BACKGROUND

[0001] Self-wicking assay devices can be used to detect the presence of a target analyte in a sample fluid, for example. These devices can be simple and can often be used typically without specialized training by the user. In some examples, self-wicking assay devices can be widely used for medical diagnostic testing, environmental sample testing, and/or in laboratories, to name a few.

BRIEF DESCRIPTION OF THE DRAWINGS

[0002] FIG. 1 graphically illustrates an example self-wicking assay device in accordance with the present disclosure;

[0003] FIG. 2 graphically illustrates an example self-wicking assay device in use in accordance with the present disclosure;

[0004] FIG. 3 graphically illustrates an example system for manufacturing a self-wicking assay device in accordance with the present disclosure;

[0005] FIG. 4 graphically illustrates an alternative example system for manufacturing a self-wicking assay device in accordance with the present disclosure; and

[0006] FIG. 5 is a flow diagram illustrating an example method of manufacturing a self-wicking assay device in accordance with the present disclosure. DETAILED DESCRIPTION

[0007] Self-wicking assay devices can permit the detection of a target analyte from a sample fluid. These devices can incorporate a porous membrane so that a sample fluid can sequentially run along the porous membrane via capillary flow. When a target analyte is present in the sample fluid, the target analyte can interact with the porous membrane differently in different regions of the device. An example of a self-wicking assay device is a lateral flow device, such as those devices commonly used for pregnancy testing. There are, however, many other applications of lateral flow devices.

[0008] Lateral flow devices typically include multiple functional regions, and as such the substrates in the various regions are typically of a different type of material, and those various substrates can be joined or laminated together at junctures, such as by lamination or other joining techniques. In accordance with the present disclosure, however, a porous membrane can be used at multiple regions, provided a region where the analyte complexes with the analyte- complexing compound can be allowed to interact and become released for downstream testing in a detecting region, e.g., the test and control lines used to detect and validate the presence of an analyte(s), for example. Benefits that can be achieved may include enhancements in reproducibility, reduction in fabrication complexity related to joining and/or overlapping of substrates, and/or more customization at the user level, e.g., selection of test and control compounds and/or placement of test and control compounds.

[0009] In accordance with an example of the present disclosure, a self- wicking assay device (“device”) can include a porous membrane for sequential flow of a sample fluid from a complexing region to a detecting region thereof. The complexing region can include a releasing compound to ameliorate binding between the porous membrane and an analyte introduced by the sample fluid, and can further include an analyte-complexing compound. The detecting region can be positioned downstream from the complexing region along the porous membrane and can include an immobilized testing compound applied to the porous membrane at a first discrete location of the detecting region and an immobilized control compound applied to the porous membrane at a second discrete location of the detecting region. In one example, a fluid receiving region can be included at or upstream from the complexing region to receive a sample fluid so that an analyte in the sample fluid while at the complexing region is available to potentially interact with the analyte-complexing compound. The porous membrane can include, for example, nitrocellulose, charge-modified nylon, polyvinylidene fluoride, or polyethersulfone. The porous membrane in the complexing region can be modified by covalent or ionic bonding, or by in situ polymerization, to form a compound including a polymer having a first moiety with an affinity for the porous membrane and a second moiety that is more hydrophilic than the porous membrane to interact with the analyte; a hydrogel; a polymer with a higher concentration oxygen-, nitrogen-, or sulfur-containing moieties than the porous membrane; a pH sensitive polymer with pH sensitive moieties; a temperature-sensitive polymer; a polymer that exhibits salt- dependent phase change; a micelle-forming polymer; or a self-immolative polymer. The porous membrane in the complexing region can be modified with a compound including an anionic surfactant, a nonionic surfactant, a polyacrylic acid hydrogel, a polyethylene glycol, an acryloyl-containing polymer, a sugar, a polyethylene oxide-polypropylene oxide-polyethylene oxide triblock copolymer, a polyvinylpyrrolidone, a polyacrylamide hydrogel, polyvinyl alcohol, derivatives thereof, or a combination thereof. In other examples, other than the presence of the testing compound and the control compound, the porous membrane at the detecting region can be otherwise unmodified in some examples.

