Patent Description:
<CIT> discloses methods for assessing one or more predetermined characteristics or properties of a microfluidic droplet within a microfluidic channel using an image sensor.

The term embodiment and like terms are intended to refer broadly to all of the subject matter of this disclosure and the claims below. Statements containing these terms should be understood not to limit the subject matter described herein or to limit the meaning or scope of the claims below. This summary is a high-level overview of various aspects of the disclosure and introduces some of the concepts that are further described in the Detailed Description section below. The subject matter should be understood by reference to appropriate portions of the entire specification of this disclosure, any or all drawings, and each claim.

Embodiments of the present disclosure include a system for detecting at least one particle or at least one droplet in a channel, as defined in claim <NUM>.

In certain embodiments, the sensor is a linear charge-coupled sensor, a linear complementary metal-oxide-semiconductor sensor, any suitable optical sensor, or any combination thereof. In some non-limiting examples, the channel is positioned in or on a substrate. In some aspects, the source of illumination comprises an optical system. In certain embodiments, the system further includes a plurality of particles or droplets and/or a plurality of channels and/or a plurality of linear sensors. Thus, in some cases, the sensor and a detector detect the movement and or position of the at least one particle or at least one droplet over a predetermined time and/or distance. As described herein, the optical system comprises a lens selected from the group consisting of a cylindrical lens, or a telecentric lens, or a spherical lens, or an aspheric lens, or any suitable lens, or any combination thereof. The sensor detects spectral information for identifying the particle or the droplet. Additionally, the system includes a sensor positioned within the region of illumination, but perpendicular to the linear axis. In some cases, the substrate and the medium have a similar refractive index. In some aspects, signals from the sensor are used to modulate a device that is separate from the system (e.g., a component for sorting the plurality of particles in a channel based on the size, position and/or other characteristics). In some embodiments, the device that is separate from the system is a component for sorting a plurality of particles in a channel based on size, position, and/or other characteristics.

Also described herein is a method of detecting at least one particle according to claim <NUM>. In some cases, the detecting is performed in real time. In some non-limiting examples, the detecting can include characterizing the at least one particle or the at least one droplet (e.g., identifying the at least one particle or the at least one droplet, recording a velocity of the at least one particle or the at least one droplet, recording a size of the at least one particle or the at least one droplet, recording an absorption spectrum of the at least one particle or the at least one droplet, recording a fluorescent spectrum of the at least one particle or the at least one droplet; recording light scattering of the at least one particle or the at least one droplet, recording a refractive index of the at least one particle or the at least one droplet, any suitable characterization technique, or any combination thereof). In some aspects, the method further includes sorting a plurality of particles or droplets in real time according to particle or droplet identification, particle or droplet size, or any suitable attribute. The sorting can be performed by actuating at least an electrode, valve or other component that may be used to change a direction of particle flow through the channel.

In the following detailed description, reference is made to the accompanying figures, which form a part hereof. In the figures, similar symbols typically identify similar components, unless context dictates otherwise. The illustrative embodiments described in the detailed description, figures, and claims are not meant to be limiting. Other embodiments may be utilized, and other changes may be made, without departing from the scope of the subject matter presented herein. It will be readily understood that the aspects of the present disclosure, as generally described herein, and illustrated in the figures, can be arranged, substituted, combined, separated, and designed in a wide variety of different configurations, all of which are explicitly contemplated herein.

Certain aspects and features of the present disclosure relate to tracking particles or droplets in a channel and in some embodiments, specifically to fluorescence activated cell sorting (FACS). Disclosed herein are systems and methods capable of identifying, tracking, and sorting particles or droplets flowing in a channel. The channel can be a microfluidic channel disposed onto or within a substrate. The channel can further include a medium in which the particles or droplets can be carried (i.e., such that the particles or droplets flow through the channel in the medium). The channel and the medium can have a similar refractive index such that they appear translucent or transparent to each other, and when illuminated by electromagnetic radiation are simultaneously translucent or transparent to the radiation. In certain cases, at least two of the substrate, the channel, and/or the medium are translucent or transparent. In some cases, at least two of the substrate, the channel, and/or the medium can have a similar refractive index such that they are, or appear translucent or transparent to each other. The particles or droplets can have a refractive index substantially different from that of the substrate, the channel, and/or the medium, such that the particles or droplets interfere with the electromagnetic radiation. A sensor can be disposed adjacent to the channel to record the electromagnetic radiation. The sensor can be attached to a system for identifying, tracking, and sorting the particles or droplets. A plurality of channels can be combined to form a network. It is noted that description embodiments described for compositions may also be incorporated in methods and/or systems and vice versa.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of ordinary skill in the art to which this disclosure belongs. All patents, applications, published applications and other publications referred to herein are incorporated by reference in their entireties. If a definition set forth in this section is contrary to or otherwise inconsistent with a definition set forth in the patents, applications, published applications and other publications that are herein incorporated by reference, the definition set forth in this section prevails over the definition that is incorporated herein by reference.

