Patent Application
20240389928
Eccrine sweat is an attractive class of biofluid suitable for non-invasive monitoring of body chemistry. Sweat contains multiple biomarkers relevant to physiological health including electrolytes, metabolites, hormones, proteins, and exogenous agents. The intermittent or continuous assessment of these and other biomarkers in sweat may offer time-dynamic insight into metabolic processes of the body, relevant to applications ranging from athletic performance to medical diagnostics.
Traditional approaches for sweat collection employ absorbent pads or microbore tubes pressed against the epidermis by virtue of bands or straps to capture sweat as it emerges from the skin. These methods suffer from numerous limitations, such as requiring trained personnel, special handling, and costly laboratory equipment; being incompatible with real-time sweat analysis; and being prone to sample contamination or loss.
More recently, skin-interfaced microfluidic systems have been developed to offer capabilities for personalized assessment of health, nutrition, and/or wellness through non-invasive analysis of sweat. Adapting concepts from traditional lab-on-chip technologies, these wearable epidermal microfluidic (or “epifluidic”) platforms may provide significant insights critical for developing personal assessments. Current device fabrication methods employ soft lithography utilizing low modulus (flexible) materials that require significant time, labor, and resource-intensive cleanroom processes to produce fluidic network designs that are restricted to planar or two-dimensional (2D) layouts. Soft lithography requires high-precision molds to form discrete, patterned layers of an elastomeric material (e.g., poly (dimethylsiloxane), PDMS) that when bonded together yield a sealed device, providing a seamless, non-irritating epidermal interface. Traditionally, producing molds with sufficient feature resolution (>20 μm) requires expensive, time-consuming processing methods (micromachining, micromilling, etc.) and access to specialized (i.e., cleanroom) environments. Such requirements result in elongated device design cycles, inequitable access to equipment necessary for innovation, and additional challenges for commercial deployment due to incompatibilities with large-scale manufacturing.
Various skin-interfaced microfluidic systems for epidermal sampling including microfluidic networks defined in flexible substrates are disclosed in U.S. Patent Application Publication No. 2021/0000395 A1 to Rogers et al. Such publication discloses microfluidic networks defined in a flexible substrate of polymeric material (e.g., polydimethylsiloxane (PDMS) among others), with the flexible substrate being used in conjunction with an adhesive layer (for adhering the device to skin) and a capping layer (for enclosing channels defined in the substrate). Each microfluidic channel defined in the flexible substrate has a fixed height, such as a channel height of 300 μm defined in a 400 μm thickness substrate. Capillary burst valves of selected burst valve pressures permit biofluid (e.g., sweat) to be supplied to different channels of a microfluidic network in a desired sequence.
The use of flexible substrates and soft lithography limits the ability to produce features of varying heights in microfluidic networks. Additionally, conventional devices have limited reservoir capacity, with limited ability to collect multiple, independent pristine samples during an extended period of active collection.
Need exists in the art for skin-interfaced microfluidic systems that address limitations associated with conventional systems.
The present disclosure relates to epidermal microfluidic devices for capture, storage, and analysis of sweat. Additive manufacturing such as three-dimensional (3D) printing may be used to produce epidermal microfluidic devices using rigid materials, with features typically not obtainable using 2D fabrication methods. In certain embodiments, epidermal microfluidic devices include microfluidic features (e.g., portions of channels, valves within channels, and/or portions of reservoirs) with gradient variation in height thereof. In certain embodiments, a rigid substrate of an epidermal microfluidic devices is designed to permit flexure during use without breakage and without distortion of microfluidic features therein, such as by providing channel-defining, reduced-width linking portions extending outward between a central portion and reservoir portions, with the reservoir portions lacking direct coupling therebetween. A method for collecting sweat with an epidermal microfluidic system is further provided, in which an epidermal interface layer comprising a flexible material and defining a first aperture is adhered to skin of a user. A first epidermal microfluidic device may be provided over the epidermal interface layer and used to collect sweat of the user. After reservoirs of the first epidermal microfluidic device are filled, the first epidermal microfluidic device may be removed from the epidermal interface layer, and a second epidermal microfluidic device may be provided over the same epidermal interface layer, and used to collect further sweat of the user. One or more adhesive gaskets may be provided between an epidermal interface layer and sequentially used epidermal microfluidic devices. Any suitable number of epidermal microfluidic devices may be used in sequence in conjunction with a single epidermal interface layer, to collect multiple independent, pristine samples during an extended period of active collection, wherein such devices may include rigid substrates embodying features disclosed herein.
In one aspect, the disclosure relates to an epidermal microfluidic device comprising a unitary rigid substrate and an adhesive layer. The unitary rigid substrate forms a body defining a fluid inlet port, a plurality of fluidic reservoirs, and a plurality of fluidic channels permitting fluidic communication between the fluid inlet port and the plurality of fluidic reservoirs, wherein portions of at least some of the fluidic reservoirs and/or at least some of the plurality of fluidic channels comprise gradient variations in height within the rigid substrate. The adhesive layer defines a first aperture positionally registered with the fluid inlet port, the adhesive layer being configured to be positioned between the rigid substrate and skin of a user.
In certain embodiments, the rigid substrate comprises an elastic modulus of at least 500 MPa.