[0010] In another example, a system for manufacturing a self-wicking device can include a first fluidjet ejector to eject a releasing agent including a releasing compound from a first reservoir when the releasing agent is loaded therein, a second fluidjet ejector to eject a testing agent including a testing compound from a second reservoir when the testing agent is loaded therein, and a third fluidjet ejector to eject a control agent including a control compound from a third reservoir when the control agent is loaded therein. The system can also include a hardware controller to generate a command to apply releasing agent onto a complexing region of a porous membrane, apply testing agent to immobilize the testing compound at a first discrete region of the detecting region, and apply control agent to immobilize the control compound at a second discrete region of the detecting region. In one example, the releasing agent, the testing agent, and the control agent are also included as part of the system. In another example, a fourth fluidjet ejector can be included to eject a complexing agent including an analyte-complexing compound from a fourth reservoir when the complexing agent is loaded therein. The complexing agent can be included as part of the system in some examples. In another example, the porous membrane can also be included as part of the system.

[0011] In another example, a method of manufacturing a self-wicking assay device can include applying releasing agent including a releasing compound to a porous membrane to leave the releasing compound at a complexing region of the porous membrane. The releasing agent in this example is not applied to a separate detecting region positioned downstream along the porous membrane relative to the complexing region. The method can also include applying a testing agent including a testing compound to the porous membrane at a first discrete location of the detecting region to provide an immobilized testing compound at the first discrete location. In further detail, the method can include applying a control agent including a control compound to the porous membrane at a second discrete location of the detecting region to provide an immobilized control compound at the second discrete location. In one example, applying the releasing compound can be carried out via a first fluidjet ejector, applying the testing agent can be carried out via a second fluidjet ejector, applying the control agent can be carried out via a third fluidjet ejector, or a combination thereof. In another example, the method can include applying a complexing agent including an analyte-complexing compound to the complexing region after applying the releasing agent. In another example, the method can include applying a second testing agent including a second testing compound to the porous membrane at a third discrete location of the detecting region to provide a second testing compound at the third discrete location, wherein the testing compound includes a molecule to bind an analyte that is complexed with the analyte-complexing compound, the second testing compound includes a second molecule to bind a second analyte that is complexed with the analyte-complexing compound, and wherein the control compound includes a third molecule to bind the analyte-complexing compound that is unbound to the analyte.

[0012] When discussing the self-wicking assay device, the system of manufacturing the self-wicking assay device, and/or the method of manufacturing the self-wicking assay device, such discussions can be considered applicable to one another whether or not they are explicitly discussed in the context of that example. Thus, for example, when discussing a complexing region in a self-wicking assay devices, such disclosure is also relevant to and directly supported in the context of the systems and/or methods, and vice versa. Furthermore, in accordance with the definitions and examples herein, FIGS 1-5 depict various self-wicking assay devices, systems, and methods. These various examples can include various common features. Thus, the reference numerals used to refer to features depicted in FIGS. 1-4 in particular may be the same throughout to avoid redundancy.

[0013] Terms used herein will be interpreted as the ordinary meaning in the relevant technical field unless specified otherwise. In some instances, there are terms defined more specifically throughout or included at the end of the present disclosure, and thus, these terms are supplemented as having a meaning described herein.

Self-wicking Assay Devices

[0014] A self-wicking assay device 100, as illustrated in FIG. 1 , can include a fluid flow pathway for fluid to pass or wick through a porous membrane 110 as well as for sequential fluid flow through multiple regions thereof. Notably, when referring to the porous membrane that includes multiple regions, the porous membrane is a single substrate in that the multiple regions use a common porous membrane. This is made possible because one (or more) of the regions along the substrate can be modified to alter properties of the porous membrane, e.g., decreased hydrophobicity, modified electrostatic properties, or modified hydrogen bonding properties. For example, the multiple regions can include a complexing region 120 that is modified with a releasing compound 162 and further includes an analyte-complexing compound 142 applied thereto that may have a functional group for binding with an analyte introduced with a sample fluid (not shown but shown at FIG. 2). The releasing compound can interact with the porous membrane in a manner so that the complexing region has suitable conditions to receive the analyte-complexing compound and the analyte (for complexing with the analyte-complexing compound), but also provide conditions to allow release of these various complexes while the sample fluid flows through the self-wicking device in the direction toward a detecting region 150. The detecting region, on the other hand, includes the same porous membrane as is present at the complexing region, but in this example, the detecting region is not modified by the releasing compound. The testing compound and the control compound can thus interact with the porous membrane in a manner so that the detecting region has suitable conditions to receive and immobilize testing compound and control compound applied thereto. The testing compound in FIG. 1 is shown as an immobilized testing compound 172 at a first discrete location and the control compound is shown as an immobilized control compound 182 at a second discrete location. In this example, the immobilized control compound is positioned downstream from the immobilized testing compound, but this may or may not be the arrangement in every circumstance. The two discrete regions (or lines in this example) can be elsewhere within the detecting region.