When introducing elements of the present disclosure or the embodiment(s) thereof, the articles "a", "an", "the" and "said" are intended to mean that there are one or more of the elements. It is understood that aspects and embodiments of the disclosure described herein include "consisting" and/or "consisting essentially of" aspects and embodiments.

The term "and/or" when used in a list of two or more items, means that any one of the listed items can be employed by itself or in combination with any one or more of the listed items. For example, the expression "A and/or B" is intended to mean either or both of A and B, i.e. A alone, B alone or A and B in combination. The expression "A, B and/or C" is intended to mean A alone, B alone, C alone, A and B in combination, A and C in combination, B and C in combination or A, B, and C in combination.

Various aspects of this disclosure are presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the disclosure. Accordingly, the description of a range should be considered to have specifically disclosed all the possible sub-ranges as well as individual numerical values within that range. For example, description of a range such as from <NUM> to <NUM> should be considered to have specifically disclosed sub-ranges such as from <NUM> to <NUM>, from <NUM> to <NUM>, from <NUM> to <NUM>, from <NUM> to <NUM>, from <NUM> to <NUM>, from <NUM> to <NUM> etc., as well as individual numbers within that range, for example, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, and <NUM>.

As used herein a translucent material is a material wherein a fraction of impinging light diffracted above <NUM>° of an incident angle (referred to as "haze") exceeds <NUM>%. Such techniques are explained fully in the literature (e.g.,<NPL>)).

As used herein a transparent material is a material wherein greater than <NUM>% of ultraviolet light, visible light and near-infrared radiation (e.g., electromagnetic radiation having a wavelength from about <NUM> to about <NUM>) can transmit through the transparent material. Such techniques are explained fully in the literature, such as, ASTM Standard D1003.

As used herein, two materials and/or media (or a material and a medium) having equal or similar refractive indices refract light similarly with respect to each other, thus allowing the light to pass through the two materials/media uninhibited, providing a substantially transparent pair. For example, glass and air have similar refractive indices, thus light passes through glass and air uninhibited. Conversely, when two media/materials have significantly different refractive indices, light transmitting from a first media/material to a second media/material is refracted in directions significantly differently as to inhibit light transmission through the system.

As used herein, a droplet is a self-contained volume of a first medium dispersed, suspended, or otherwise included in a second medium. Generally, the first medium of the droplet is a liquid and the second medium is either a liquid or a gas, though it need not be. The droplet may be homogeneous or heterogeneous (i.e., include other materials within the droplet such as solid particles, gas bubbles, miscible liquids, immiscible liquids, or the like). For example, in various non-limiting embodiments the droplet may be an oil droplet in an aqueous medium, a water droplet in an oil medium, a soap bubble in air, or the like).

Other objects, advantages and features of the present disclosure will become apparent from the following specification taken in conjunction with the accompanying drawings.

In some non-limiting examples, disclosed herein is a system for detecting at least one particle or at least one droplet in a channel, including a channel having at least one particle or at least one droplet dispersed in a medium, such that the at least one particle or the at least one droplet (i.e., the particle or the droplet) is moving from a first end of the channel to a second end of the channel, a source of electromagnetic radiation that illuminates at least a portion of the channel, a sensor to detect the particle or the droplet, wherein the sensor is positioned along the linear axis of the illuminated portion of the channel such that the sensor is substantially parallel to the direction of movement of the particle through the channel, and an optical system that focuses and aligns the illuminated portion of the channel to the sensor.