In certain embodiments, the epidermal microfluidic device further comprises a reservoir capping layer arranged between the rigid substrate and the adhesive layer, wherein the reservoir capping layer comprises a second aperture positionally registered with the first aperture and the fluid inlet port.
In certain embodiments, the epidermal microfluidic device further comprises: an epidermal interface layer comprising a flexible material, configured to be positioned between the adhesive layer and the rigid substrate, and defining a third aperture; and an adhesive gasket configured to be positioned between the epidermal interface layer and the rigid substrate, and defining a fourth aperture; wherein the third aperture and the fourth aperture are positionally registered with the first aperture and the fluid inlet port.
In certain embodiments, the adhesive gasket comprises a maximum width that is smaller than a maximum width of the adhesive layer.
In certain embodiments, the body of the rigid substrate comprises a plurality of fused dots, rods, or layers.
In certain embodiments, the plurality of fluidic channels comprises one or more capillary burst valves that comprise gradient variations in height within the rigid substrate.
In certain embodiments, portions of at least some of the fluidic reservoirs comprise gradient variations in height within the rigid substrate.
In another aspect, the disclosure relates to an epidermal microfluidic device comprising a unitary rigid substrate and an adhesive layer. The unitary rigid substrate forms a body comprising a central portion, a plurality of distal portions, and a plurality of linking portions extending outward from the central portion and coupling the central portion to the plurality of distal portions, wherein the central portions defines a fluid inlet port, the plurality of distal portions define a corresponding plurality of fluidic reservoirs, and a plurality of fluid channels extend through the plurality of linking portions to provide fluid communication between the fluid inlet port and the plurality of fluidic reservoirs. Each distal portion is joined by a single corresponding linking portion to the central portion, and each linking portion has a maximum width that is less than a maximum width of each distal portion. The rigid substrate is devoid of material joining any linking portion to any other linking portion except through the central portion, and the rigid substrate is devoid of material joining any distal portion to any other distal portion except through the central portion. The adhesive layer defines a first aperture positionally registered with the fluid inlet port, the adhesive layer being configured to be positioned between the rigid substrate and skin of a user.
In certain embodiments, each linking portion of the plurality of linking portions comprises a serpentine shape.
In certain embodiments, each distal portion of the plurality of distal portions comprises a ventilation region configured to ventilate a fluidic reservoir of the defined in the distal portion.
In certain embodiments, the plurality of fluidic channels comprises one or more capillary burst valves.
In another aspect, the disclosure relates to a method for collecting sweat with an epidermal microfluidic system, the method comprising multiple steps. One step includes adhering an epidermal interface layer comprising a flexible material to skin of a user, the epidermal interface layer defining a first aperture. Another step includes providing a first epidermal microfluidic device over the epidermal interface layer, the first epidermal microfluidic device comprising a first body defining a first fluid inlet port, a plurality of first fluidic reservoirs, and a plurality of first fluidic channels permitting fluidic communication between the first fluid inlet port and the plurality of first fluidic reservoirs, wherein at least one adhesive gasket defining at least one gasket aperture is arranged between the first epidermal microfluidic device and the epidermal interface layer, with the first aperture and the at least one gasket aperture being positionally registered with the fluid inlet port. Another step includes collecting sweat of the user supplied through the first aperture, the at least one gasket aperture, the first fluid inlet port, and the plurality of first fluidic channels into the plurality of first fluidic reservoirs. Another step includes removing the first epidermal microfluidic device from the epidermal interface layer. Another step includes providing a second epidermal microfluidic device over the epidermal interface layer, the second epidermal microfluidic device comprising a second body defining a second fluid inlet port, a plurality of second fluidic reservoirs, and a plurality of second fluidic channels permitting fluidic communication between the second fluid inlet port and the plurality of second fluidic reservoirs, wherein at least one adhesive gasket defining a second aperture is arranged between the second epidermal microfluidic device and the epidermal interface layer, with the second aperture and the at least one gasket aperture being positionally registered with the second fluid inlet port. Another step includes collecting sweat of the user supplied through the second aperture, the at least one gasket aperture, the second fluid inlet port, and the plurality of second fluidic channels into the plurality of second fluidic reservoirs.
In certain embodiments, the first body is defined by a first unitary rigid substrate, in which portions of at least some of the first fluidic reservoirs and/or at least some of the plurality of first fluidic channels comprise gradient variations in height within the first unitary rigid substrate, and the second body is defined by a second unitary rigid substrate, in which portions of at least some of the second fluidic reservoirs and/or at least some of the plurality of second fluidic channels comprise gradient variations in height within the second unitary rigid substrate.
In certain embodiments, the first body is defined by a first unitary rigid substrate and comprises (i) a first central portion defining the first fluid inlet port, (ii) a plurality of distal portions defining the plurality of first fluidic reservoirs, and (iii) a plurality of first linking portions extending outward from the first central portion, defining the plurality of first fluidic channels, and coupling the first central portion to the plurality of first distal portions, wherein each first distal portion is joined by a single corresponding first linking portion to the first central portion, each first linking portion has a maximum width that is less than a maximum width of each distal portion, the first unitary rigid substrate is devoid of material joining any first linking portion to any other first linking portion except through the first central portion, and the first unitary rigid substrate is devoid of material joining any first distal portion to any other first distal portion except through the first central portion; and the second body is defined by a second unitary rigid substrate and comprises (i) a second central portion defining the second fluid inlet port, (ii) a plurality of distal portions defining the plurality of second fluidic reservoirs, and (iii) a plurality of second linking portions extending outward from the second central portion, defining the plurality of second fluidic channels, and coupling the second central portion to the plurality of second distal portions, wherein each second distal portion is joined by a single corresponding second linking portion to the second central portion, each second linking portion has a maximum width that is less than a maximum width of each distal portion, the second unitary rigid substrate is devoid of material joining any second linking portion to any other second linking portion except through the second central portion, and the second unitary rigid substrate is devoid of material joining any second distal portion to any other second distal portion except through the second central portion.