[0015] The term “discrete location” means a discrete location relative to the detecting region, e.g., having a smaller footprint than the detecting region, but does not imply that the immobilized testing compound and the immobilized control compound are necessarily completely discrete relative to one another. For example, the immobilized testing compound may be applied in a line that partially overlaps with the immobilized control compound, e.g., two lines may intersect to form a “plus” pattern.

[0016] The term “immobilized” includes any material that can be applied and will not wick away, at least for long enough for analyte testing to occur, while a sample fluid is passed through the self-wicking device. Thus, the immobilized testing compound or the immobilized control compound may be held in place by covalent attachment, hydrophobic attraction, electrostatic attraction, hydrogen bonding, physical trapping, or the like.

[0017] In further detail, the complexing region 120 in this example can include a releasing compound 162 that is applied on the porous membrane 110. As described, the releasing compound can interact with the porous membrane in a manner so that the complexing region has suitable conditions to receive the analyte-complexing compound and the analyte (for complexing with the analyte- complexing compound), but also provide conditions to allow release or the rate of release of these various complexes while the sample fluid flows through the self-wicking device. More details regarding the releasing compound and application thereof is described hereinafter.

[0018] The complexing region can also be loaded, e.g., adsorbed or impregnated with, an analyte-complexing compound 142 having a functional group that can bind with an analyte in a sample fluid to form an analyte complex. The complexing region can release the analyte-complexing compound having the functional group to bind with the analyte in the sample upon application and movement of a sample fluid therethrough. The analyte- complexing compound identity may vary based on the analyte and the purpose of the self-wicking assay device. In some examples, the analyte-complexing compound can include a colloidal gold, a colored latex particle, a fluorescent latex particle, a paramagnetic latex particle, a cellulose nanobead, a fluorescent tag, or a combination thereof. The analyte-complexing compound (unbound to the analyte, for example) can include a detection moiety that can be detected by the immobilized control compound also in the detecting region, thereby indicating that a sample fluid has passed through the self-wicking assay device. This is one way of determining that the test has run its course if the results are negative, e.g., the immobilized testing compound did not detect an analyte complex.

[0019] Notably, as shown in FIG. 1 , there are two additional regions shown, namely an upstream region 112 and a downstream region 114. These regions may be of a different substrate material joined with the porous membrane or may be of the same substrate material extended into regions 112 and/or 114 beyond what is specifically shown at reference numeral 110. Example functions that may occur at upstream region 112 may include receiving sample fluid to a sample pad, fluid processing prior to reaching the complexing region 120, etc. Example functions that may occur at downstream region 112 may include fluid absorption to an absorbent pad. Notably, there may also be intervening regions between the complexing region and the detecting region 150, such as concentrating regions or other fluid processing regions.

[0020] Referring now to FIG. 2, the same self-wicking assay device 100 is presented as shown in FIG. 1 , including features such as the porous membrane 110, the complexing region 120 including releasing compound 162 and the analyte-complexing compound 142, and the detecting region 150 including the immobilized testing compound 172 and the immobilized control compound 182. However, in this example, the device is shown in the context of a system coupled with sample fluid 130 that contains an analyte 132 to be tested using the self-wicking assay device. As mentioned, the analyte should be capable of binding with the analyte-complexing compound in the complexing region and allowed to pass therethrough to be received at the detecting region positioned downstream relative to the complexing region.

[0021] An example method detecting the presence of an analyte 132 in a sample fluid 130 using the self-wicking device 100 shown in FIG. 2 may be carried out as follows. In one example, a method may include flowing the sample fluid through a complexing region 120 of a porous membrane 110 of the self-wicking assay device. The complexing region in this example can include a releasing compound 162 to provide for release of an analyte-complexing compound (bound or unbound with analyte) from the porous membrane during fluid flow, as well as the analyte-complexing compound 142 to bind with the analyte (or not if the analyte is not present) of the sample fluid to form an analyte complex, shown at 144. The method may further include flowing the sample fluid, including the analyte complex formed in the complexing region, into a detecting region 150 that includes an assay component, which in this case includes a test line in the form of an immobilized testing compound 172 and a control line in the form of an immobilized control compound 182. Because of the nature, e.g., properties, of the porous membrane, the immobilized testing compound and the immobilized control compound can be immobilized at discrete locations within the detection regions.