In some non-limiting examples, the channel is positioned in or on a substrate. In some cases, the substrate can be a silicon wafer substrate, a polymer substrate (e.g., a poly(dimethylsiloxane) (PDMS) substrate), a gallium arsenide wafer substrate, a glass substrate, a ceramic substrate (e.g., a yttrium stabilized zirconia (YSZ) substrate), or any suitable substrate. In some non-limiting examples, the channel can be positioned within the substrate. For example, the channel can be created by creating at least a first half of a channel in a first substrate and creating a second half of a channel in a second substrate, and aligning and joining the first substrate to the second substrate such that the first half of the channel and the second half of the channel form a complete channel encased within the first substrate and the second substrate. In some other examples, the channel can be at least partially exposed to the environment outside of the substrate. For example, a portion of the substrate can be removed in a predetermined pattern creating an exposed channel positioned at least partially within the substrate, such that any medium and/or particles or droplets (e.g., cells, liposomes, or the like) are exposed to the environment outside of the substrate when flowing through the channel. The portion of the substrate can be removed by any one of reactive ion etching (i.e., dry etching), wet chemical etching (i.e., wet etching), electron beam (E-beam) lithography, photolithography (e.g., photolithography employing dry etching and/or wet etching), laser etching, any suitable material removal technique, or any combination thereof. In some further examples, the channel can be fabricated on the substrate. For example, the channel can be created by depositing a material onto the substrate, removing at least a portion of the material in a predetermined pattern (e.g., a channel or a network of channels) to create a channel within the material deposited onto the substrate. The portion of the material deposited onto the substrate can be removed by any one of reactive ion etching (i.e., dry etching), wet chemical etching (i.e., wet etching), electron beam (E-beam) lithography, photolithography (e.g., photolithography employing dry etching and/or wet etching), laser etching, soft lithography, two-photon lithography, forming the channel around a sacrificial template, any suitable material removal technique, or any combination thereof. In certain embodiments, the channel can be created by three dimensional (3D) printing.

Also, as noted herein, the channel may be configured in a variety of shapes. The channel can have a square shape, a rectangular shape, a triangular shape, a circular shape, an elliptical shape, or any suitable shape. In further embodiments, for example, the channel can have any two dimensional (2D) cross section and/or three dimensional (3D) shape. Thus, the channel cross section can be a rectangle, square, circle, ellipse, polygon, parallelogram, triangle, any combination thereof, or any suitable shape.

The channels disclosed herein may be configured in a variety of sizes. Round channels can have a diameter of from about <NUM> to about <NUM> (e.g., about <NUM> to about <NUM>, about <NUM> to about <NUM>, about <NUM> to about <NUM> microns (µm), about <NUM> to about <NUM>, or about <NUM> to about <NUM>). For example, round channels can have a diameter of <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, or anywhere in between. Rectangular channels can have a width of from about <NUM> to about <NUM> (e.g., about <NUM> to about <NUM>, about <NUM> to about <NUM>, about <NUM> to about <NUM> microns, about <NUM> to about <NUM>, or about <NUM> to about <NUM>). For example, rectangular channels can have a width of <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, or anywhere in between. Rectangular channels can have a depth of from about <NUM> to about <NUM> (e.g., about <NUM> to about <NUM>, about <NUM> to about <NUM>, about <NUM> to about <NUM> microns, about <NUM> to about <NUM>, or about <NUM> to about <NUM>). For example, rectangular channels can have a depth of <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, <NUM>, or anywhere in between.

The medium can be at least partially contained within a channel, wherein the channel can be fashioned as a plurality of channels, a network of channels, a reservoir, an inlet, an outlet, a source, a drain, or any combination thereof. The channel can be contained within the substrate. The channel or plurality of channels can be disposed on a surface of the substrate such that the medium can be exposed to any environment in which the substrate can be placed.

The channel created in the substrate can have at least a first end and a second end. In some examples, the substrate can have a first port disposed on a surface of the substrate, wherein the first port can be an inlet. The inlet can expose at least part of a channel disposed within the substrate to the exterior of the substrate, enabling filling the channel with a medium (e.g., oil, water, any suitable medium, or any combination thereof) and/or the particles or droplets. The inlet can optionally be sealed after filling the channel with the medium and/or the particles or droplets. Sealing the inlet can include gluing, pinching, clamping, recasting (e.g., melting the inlet material and allowing the material to solidify in a sealed state), or plugging. Optionally, the substrate can have a second port disposed on a surface of the substrate, wherein the second port can be an outlet. The outlet can expose at least part of a channel disposed within the substrate to the exterior of the substrate, enabling draining the channel of the medium and/or the particles or droplets. The outlet can optionally be opened after filling the channel with the medium and/or the particles or droplets to drain the channel. Opening the outlet can include dissolving glue, unpinching, unclamping, melting, piercing, or unplugging.