In certain embodiments, each first linking portion, and each second linking portion, comprises a serpentine shape.
In certain embodiments, each of the first unitary rigid substrate and the second unitary rigid substrate comprises an elastic modulus of at least 500 MPa.
In certain embodiments, the first body of the first rigid substrate and the second body of the second rigid substrate comprises plurality of fused dots, rods, or layers.
In certain embodiments, the plurality of first fluidic channels comprises a plurality of first capillary burst valves, and the plurality of second fluidic channels comprises a plurality of second capillary burst valves.
In another aspect, any two or more features of aspects and/or embodiments disclosed herein may be combined for additional advantage.
Additional features and advantages will be set forth in the detailed description which follows, and in part will be readily apparent to those skilled in the technical fields of microfluidic and epifluidic devices. It is to be understood that the foregoing general description, the following detailed description, and the accompanying drawings are merely exemplary and intended to provide an overview or framework to understand the nature and character of the claims.
Aspects of the present disclosure relate to epidermal microfluidic devices for capture, storage, and analysis of sweat. Additive manufacturing such as three-dimensional (3D) printing may be used to produce epidermal microfluidic devices using rigid materials, with microfluidic features typically not obtainable using 2D fabrication methods. Such microfluidic features with gradient variation in height may include portions of channels, capillary burst valves within channels, and/or portions of reservoirs. Rigid substrates permit features with gradient variation in height to be fabricated, and (in contrast to conventional devices fabricated of flexible materials) promote dimensional stability of microfluidic features with gradient variation in height even if a rigid substrate is subjected to flexure. In certain embodiments, a rigid substrate of an epidermal microfluidic devices is designed to permit flexure during use without breakage and without distortion of microfluidic features therein, such as by providing channel-defining, reduced-width linking portions extending outward between a central portion and reservoir portions, with the reservoir portions lacking direct coupling therebetween.
In certain embodiments, a rigid substrate is produced by an additive manufacturing technique such as 3D printing. In general, these methods create solid objects in a sequential, layer-by-layer manner directly from a computer-aided design (CAD) file, typically involving deposition of liquid and/or solid materials that are cured, wherein a resulting 3D printed structure comprises a plurality of fused dots, rods, or layers. Examples of 3D printing techniques include vat photopolymerization techniques, extrusion, fused deposition modeling, direct ink writing, and the like. Although 3D printer manufacturers advertise printers with high resolution (e.g., x-y resolution of 50 microns, and z resolution of 5 microns), in practice it has historically been challenging to reliably obtain complex devices with channel dimensions of less than 100 microns that simultaneously meet various application specific requirements such as biocompatibility and/or optical clarity, while preserving printability. Careful attention to printer-dependent parameters and chemistry of printer materials improves reproducibility of high resolution microfluidic device designs capable of meeting application specific requirements.
The term “rigid substrate” as used herein refers to a substrate having an elastic modulus (a/k/a Young's modulus) value of at least 500 MPa, at least 600 MPa, at least 700 MPa, at least 800 MPa, at least 900 MPa, at least 1000 MPa, or a value within a range of from 500 to 3,000 MPa, or a value within a range of from 600 to 1,500 MPa, or a value of 700 to 1,200 MPa, or a value of about 975 MPa, in certain embodiments. In certain embodiments, a rigid substrate is produced by 3D printing and has sufficient rigidity to resist deformation of microfluidic features (e.g., channels, capillary burst valves, reservoirs, etc.) when the substrate is subjected to moderate flexural forces, but is sufficiently pliable for the substrate to resist breakage when subjected to such forces. Moreover, a rigid substrate is preferably resistive to uncontrolled fluid flow during physical handling (e.g., exertion of finger pressure).
To enhance pliability of a rigid substrate having microfluidic features defined therein, in certain embodiments a substrate may include: a central portion having a fluid inlet port, multiple distal portions each defining a corresponding fluidic reservoir, and multiple linking portions defining fluidic channels that provide fluid communication between the fluid inlet port and the multiple fluidic reservoirs, wherein each distal portion is joined by a single corresponding linking portion to the central portion, each linking portion is narrower than each distal portion (and optionally may comprise a serpentine shape), the substrate is devoid of material joining any linking portion to any other linking portion except through the central portion, and the substrate is devoid of material joining any distal portion to any other distal portion except through the central portion. Restated, providing narrow linking portions between a central portion and distal portions of a rigid substrate, without rigid substrate material (other than the central portion) coupling the respective linking portions and respective distal portions, enables the distal portions to be flexed or twisted away from a common plane to better conform to non-planar skin surfaces. The foregoing arrangement utilizes rigid substrate material only where necessary to support microfluidic structures (e.g., ports, channels, reservoirs, etc.), and permits the rigid substrate to withstand being subjected to at least moderate torsion and/or flexure without breakage, thereby permitting a rigid substrate to be used as part of an epidermal microfluidic device that conforms to non-planar skin of a user. Use of a rigid substrate instead of flexible substrate materials permits microfluidic features to remain undeformed or only minimally deformed when an epidermal microfluidic device is subjected to flexural or torsional forces, thereby eliminating concerns of unconstrained fluid flow if flexible substrate materials were employed instead.
