed of or include a pattern or grating that interrupts the illumination excitation light. In some embodiments, the structures may include a pattern of stripes (patterns of fringes). Reflection or transmission diffraction occurs by irradiating a light beam onto the diffraction grating to produce fringes of light. Structured light may be projected onto the sample, illuminating the sample according to respective stripes, which may occur with a certain periodicity. To reconstruct an image using a SIM, two or more patterned images are used with the patterns of excitation illumination at different phase angles with respect to each other. For example, images of the sample may be acquired at different stages of the fringes of the structured light (sometimes referred to as various pattern stages of the image). This may allow multiple locations on the sample to be exposed to multiple illumination intensities. The resulting set of emitted light images can be combined to reconstruct a higher resolution image.

The sample material in flow cell 1936 is contacted with a fluorescent dye that is coupled to the corresponding nucleotide. The fluorescent dye emits fluorescent illumination upon being illuminated by corresponding excitation illumination incident on flow cell 1936 from objective lens 1934. The emitted illumination is identified by wavelength bands, each of which may be classified into a respective color channel. The fluorescent dyes are chemically bound to the respective nucleotides (e.g., containing the respective nucleobases). In this manner, dntps labeled with fluorescent dyes can be identified based on the wavelengths of light emitted within the corresponding wavelength bands when detected by the image detectors 1946, 1954.

Objective lens 1934 captures fluorescence emitted by fluorescent dye molecules in flow cell 1936. After capturing the emitted light, the objective lens 1934 collects and transmits collimated light (collimated light). This emitted light then propagates back along the path where the original excitation illumination came from the light source assembly (assembly) 1910. It should be noted that there is little or no expected interference along this path between the emitted and exciting illumination due to the lack of coherence between the emitted and exciting illumination. That is, the emitted light is the result of a separate light source, i.e., the result of the fluorescent dye contacting the sample material in flow cell 1936.

Upon reflection by mirror 1932, the emitted light is incident on an excitation/emission dichroic filter 1930. The filter 1930 passes the emitted light into a dichroic filter 1938.

In some implementations, the dichroic filter 1938 transmits illumination associated with the blue channel and reflects illumination associated with the green channel. In some embodiments, the dichroic filter 1938 is selected such that the dichroic filter 1938 reflects emitted illumination to optical subsystem 1956 within the defined green wavelength band and transmits emitted illumination to optical subsystem 1958 within the defined blue wavelength band, as discussed above. Optical subsystem 1956 includes tube lens 1942, filter 1944, and image detector 1946. Optical subsystem 1958 includes tube lens 1948, filter 1950, and image detector 1954.

In some implementations, the dichroic filter 1938 and the dichroic filter 1920 operate similarly to each other (e.g., both can reflect one color of light and transmit another color of light). In other embodiments, the dichroic filter 1938 and the dichroic filter 1920 operate differently from each other (e.g., the dichroic filter 1938 may transmit light having a color reflected by the dichroic filter 1920, or vice versa).

In some embodiments, the emitted illumination encounters the mirror 1952 before the image detector 1954. In the illustrated embodiment, the optical paths in optical subsystem 1958 are angled so that the illumination system 1900 as a whole can meet space or volume requirements. In some embodiments, such subsystems 1956 and 1958 both have angled optical paths. In some embodiments, none of the optical paths in either subsystem 1956 or 1958 are angled. As such, one or more of the plurality of optical subsystems may have at least one angled optical path.

Each tube lens 1942 and 1948 focuses the emitted illumination incident thereon onto a respective image detector 1946 and 1954. In some embodiments, each detector 1946 and 1954 includes a Charge Coupled Device (CCD) array. In some embodiments, each image detector 1946 and 1954 includes a Complementary Metal Oxide Semiconductor (CMOS) sensor.

The illumination system 1900 need not be as shown in fig. 19. For example, each mirror 1928, 1932, 1940 may be replaced by a prism or some other optical device that changes the direction of illumination. Each lens may be replaced by a diffraction grating, diffractive optics, a fresnel lens, or some other optical device that produces collimated or focused illumination from incident illumination.

The terms "substantially" and "about" are used throughout the specification to describe and explain small fluctuations, such as fluctuations due to variations in processing. For example, they may refer to less than or equal to ± 5%, such as less than or equal to ± 2%, such as less than or equal to ± 1%, such as less than or equal to ± 0.5%, such as less than or equal to ± 0.2%, such as less than or equal to ± 0.1%, such as less than or equal to ± 0.05%. Also, as used herein, an indefinite article such as "a" or "an" means "at least one".

It should be understood that all combinations of the foregoing concepts and additional concepts discussed in greater detail below (provided that such concepts do not contradict each other) are considered a part of the inventive subject matter disclosed herein. In particular, all combinations of claimed subject matter appearing at the end of this disclosure are considered part of the inventive subject matter disclosed herein.

A number of embodiments have been described. Nevertheless, it will be understood that various modifications may be made without departing from the spirit and scope of the specification.

In addition, the logic flows depicted in the figures do not require the particular order shown, or sequential order, to achieve desirable results. In addition, other processes may be provided, or eliminated, from the described flows, and other components may be added to, or removed from, the described systems. Accordingly, other implementations are within the scope of the following claims.

While certain features of the described embodiments have been illustrated as described herein, many modifications, substitutions, changes, and equivalents will now occur to those skilled in the art. It is, therefore, to be understood that the appended claims are intended to cover all such modifications and changes as fall within the scope of the implementations. It is to be understood that they have been presented by way of example only, and not limitation, and various changes in form and details may be made. Any portions of the devices and/or methods described herein may be combined in any combination, except mutually exclusive combinations. The embodiments described herein may include various combinations and/or subcombinations of the functions, components and/or features of the different embodiments described.