In some cases, the substrate and the medium have a similar refractive index. In certain embodiments, the substrate and the channel have a similar refractive index. For example, the substrate can have a refractive index that is similar to a gas filling the channel (e.g., air, an inert gas, a processing gas, or any combination thereof). In some examples, the medium and the channel have a similar refractive index (e.g., when the channel is partially filled with the medium, the medium and the gas filling a remainder of the channel can have a similar refractive index). In some cases all, or at least two of the substrate, the medium, and/or the channel all have a similar refractive index and/or are translucent and/or transparent. For example, when the substrate, the medium, and the channel have a similar refractive index, the system can be substantially translucent and/or substantially transparent. In some cases, the refractive indices of the substrate, the channel, and the medium can be within about up to <NUM> % of each other. For example, the refractive indices of the substrate and the medium can be within about <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> %, <NUM> % (e.g., matching refractive indices), or anywhere in between.

In some embodiments, a plurality of channels can be formed in a substrate to create a network of channels. The channels can intersect in 2D and/or in 3D. For example, the channels can intersect at any suitable angle (e.g., about <NUM>° to about <NUM>°, or anywhere in between) in a single plane. In some further examples, the channels can intersect across a plurality of planes (i.e., the channels can be formed into interplanar interconnects). In a still further example, the channels can intersect within a single plane and across a plurality of planes. In still further examples, the channel can have a 3D shape. For example, the channel can be a coil, a toroid, an arc, or a helix. For example, the sensor can be placed along and/or within a linear axis of a coil or a toroid. In certain embodiments, the channel is a microfluidic channel.

In certain embodiments, the sensor is a linear charge-coupled sensor, or a linear complementary metal-oxide-semiconductor sensor, though it need not be. In some aspects, the sensor can be placed adjacent to the microfluidic channel. For example, the sensor can be suspended above the microfluidic channel (e.g., when the microfluidic channel is placed partially within the substrate or onto the substrate), the sensor can be placed beneath the substrate (e.g., the substrate can be placed onto the sensor. The sensor detects spectral information about the particle or droplet. For example, the sensor can be coupled to a fluorescence spectrometer, an absorption spectrometer, an optical spectrometer, any suitable spectrophotometer, or any combination thereof (e.g., when employed in a fluorescence activated cell sorting (FACS) system). In some aspects, the illumination source can provide excitation energy. For example, when the particle or droplet is a fluorescent particle or droplet, the illumination source can provide EM radiation sufficient to excite the fluorescent particle or droplet such that the fluorescent particle or droplet fluoresces. Thus, the sensor can characterize the fluorescence of the particle or droplet. Spectral information of the particle or droplet is employed to identify the particle or droplet. In certain other embodiments, the sensor can be configured to detect partial spectral information about the particle or droplet. For example, a filter can be placed in front of the sensor to allow only a desired wavelength of electromagnetic radiation to the sensor. In further embodiments, the excitation energy can be tuned to excite at least a portion of the particle or droplet. For example, the particle or droplet can contain several different species (e.g., small molecule pendant groups attached to a main backbone, several different dye species, several different ligands, or the like) wherein a first subset of particles or droplets can be sorted from a second subset of particles or droplets.

According to the invention, a line sensor is employed to detect the particle or the droplet crossing a point or a channel cross-section by positioning the line sensor so that the array is perpendicular to the linear axis of the channel. Additionally, Z the perpendicular sensor can be used to determine a position of the particle or the droplet within the channel.

In some aspects, the source of illumination comprises an optical system. The optical system can comprise any suitable arrangement of optical components such that a predetermined amount and kind of electromagnetic radiation is employed. For example, the optical system can include an electromagnetic (EM) radiation source (e.g., a visible light source, an ultraviolet light source, an infrared light (IR) source, any suitable EM radiation source, or any combination thereof), a lens (e.g., a spherical lens, an aspherical lens, an axisymmetric lens, a non-axisymmetric lens, a cylindrical lens, a telecentric lens, an anamorphic lens, a collimating lens, any suitable lens, or any combination thereof), a mirror (e.g., a full silvered mirror, a half silvered mirror, any suitable mirror, or any combination thereof), a prism, a waveguide, optical fiber, any suitable optical component, or any combination thereof. In certain embodiments, a variety of different lenses can be employed in the optical system to improve performance. For example, a cylindrical lens can expand or contract an image of the channel along an axis (e.g., the linear axis of the channel), a very fast cylindrical lens can be used to account for any residual tilt in the axis that might be greater than the height of a pixel, and/or a telecentric lens (e.g., a lens that preserves distances at different focal points) can be used to prevent errors in length measurements.