In certain embodiments, at least some microfluidic features (e.g., reservoirs, channels, and/or capillary burst valves in channels) within a rigid substrate may comprise gradient variations in height. Although it is commonplace to provide gradient variation in width of microfluidic features, it is generally not feasible to produce gradient variation in height of microfluidic features using conventional 2D fabrication techniques. Gradient variations in height of microfluidic features within a rigid substrate may be produced by 3D printing, which may yield features having a plurality of fused dots, rods, or layers. Such features may appear to have continuous variation in height, but under a microscope may exhibit steps. Accordingly, the term “gradient variation in height” as used herein may refer to height variations produced by steps of less than 30 μm, less than 25 μm, less than 20 μm, less than 15 μm, less than 10 μm, less than 5 μm, or less than 2 μm in certain embodiments.
In certain embodiments, a fluid inlet port of a rigid substrate is configured to receive sweat of a user, where the sweat flows through at least one microfluidic channel to a series of capillary burst valves (CBVs) and corresponding reservoirs. A CBV at the ingress of each reservoir permits fluid flow only after a set pressure is exceeded, thereby enabling time-sequential sweat collection. Operating without use of actuation or moving components, capillary burst valve burst pressure is governed by valve geometry. For a microfluidic channel with fixed dimensions, the burst pressure of a CBV becomes a function of channel diverging angles, and is also inversely proportional to channel size. In certain embodiments, integrated ventilation holes are provided proximate to each reservoir to eliminate backpressure that would be generated by trapped air and impede ingress of sweat into the reservoirs.
In certain embodiments, at least one adhesive layer is provided between a rigid substrate and skin of a user, wherein the adhesive layer defines a first aperture positionally registered (i.e., aligned with) a fluid inlet port of the substrate, wherein the first aperture may be defined by laser patterning (e.g., laser ablation). The first aperture serves as a sweat collection area may have any suitable size and shape, such as round, square, star-shaped, etc.). In certain embodiments, the first aperture comprises a patterned opening that is larger than the fluid inlet port of the substrate. Although any suitable size and area of the first aperture may be selected, in certain embodiments the first aperture comprises an area in a range of 100 mm2 to 400 mm2, or in a range of 150 mm2 to 300 mm2, or in a range of about 175 mm2 to about 250 mm2, or a value of about 180 mm2 or about 200 mm2. In certain embodiments, the at least one adhesive comprises a biocompatible, medical grade adhesive layer, optionally being transparent, such as a fiber-reinforced adhesive polymeric (e.g., polyester) transfer tape material. One example of such a material is 3M Medical Transfer Adhesive 1524 (80 μm thickness) including randomly oriented polyester fibers and pressure sensitive acrylic adhesive (3M Company, St. Paul, MN, US). In certain embodiments, a single adhesive layer is provided; in other embodiments, multiple adhesive layers may be provided. In certain embodiments, one adhesive layer forms an aperture-defining gasket proximate to a central portion of a rigid substrate, and another aperture adhesive layer is larger in width and directly contacts an epidermal (skin) surface of a user, wherein an epidermal interface layer may be arranged between the foregoing two adhesive layers.
In certain embodiments, a rigid substrate comprises a skin-facing surface and a skin-opposing surface, wherein reservoir openings may be provided in the distal portions of the rigid substrate along the skin-facing surface of the rigid substrate, and the reservoir openings may be covered by a reservoir capping layer, which may be continuous or discontinuous in character. In certain embodiments, a reservoir capping layer comprises a soft material such as PDMS. In certain embodiments, a capping layer comprises a thickness in a range of from 20 μm to 50 μm, or a range of from 25 μm to 40 μm, or a thickness of about 30 μm. In certain embodiments, a reservoir capping layer may be formed directly in or on a substrate by three-dimensional printing and/or spin coating, or may be prefabricated and physically placed over at least portions of a skin-facing surface of a rigid substrate. In certain embodiments, a reservoir capping layer may be formed by pouring liquid polymer precursor (e.g., liquid PDMS with a curing agent, optionally including a pigment) onto a sacrificial film, spin coating, and curing by heat and/or other means, followed by laser cutting. Any suitable method for bonding a reservoir capping layer to a reservoir may be used, including surface modification of a rigid substrate (e.g., corona treatment with air plasma) followed by thermal bonding (e.g., lamination) between a substrate and a reservoir capping layer; chemical bonding; or adhesive bonding. In certain embodiments, a single unitary reservoir capping layer may be provided; in other embodiments, discrete (disconnected) reservoir capping layer portions may be individually provided in contact with corresponding distal portions of a rigid substrate. If a reservoir capping layer is arranged in contact (e.g., underlying) a central portion of a rigid substrate, then the reservoir capping layer may comprise a second aperture that is positionally registered (i.e., aligned) with an aperture (e.g., first aperture) of an adhesive layer as well as with the fluid inlet port of a rigid substrate.
In certain embodiments, an epidermal interface layer defining a third aperture may be provided between a skin-contacting adhesive layer and a reservoir capping layer (or between a skin-contacting adhesive layer and a rigid substrate) of an epidermal microfluidic device. In certain embodiments, an epidermal interface layer is configured to permit reversible adhesion (optionally aided by an adhesive gasket) between a skin-contacting adhesive layer and a capping layer, enabling a first rigid substrate (with associated capping layer) to replaced, after reservoirs thereof are filled, with a second rigid substrate (and associated capping layer), thereby permitting multiple epidermal microfluidic devices (or portions thereof) to collect multiple independent pristine sweat samples during an extended period of active collection. Utilizing an epidermal interface layer provides biocompatible fluid-tight interface with the epidermis, which provides a significant technical benefit when switching rigid substrates because it avoids adhesion challenges and potential contamination that would result in trying to adhere second and subsequent substrates to wet (i.e., sweaty) skin. In certain embodiments, an epidermal interface layer comprises PDMS or another biocompatible polymeric material, and has a thickness in a range of 200 μm to 600 μm, or in a range of 300 μm to 500 μm, or about 400 μm.
In certain embodiments, one or more functional constituents configured to interact with sweat may be arranged in or on microfluidic features of a unitary rigid substrate (e.g., supplied during fabrication, such as prior to bonding of a reservoir capping layer in certain embodiments). In certain embodiments, colorimetric assay chemicals and/or biological moieties may be arranged in or on microfluidic features of a rigid substrate to enable concentration analyses of one or more sweat constituents (e.g., chloride). In certain embodiments, dye may be arranged in or on microfluidic features of a rigid substrate to enable sweat to be visualized.
Various examples of epidermal microfluidic device designs are disclosed herein, wherein it is to be recognized that features of any one or more device designs may be combined or substituted with other device designs herein, unless indicated to the contrary.
With continued reference to
Successful fabrication of a fully-enclosed microfluidic channel with feature sizes at the x-y plane resolution limit of current DLP printers (˜30 μm to 50 μm) depends on several related factors including: design aspects (e.g., channel vertical position), print process parameters (e.g., layer height, layer cure time (LCT), print speed), and printer hardware (e.g., projector light power, wavelength). Optimization of user-adjustable factors results in a robust print process suitable for producing microfluidic devices with sufficient optical clarity, dimensional fidelity, and mechanical performance for use in epidermal microfluidic applications.
As expected, epidermal microfluidic device performance is dependent on the dimensional accuracy of a 3D fabrication process. If not quantified, unintended deviation from designed dimensions can adversely affect component performance (e.g., CBV burst pressure) or measurement accuracy (e.g., sweat volume, sweat rate, etc.). To evaluation dimensional accuracy of a 3D fabrication process, nine test channels of substantially square cross-sectional shapes ranging in dimensions from 100 μm to 900 μm (in both width and height) and 5 mm long were fabricated by 3D printing, to permit determination of the minimum repeatable printable channel dimensions and sidewall thickness (minimum 50 μm).
Experimental studies also have revealed a strong influence of layer cure time (LCT) on 3D print success using a DLP-based 3D printer and resulting quality of DLP 3D printed substrates of epidermal microfluidic devices. The LCT defines the energy dose used to crosslink a photopolymer used in DLP-based 3D printing, given in time (seconds). The projector wavelength is defined by hardware (e.g., 385 nm in production of the test channels 70A-70I) and varying projector power is not possible by a user of commercially available DLP-based 3D printers. Systematic studies of four LCT settings (i.e., from tested minimum (0.54 s) to maximum values (2.0 s; 0.6 s interval) beyond which channels could not be fabricated successfully) establish a relationship between 3D print performance (i.e., for successful channel printing), dimensional accuracy, and optical clarity. Measurement results obtained from optical microscope are plotted in
Additional systematic experiments establish the DLP-printable design space for dimensions relevant to epidermal microfluidic devices (e.g., 100 μm-600 μm). An encapsulated microfluidic channel capable of supporting unrestricted fluid flow, in contrast to a sealed or partially-restricted channel, defines a successful print. Intuitively, print failure rate increases as the enclosed channel dimensions approach the printer x-y plane resolution limit (e.g., ˜32 μm square pixel for the selected DLP-based printer). Experimental results showed that channel dimensions of 100 μm (either width or height) correspond to the lower limit for a successful printed device.
The optical transparency of a 3D-printed microfluidic device depends on several factors including material selection, printer hardware (e.g., build plate, vat surface material), post processing, and surface roughness. In contrast to the typical surface roughness feature size necessary for optical transparency (<10 nm), DLP printers produce parts with microscale surface roughness resulting in a semi-translucent appearance.