In some cases, the particle or the droplet is a cell, a liposome, a bead, an emulsion, a colloid, a cell fragment, a cell cluster, a vesicle, an extracellular vesicle, an exosome, a virus, a quantum dot, a nanoparticle, a surface-enhanced Raman spectroscopy (SERS) particle, any suitable material dispersed in a medium, or any combination thereof. In certain embodiments, the particle or the droplet can have a diameter of from about <NUM> to about <NUM> (e.g., about <NUM> to about <NUM>). For example, the particle or the droplet can have a diameter of about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, about <NUM>, or anywhere in between.

In some aspects, the particle or the droplet can move through the channel at a rate of from about <NUM> meters per second (m/s) to about <NUM>/s. For example, the particle or droplet can move at a rate of about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, or anywhere in between.

In certain embodiments, a plurality of particles or droplets can be detected by the system. In some cases, the plurality of particles or droplets can be produced at a rate of from about <NUM> per second to about <NUM>,<NUM> per second (/s). For example, the particles or droplets can be produced at a rate of about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>/s, about <NUM>,<NUM>/s, or anywhere in between.

In some aspects, signals from the sensor are used to modulate a device that is separate from the system (e.g., a component for sorting the plurality of particles in a channel based on the size, position and/or other characteristics). In some non-limiting examples, the device that is separate from the system is an electrode, a valve (e.g., an electronic valve, a pneumatic valve, a hydraulic valve, any suitable valve, or any combination thereof), a switch, a magnet, any suitable particle directing device, or any combination thereof. In some cases, the device that is separate from the system is an analytical tool (e.g., a spectrophotometer), a display (e.g., for reporting data, or for providing analysis by a lab-on-a-chip device), a pressure modulator (e.g., for maintaining or controlling pressure in a closed-loop system), any suitable device, or any combination thereof.

In certain embodiments, the system further includes a plurality of particles or droplets, and/or a plurality of channels, and/or a plurality of sensors. In some embodiments, a plurality of substrates can be combined to form a network system. Any suitable system design can be accomplished by combining the plurality of substrates.

Described herein is a method of detecting at least one particle or at least one droplet in a channel comprising allowing the at least one particle or the at least one droplet (i.e., the particle or the droplet)to flow through the channel (the channel having a substantially linear axis perpendicular to the circumference of the channel), aligning a linear sensor along the substantially linear axis of the channel, illuminating the channel with electromagnetic radiation along at least the linear axis (wherein at least the particle or the droplet changes optical properties of the electromagnetic radiation transmitted to the sensor), and detecting, with the linear sensor, the electromagnetic radiation as transmitted along the linear axis of the channel.

In certain embodiments, allowing the particle or the droplet to flow through the channel can be a laboratory method (e.g., analyzing blood or conducting research), a lab-on-a-chip method (e.g., analyzing fluids in an emergency), any suitable method wherein a plurality of particles or droplets are suspended in a medium and require analysis, or any combination thereof.

In some cases, the detecting is performed in real time (e.g., detecting the particle or the droplet can be instantaneous such that a desired action can be taken in response to the identification of the particle or the droplet). Aligning a single line sensor (i.e., a sensor that has a line of pixels that takes a line scan along the linear axis of the channel and uses, in some examples, contrast changes to determine the location of the particle or the droplet along the channel) can provide a constant stream of information about the channel and the particle or the droplet in the channel. In certain embodiments, the sensor is aligned with the linear axis of the channel at a fixed location above the channel, thus allowing the particle or the droplet to flow past the sensor. Illuminating the channel from a side opposite the sensor can allow the sensor to detect light transmitting through the channel and the medium in which the particle or the droplet is suspended. In some aspects, the sensor is aligned with the linear axis of the channel such that, in the line of pixels, a first pixel in the line of pixels is positioned over a first area being observed by the sensor, and each subsequent pixel is positioned over a subsequent area being observed. Thus, allowing the particle or the droplet to flow past the sensor further allows the particle or the droplet to interfere with the light transmitting through the channel and the medium, further allowing detection of the particle or the droplet.

In some non-limiting examples, all, or at least two of the substrate, the channel, and/or the medium have similar refractive indices such that neither the substrate, the channel, nor the medium interfere with the light transmitting through with respect to each other. In some non-limiting examples, all, or at least two of, the substrate, the channel, and the medium are translucent or transparent. Accordingly, the particle or the droplet can have a different refractive index, or, in some examples, be opaque, and interfere with the light transmitting through the system, allowing the sensor to detect the particle or the droplet.