As mentioned previously, pixel size of a digital micromirror device (DMD) governs the x-y plane resolution of a DLP 3D printer. Minute gaps between individual DMD elements locally reduce reflected light intensity, yielding a surface roughness with features corresponding to DMD pixel size and layer height. Although specialized printing methods (e.g., grayscale) or printer hardware (e.g., oscillating lenses) offer sophisticated strategies to reduce aliasing and improve surface roughness, the fundamental approach to eliminating this defect mode is enhancing the uniformity of projected light to ensure complete photopolymerization. UV-Vis spectroscopy experiments examined the transmission properties of 3D-printed microcuvettes produced by such method in comparison to a commercial plastic cuvette. In particular, a UV-Vis spectrophotometer (UV-1900i, Shimadzu, Japan) enabled quantification of the optical transmission properties of the 3D printed devices (300 nm-1000 nm, 0.5 nm interval). A commercial plastic cuvette (10 mm pathlength, Shimadzu) served as a reference (control). Four (4) sets of 3D-printed cuvettes (N=3 per set) utilizing a different LCT setting (0.54 s, 0.8 s, 1.4 s, 2 s) enable quantification of the relationship between LCT and optical transmission (dimensions: 50 mm height, 8 mm width, 1 mm pathlength, ˜21 μL volume). The inventors have found that increasing the exposure dose by lengthening the LCT eliminated the observed grid pattern defects (from DMD element gaps) and improved optical transparency.
In addition to LCT, layer height affects both overall device quality (e.g., vertical resolution, optical clarity, channel roughness) and print time, which corresponds to device yield. Conventional approaches to vat photopolymerization utilize constant values for a given print run (i.e. fixed layer height, LCT). At present, only one manufacturer (i.e., Formlabs) supports an adaptive layer height process to increase print speed by adjusting layer height as a function of model detail (i.e., small layers for fine features, thick layers for coarse features). Adaptive printing is an attractive process for obtaining expanded design flexibility for 3D-printed epidermal microfluidic systems. Although not supported by default, a combination of custom software and manual G-code (geometric code) programming enables definition of both layer height and LCT as a function of model dimensions. By comparison to a constant LCT and layer height setting printing process, this approach enables successful, time-efficient fabrication of epidermal microfluidic with complex geometries and high device quality.
Colorimetric assays facilitate passive, battery-free in situ quantitative measurement of sweat biomarkers. A chemical reagent reacts with a target species to generate an optical signal proportional to analyte concentration. Accurate colorimetric analysis requires channels with uniform height (i.e., path length), a high degree of optical transparency, and integrated color reference markers to support reliable image processing under variable ambient lighting conditions. The layer-by-layer control over LCT and layer height parameters during enabled by an adaptive printing process is highly desirable for fabricating microfluidic devices with the requisite surface finish and optical transparency to support colorimetric analysis. The inventors have found that an adaptive LCT print process beneficially improves optical transparency of microfluidic features in 3D printed substrates, wherein optical clarity for two representative epidermal microfluidic device reservoirs manufactured utilizing a layer-constant LCT (0.54 s, 2 s) was found to increase with longer LCT. While beneficial for reducing nonuniform illumination, the increased UV dose results in undesirable curing of resin in enclosed features (channels, CBVs). By comparison, an adaptive printing process (AP1) utilizing an LCT of 0.54 s for the reservoir surface and an LCT of 2 s for subsequent layers enables fabrication of microfluidic substrate with a translucent imaging plane, transparent faces, and preservation of internal channel features. An inverse adaptive printing process (AP2; base LCT: 2 s, subsequent layer LCT: 0.54 s) results in an optically transparent imaging plane and a translucent device.
Systematic benchtop experiments evaluate the suitability of devices fabricated by adaptive printing for colorimetric analysis. The colorimetric assay silver chloranilate produces a dark violet color response proportional to chloride concentration. Imaging the device with a smartphone camera enables color extraction and subsequent quantification of color response. The inclusion of a color balance chart facilitates color calibration for each image. As previously reported by others, converting images from native red-green-blue (RGB) color space to CIELAB color space (which expresses color as lightness (L), amount of green to red (a*), and amount of yellow to blue (b*)) ensures device-independent color sampling. Conversion of the a* and b* components to chroma (C*) by the relation
yields a calibration curve with chloride concentration by a power-law relation). Calibration charts were created from 3D-printed epidermal microfluidic device substrates with different LCT parameters and reference colorimetric assay solutions, revealing that the improvement in optical clarity with increasing LCT provides a corresponding enhancement in the range of detectable color measurements. Thus, an adaptive printing process is helpful for fabricating epidermal microfluidic device substrates with an optical transparency sufficient to support colorimetric analysis.
CBVs are a key component for the sequential analysis of sweat biomarkers in many epifluidic platforms. Time-dynamic variations in sweat rate arising from physical (e.g., sweat gland density), physiological (e.g., exertion, emotion), and external factors (e.g., temperature, pH) result in corresponding changes in analyte concentration. As previously described, CBVs prevent flow for fluid pressure conditions below a designed threshold (bursting pressure, BP); when the fluid pressure exceeds the BP, the CBV immediately bursts. Operating without use of actuation or moving components, CBV BP is governed by valve geometry
The Young-Laplace equation describes the BP for a CBV (rectangular channel) as
where σ is the fluid surface tension; θ+ is the critical advancing contact angle for the channel (material dependent, θ+=120° for PDMS); θI* is the minimum of either θ+β or 180°; β is the (horizontal) channel diverging angle; b and h are the diverging channel width and height, respectively. As the second term of Equation 2 is constant for a planar (i.e., two-dimensional or 2D) CBV, channel width and diverging angle govern the BP for a given CBV. In practice, epidermal microfluidic device designs utilize geometric restrictions (i.e., modifications to channel width) to control valve BP.