In certain embodiments, illuminating the channel is performed with bright-field illumination (e.g., white light) and the sensor is an optical sensor. Due to the difference in refractive index between the substrate, the channel, and the medium (e.g., oil) (i.e., the system), and a dispersed droplet (e.g., water), the boundary of the droplet stands out as circle of dark pixels as shown in <FIG>.

<FIG> shows a system <NUM> placed on a substrate bearing a microfluidic channel <NUM>. The microfluidic channel <NUM> was filled with oil. Water droplets <NUM> were dispersed in the oil and forced to flow through the microfluidic channel <NUM>. Alignment marks <NUM> were employed to ensure proper positioning of the sensor. Scale bars <NUM> were employed to verify sizing the particles. For example, the largest scale bar <NUM> is <NUM> wide.

<FIG> shows a system for performing a method as described herein. In some non-limiting examples, a linear sensor can be aligned with a linear axis <NUM> of the channel <NUM>. An illumination source can be positioned such that light can be transmitted through the substrate and channel <NUM>. In some cases, the droplet <NUM> can have a different refractive index than the substrate, and/or the medium, and/or the channel. In some aspects, the droplet <NUM> can be opaque. In some further cases, the droplet <NUM> can have a similar refractive index to the substrate, the medium and/or the channel, however, the edges of the droplet <NUM> can still effectively interfere with the light, allowing for detection of the droplet <NUM> flowing through the channel <NUM>.

In certain embodiments, when the sensor is a single line sensor (e.g., a sensor that employs an array of pixels arranged in a line), a line scan can be taken along the linear axis <NUM> of the channel <NUM>. Thus, contrast changes can be recorded to determine the location of the droplet <NUM>, a particle, a liposome, or any suitable material dispersed in a medium flowing within the channel <NUM>.

<FIG> is a schematic depicting the progression of the droplet <NUM> along the linear axis <NUM> of the channel <NUM> and the sensor. In certain embodiments, row <NUM> (top row in <FIG>) depicts a first time or line scan, row <NUM> (center row in <FIG>) depicts a second time or line scan, and row <NUM> (bottom row in <FIG>) depicts a third time or line scan. Darkened pixel depictions illustrate the boundary of the droplet <NUM> interfering with light transmitting through the system. In the example of <FIG>, row <NUM> illustrates the boundary of the droplet <NUM> blocking light from the first and third pixels and light is allowed to transmit to the second pixel (e.g., the center of the droplet <NUM>), as well as the fourth through eighth pixels (e.g., no droplet <NUM> is present). In the second time (see row <NUM>), as the droplet <NUM> progresses, light transmitting through the system is blocked from the second and fourth pixels of the sensor by the boundary of the droplet <NUM>. Accordingly, in the third time (see row <NUM>), light transmitting through the system is blocked from the third and fifth pixels as in the example of <FIG>. Thus, size, velocity, and/or acceleration of the droplet <NUM> can be provided from information detected by the sensor.

In certain embodiments, velocity can be provided by evaluating multiple contiguous line scans, for example, at least two line scans. Accordingly, dividing a distance traveled by the droplet <NUM> (i.e., a position recorded by a first line scan subtracted from a position recorded by a second line scan) by the time between the two scans can provide a velocity. In a further example, employing at least a third line scan can provide an acceleration of the droplet <NUM> when needed or desired.

In some cases, data recorded from the sensor can be presented as a kymograph (see <FIG>). In the example of <FIG>, the y-axis represents time and the x-axis represents position of the droplet <NUM>. The kymograph represents the progression of the droplet <NUM> as recorded by the sensor in the example of <FIG>. The progression of the droplet <NUM> is depicted as a single line, thus the kymograph in the example of <FIG> depicts the progression of <NUM> droplets. Velocity of the droplet <NUM> is represented by the slope of the line (e.g., a steeper slope indicates a slower droplet). In the example of <FIG>, velocity of the droplet <NUM> was varied as it progressed through the channel <NUM>.

In some aspects, a plurality of particles or droplets can be detected simultaneously. In certain embodiments, the sensor can perform the line scans constantly, providing constant real time analysis of the particles or droplets flowing in the channel. In some examples, the sensor can perform greater than <NUM> line scans per second (e.g., greater than <NUM> line scans per second, greater than <NUM> line scans per second, greater than <NUM> line scans per second, greater than <NUM> line scans per second, greater than <NUM> line scans per second, greater than <NUM> line scans per second, greater than <NUM> line scans per second, greater than <NUM> line scans per second, greater than <NUM> line scans per second, greater than <NUM> line scans per second, or anywhere in between). Thus, the line scan data can be used to identify and/or track the particles or droplets and a signal can be sent to device external to the system to perform a desired action (e.g., characterizing, sorting, or any suitable action).

In certain embodiments, the desired action can be characterizing the particle or droplet, sorting the particle or droplet, modifying the particle or droplet, treating the particle or droplet, any suitable action, or any combination thereof. In some examples, sorting the particle or droplet can be performed by activating an external device, including an electrode, a valve, any flow control mechanism, or any combination thereof. In certain embodiments, modifying the particle or droplet can include releasing a chemical into the channel, emitting radiation to the particle or droplet, or the like.

In some non-limiting examples, characterizing the particle can include identifying the particle or droplet, recording a velocity of the particle or droplet, recording an acceleration of the particle or droplet, recording a size of the particle or droplet, or any combination thereof). Characterizing the particle can be performed employing any suitable characterization systems or methods able to use information captured by the sensor. For example, illuminating the channel with electromagnetic radiation can include transmitting a wavelength of light that can excite an aspect of the droplet or particle and stimulate fluorescence. The fluorescence can be recorded by a spectrophotometer coupled to the sensor and the particle or droplet can be identified by its fluorescent spectrum.

In some aspects, the method further includes sorting a plurality of particles or droplets in real time according to particle or droplet identification, particle or droplet size, or any suitable attribute. In certain embodiments, the plurality of particles or droplets can include various different particles or droplets requiring sorting for analytical, research, or any suitable purpose.

The sorting can be performed by actuating at least an electrode, valve or other component that may be used to modulate flow through the channel. In some aspects, the sorting can be performed based on information detected by the sensor. In certain embodiments, actuating an electrode can produce an electric field within and/or across the channel capable of redirecting the particle or droplet flowing in the channel into a secondary channel or reservoir (e.g., as in a network system including a plurality of channels or the like).

In certain embodiments, the system described herein can react to the velocity of the particles or droplets. For example, the system can be employed to actuate a pressure control mechanism to control the velocity of the particles or droplets and/or the pressure of the system (e.g., in a closed-loop system having a predetermined pressure constraint).

In some embodiments, a plurality of channels can be formed in a substrate to create a network of channels. The channels can intersect in 2D and/or in 3D. For example, the channels can intersect at any suitable angle (e.g., about <NUM>° to about <NUM>°, or anywhere in between) in a single plane. In some further examples, the channels can intersect across a plurality of planes (i.e., the channels can be formed into interplanar interconnects within a substrate, or between a plurality of substrates). In a still further example, the channels can intersect within a single plane and across a plurality of planes. In still further examples, the channel can have a 3D shape. For example, the channel can be a coil, a toroid, an arc, or a helix.

The method of producing the channel, creating an inlet or an outlet, creating channels and otherwise preparing a substrate for implementation of channels can further include the use of at least one of photolithography, wet etching, reactive ion etching, soft lithography, two-photon lithography, 3D printing, or forming the channel around a sacrificial template.

These illustrative examples are given to introduce the reader to the general subject matter discussed here and are not intended to limit the scope of the disclosed concepts. The following sections describe various additional features and examples with reference to the drawings in which like numerals indicate like elements, and directional descriptions are used to describe the illustrative embodiments but, like the illustrative embodiments, should not be used to limit the present disclosure. The elements included in the illustrations herein may not be drawn to scale.

<FIG> is an illustration of a line scan <NUM> having a scan width of one pixel (depicted as image height in the example of <FIG>) and a scan length of about <NUM> pixels (depicted as image width in the example of <FIG>). Droplets <NUM> have a dark boundary and a lighter interior in bright-field illumination (e.g., white light) that makes the droplets <NUM> detectable to the sensor. In the example of <FIG>, a single-pixel wide area of a full frame camera image was extracted from the full frame image for analysis. Accordingly, higher frame rates (e.g., greater than about <NUM> frames per second (fps)) and real time processing was possible. The system included a beam splitter to send some light to a pointgrey camera (e.g., a camera that performed line scans) as well as a high-speed camera (e.g., a fast camera that performed full frame imaging). The images were processed and data <NUM> was provided (see <FIG> is a graph showing light intensity (referred to as "Mean Pixel Intensity") as a function of position (indicated by Pixel number across the image provided in the example of <FIG>). Areas of higher light intensity indicated either no droplets <NUM> present or the center of the droplets <NUM> was observed. Decreased light intensity indicated the edges of the droplets <NUM>, as shown as darker areas in the example of <FIG>.