Utilization of 3D printing to produce substrates of epidermal microfluidic devices as disclosed herein expands CBV capabilities by enabling a full 3D CBV designs. As a consequence, for a 3D CBV, Equation 2 can be written as
where θ* is the minimum of either θ+γ or 180° and γ is the channel diverging angle (z-axis). It follows that for a microfluidic channel with fixed dimensions, the CBV BP becomes a function of the channel diverging angles (β, γ). Computational predictions of four representative CBV valve designs, presented schematically in
In addition to CBV design parameters, Table 2 provides theoretical CBV BPs and effective theoretical BPs, which considers the theoretical CBV BP and fluidic resistance of the microfluidic channel network. Imperfections resulting from the 3D-printing process result in experimental burst pressure values below theoretical limits.
Benchtop experiments yield measurements of CBV BPs by means of a positive pressure displacement pump apparatus that perfuses water (dyed blue for visualization) into the microfluidic network at defined pressures. A digital microscope (VHX-7100, Keyence Corp., Japan) produced micrographs of the devices. An optical camera (Canon 90D, Canon EF 100 mm f/2.8 L USM lens) provided video capture capabilities (30 frames per second) for device analysis. Measurement of the CBV burst pressure consisted of a “fill test” in which water (dyed blue for visualization) entered a device until flow stopped the CBV. A modular, calibrated pressure displacement flow system (Flow EZ, Fluigent, France) controlled the fluid pressure and permitted near-instantaneous stepwise increase in pressure (0.1 mbar interval, 10 s dwell time). Video observation identified the pressure threshold for fluid to burst each CBV.
Each 3D epifluidic device design described herein was created using computer-aided design (CAD) software (AutoCAD 2019, Autodesk, CA, USA). Subsequent export to a stereolithography readable file (.stl) format yielded a file suitable for direct use by the digital light processing (DLP) resin printer (Prime 110, 385 nm, MiiCraft, Taiwan and Creative CADworks, Ontario, Canada). The included printer control software (Utility, v 6.3.0.t3) provided direct control over print parameters for each file including layer height (5 μm to 50 μm), dose, and lamp power. High-fidelity printing was achieved by application of a removable Kapton polyimide tape over the surface of a polished aluminum build plate. The applied tape was free of bubbles and wrinkles to ensure a smooth build surface free of defects.
Devices were printed using transparent resin (MiiCraft BV-007A, Creative CADworks, Ontario, Canada) and a 10 μm layer height (6 devices per build plate, ˜20 min total print time). Gentle removal of printed parts from the build plate, soaking in 1% detergent solution (Alconox-1232-1, Alconox, NY, USA) under sonication (CPX2800, Fisher Scientific, PA, USA) for 10 min, drying of device using clean dry air (CDA), post-print UV cure for 10 min (CureZone, MiiCraft, Taiwan), and post-cure bake at 70° C. for 30 min (Model 40E Lab Oven, Quincy Lab Inc., IL, USA) yielded a 3D-printed epifluidic device suitable for direct use or integration with PDMS (e.g., a PDMS epidermal interface layer, optionally in combination with a PDMS reservoir capping layer).
A three-step process facilitated printing fully enclosed 3D-printed devices. Printing epifluidic substrates with open reservoirs (Step 1) and post-print removal of uncured liquid resin by CDA (Step 2) enabled enclosure of the substrates with a thin reservoir capping layer (30 μm) by means of a second print process (Step 3). The printed device remained fixed to the build plate during the “print-pause-print” process to ensure feature alignment. Execution of the above-described post processing steps yielded a fully enclosed epifluidic substrate.
To form PDMS portions (capping layer and epidermal interface layer), liquid PDMS (10:1 base: curing agent, Sylgard 184, Dow Inc., MI, USA) with white pigment (3% w/w, Ignite White, Smooth-On, Inc., PA, USA) was poured onto a sacrificial mylar film (2 mil thickness), spin coated for 30 s (400 rpm for reservoir capping layer; 200 rpm for epidermal interface layer), and cured in an oven (70° C., 2 h) to form films with thicknesses of 200 μm and 400 μm, respectively. A CO2 laser cutter (30W Epilog Mini 24, Epilog Laser, Colorado, USA) patterned the PDMS films into the final geometries used in the epifluidic devices. A medical-grade adhesive (1524, 3M Inc., MN, USA) patterned in the same manner and bonded to the PDMS interfacial layer, was provided below the epidermal interface layer for bonding to skin of a user.