<FIG> is an illustration of a line scan <NUM> having a scan width of two pixels (depicted as image height in the example of <FIG>) and a scan length of about <NUM> pixels (depicted as image width in the example of <FIG>). Droplets <NUM> have a dark boundary <NUM> and a lighter interior <NUM> in bright-field illumination (e.g., white light) that makes the droplets <NUM> detectable to the sensor. In the example of <FIG>, a two pixel wide area of a full frame camera image was extracted from the full frame image for analysis as described above. The images were processed and data <NUM> was provided (see <FIG> is a graph showing light intensity (referred to as "Mean Pixel Intensity") as a function of position (indicated by Pixel number across the image provided in the example of <FIG>). Areas of higher light intensity indicated either no droplets <NUM> present or the center of the droplets <NUM> was observed. Decreased light intensity indicated the dark boundary <NUM> of the droplets <NUM>, as shown as darker areas in the example of <FIG>.

<FIG> are illustrations of a first line scan <NUM> and its accompanying first light intensity graph <NUM>, and a second consecutive line scan <NUM> and its accompanying second light intensity graph <NUM>. In the example of <FIG>, sorting was performed by analyzing the light intensity signal (e.g., recorded brightness) obtained from the first line scan <NUM> and the second line scan <NUM>. Each pixel of both line scans <NUM>, <NUM> was aligned to a predetermined position along the channel. In the example of <FIG>, the pixel intensity was calculated as an average pixel intensity since the line consisted of more than one pixel in width (e.g., a <NUM> pixel width line scan as in the example of <FIG>). Droplets moving along a main channel <NUM> show up in both light intensity graphs <NUM>, <NUM> as a decrease in light intensity relative to the brightness of adjacent pixels, as in the examples of <FIG> and <FIG>. In the example of <FIG>, a side channel was attached to the main channel <NUM> at a position between pixel <NUM> and pixel <NUM>. Additionally, an electrode was positioned next to the side channel. When actuated by the light intensity signal, a voltage was applied to the electrode, forcing a droplet into the side channel. Actuating the electrode was based on dielectrophoresis. In the example of <FIG>, a non-uniform electric field of the electrode was combined with the difference between the dielectric constant of the medium and the dielectric constant of a droplet <NUM>. The electric field interaction (i.e., dielectrophoretic effect) caused a force on the droplet <NUM>, forcing it into the side channel. A reference droplet <NUM> was following the droplet <NUM>. The reference droplet <NUM> is seen in the first line scan <NUM> and the second line scan <NUM>, wherein the droplet <NUM> is seen in the first line scan <NUM> and was sorted into the side channel and is not visible in the second line scan <NUM>. The droplet <NUM> was forced into the side channel because its dielectric constant triggered the dielectrophoretic effect, placing the electric field of the droplet <NUM> into the working range of the electrode. If no voltage was applied (i.e., no dielectrophoretic effect occurred), the channel configuration ensured that a droplet not intended to be sorted (e.g., the reference droplet <NUM>) was not forced into the side channel, but maintained flow along the main channel <NUM>. When a droplet was observed at a position just before the side channel, a decision was made in real time by a program whether or not a voltage was applied to deflect the droplet or not. Sorting was further verified by confirming that there was no decrease in the light intensity signal in the second light intensity graph <NUM>.

Claim 1:
A system for detecting a particle in a channel (<NUM>), comprising:
a channel for comprising at least one particle dispersed in a medium, such that the at least one particle is moving from a first end of the channel (<NUM>) to a second end of the channel;
a source of electromagnetic radiation that illuminates at least a portion of the channel;
a first line sensor configured to detect spectral information for identifying the at least one particle, wherein the first line sensor is positioned along a linear axis (<NUM>) of the illuminated portion of the channel (<NUM>) such that the first line sensor is substantially parallel to a direction of movement of the at least one particle through the channel (<NUM>); and
a first optical system that focuses and aligns the illuminated portion of the channel (<NUM>) to the first line sensor characterized in that:
the system further comprises a second '<NUM>. line sensor positioned within the Z. region of illumination, wherein the second line sensor is disposed perpendicular to the linear axis (<NUM>) of the channel to determine a position of the particle within the channel (<NUM>).