Hybrid 3D-printed epifluidic devices utilize bonded PDMS capping layers to enclose 3D-printed microfluidic reservoirs. Modification of a previously reported method facilitated a strong bond between PDMS and the printed device. Specifically, rinsing with isopropyl alcohol (2-propanol, A416, Fisher Scientific, Massachusetts, USA), soaking in DI water (Direct-Q 3 UV Water Purification System, MilliporeSigma, Missouri, USA) for 30 min, corona treating with air plasma (BD-20, Electro-Technic, Illinois, USA) for 30 s followed by immediate immersion in a 12% v/v solution of (3-aminopropyl)triethoxysilane (APTES, 440140, MilliporeSigma, MO, USA) for at least 20 min, rinsing in DI water, and drying with CDA prepared the oven-baked 3D-printed substrate for bonding to PDMS. Pipetting colorimetric reagents or flow visualization dye (Soft Gel Paste, AmeriColor Corp., CA, USA) into pre-determined regions occurred prior to sealing of the 3D-printed device substrate. After a 30 s corona treatment, laminating the PDMS capping layer to the APTES-modified printed surface sealed the epifluidic substrate. Heat treating the assembled substrate and PDMS layer on a hotplate (70° C.) under applied weight (3 kg) for 30 min to form a permanent bond. Removal of the sacrificial mylar layer, release from excess PDMS via laser-cutting, and opening the central sweat ingress points using a 1.5 mm diameter circular punch (reusable biopsy punch, World Precision Instruments) yielded a final hybrid epifluidic device.
A study was performed to evaluate the performance of 3D-printed epifluidic devices (according to the design of
Cleaning of the forearm of each individual with an alcohol wipe prepared the skin for robust adhesion to an epifluidic device including a 3D printed substrate, a reservoir capping layer, an adhesive gasket, an epidermal interface layer, and a skin-facing adhesive layer. The exercise regime comprised stationary cycling under approximately constant working load for 50 min in a controlled laboratory environment (22° C., 55% RH).
Evaluation of sequential generation of aliquots of sweat required periodic monitoring of the filling of an epifluidic device. Once all reservoirs filled, as determined by visual observation, a first (initially attached) device portion (including a first 3D printed substrate, first reservoir capping layer, and first adhesive gasket) was removed from the epidermal interfacial layer and replaced with a second device portion (including a second 3D printed substrate, second reservoir capping layer, and second adhesive gasket, all being identical to their counterparts in the first device portion), was applied to the epidermal interface layer while the volunteer continued to exercise. Evaluation of colorimetric performance of the epifluidic devices required individual subjects to wear two separate epifluidic devices (i.e., one colorimetric and one collection (as a control)), located in close proximity on the same arm. Prior to device removal, a photograph of the colorimetric epifluidic device was recorded at the conclusion of the collection period for image processing and chloride analysis. Extraction of sweat from the individual reservoirs of the collection epifluidic device at the conclusion of the exercise period facilitated chloride measurements using a ChloroChek Chloridometer.
A chloride colorimetric assay solution was produced by thoroughly vortexing 50 mg of silver chloranilate (MP Biomedicals, CA, USA) in 200 μL of a solution of 2% (w/v) polyhydroxyethylmethacrylate (pHEMA, 529265, MilliporeSigma, Missouri, USA) in methanol (A412, Fisher Scientific, Massachusetts, USA) to yield a homogenous suspension. Spotting 2 μL of this solution via laboratory pipette onto the 3D-printed device near the central sweat ingress point, followed by drying in an oven for 30 min prior to encapsulation, prepared the epifluidic device for colorimetric chloride measurements.
To develop standard color and color reference markers, sodium chloride (S271, Fisher Scientific, MA, USA) in DI water was mixed to produced standard test solutions (0, 10, 20, 30, 50, 75, 90, 110, 130, 150 mM), and clinical-grade chloridometer measurements (ChloroChek, ELITech Group, Inc.) yielded validated test solution concentrations. Digital imaging and analysis of sample reservoirs containing one standard solution reacted with the silver chloranilate assay under uniform illumination formed a set of reference images. The sample reservoirs were of the same depth as the epifluidic channels to ensure accurate color representation.
A digital smartphone camera captured images during on-body field tests. A color calibration card (ColorChecker Classic, X-Rite, MI, USA) in the frame of each image facilitated accurate color extraction under various illumination conditions. An open-source photography software package (Darktable 3.0.0, Darktable.org) served as the platform for calibrating images using the color reference card. Analysis of calibrated images using MATLAB (R2019b, The MathWorks Inc., MA, USA) enabled cropping multiple regions of interest from images and extraction of CIELAB color values (L, A, B) for chroma analysis. Mapping of chroma values from colorimetric samples of known reference chloride solutions yielded colorimetric calibration charts with a power-law relationship. The obtained calibration chart supported quantification of the sweat chloride concentration in on-body field testing.
The present disclosure thus provides skin-interfaced wearable systems with integrated microfluidic structures (channel, valve, reservoir) and sensing capabilities (e.g., optical, electrochemical), offering powerful platforms for monitoring the signals arising from natural physiological processes, and providing utility not offered with epifluidic devices having two-dimensional structures produced with flexible substrates.
Those skilled in the art will recognize improvements and modifications to the preferred embodiments of the present disclosure. All such improvements and modifications are considered within the scope of the concepts disclosed herein and the claims that follow. Any of the various features and elements as disclosed herein may be combined with one or more other disclosed features and elements unless indicated to the contrary herein
This application claims benefit of U.S. Provisional Patent Application No. 63/247,894 filed on Sep. 24, 2021, wherein the contents of the foregoing application are hereby incorporated by reference herein.
This invention was made with government support under P20 GM113134 awarded by the National Institutes of Health. The government has certain rights in the invention.
| Filing Document | Filing Date | Country | Kind |
|---|---|---|---|
| PCT/US2022/044543 | 9/23/2022 | WO |
| Number | Date | Country | |
|---|---|---|---|
| 63247894 | Sep 2021 | US |