[0022] The porous membranes in these and other examples can include, for example, nitrocellulose, polyvinylidene fluoride, charge-modified nylon, or polyethersulfone. Other hydrophobic materials can likewise be used in addition to these listed, such as those in which a relatively homogeneous or homogenous porosity can be obtained with pore sizes from about 500 nm to about 10 μm or other subrange of pore sizes set forth herein. The binding mechanism that may assist with immobilizing the testing compound and/or the control compound to form the testing and control lines in the detection region may include electrostatic (ionic), hydrophobic, or covalent, for example. Likewise, some of these same mechanisms can be beneficial in the complexing region when applying the releasing compound thereto. In many examples, the porous membrane can have a hydrophobic character, in that they are made from primarily hydrophobic components, e.g., nitrocellulose. However, if treated with releasing compound, such as anionic surfactant and/or certain polymers, the surfaces can be made to be more hydrophilic, allowing for wetting and wicking away of the sample fluid through the complexing region.

[0023] When the sample fluid is applied to the complexing region, the analyte should be capable of binding with the analyte-complexing compound and allowed to pass therethrough to be received at the detecting region positioned downstream relative to the complexing region, e.g., flowing with the sample fluid flows. For control purposes, unbound analyte-complexing compound can also be released to interact with the control compound, for example.

[0024] Pores of the porous membrane can have an average pore size ranging from about 500 nm to about 10 μm, from about 1 μm to about 10 μm, from about 5 μm to about 10 μm, from about 500 nm to about 5 μm, or from about 2 μm to about 8 μm. A thickness of the porous membrane can range from about 0.1 mm to about 2 mm, from about 0.5 mm to about 1 mm, or from about 0.2 mm to about 0.8 mm. A length of the porous membrane can range from about 1 mm to about 200 mm, from about 1 mm to about 100 mm, from about 1 mm to about 50 mm, from about 1 mm to about 10 mm, from about 5 mm to about 10 mm, from about 1 mm to about 5 mm, or from about 2 mm to about 8 mm. A width of the porous membrane can range from about 1 mm to about 50 mm, from about 1 mm to about 20 mm, from about 5 mm to about 15 mm, or from about 8 mm to about 16 mm.

[0025] The self-wicking assay devices can be positioned in some instances in a housing (not shown) with a fluid flow channel therein. In some examples, the fluid flow channel can include a negative space that can be etched, molded, or engraved from a material of a housing and may surround the porous membrane of a self-wicking assay device. The housing can be a casing that the porous membrane may be disposed within. In some examples, the housing may further include a viewing window over the detecting region. The viewing window may be an opening in the housing or may include an optically transparent material in the area of the detecting region to allow a user to view the results of a self-wicking test.

[0026] The fluid flow channel through the housing can have a channel size perpendicular to fluid flow through the channel that can range from about 5 μm to about 15 mm. In yet other examples, the fluid flow channel can have a size that can range from about 5 μm to about 15 mm, from about 5 μm to about 2 mm, from about 1 mm to about 15 mm, from about 100 μm to about 500 μm, from about 500 μm to about 1 mm, or from about 5 mm to about 10 mm, etc. The fluid flow channel may include a pathway. The pathway may be a linear pathway, a curved path, a pathway with turns, a branched pathway, a serpentine pathway, or any other pathway configuration. In some examples, the pathway may be linear and/or branched. The porous membrane may be present in a portion of or throughout the entire length of the fluid flow channel.

[0027] In some examples, the self-wicking assay device may further include other components. For example, the device may include a fluid input port. The fluid input port may be used to access the complexing region so that a sample fluid, an elution fluid, or a combination thereof may be applied through the fluid flow channel and onto the porous membrane. The self-wicking assay device may also include an elution fluid input port. The elution fluid input port may be positioned at or upstream of the concentrating region. The elution fluid input port may also be used to add other charge-modifying fluids into the self- wicking assay device.

[0028] In some examples, the self-wicking assay device can further include a backing. The backing can support the porous membrane. In some examples, the backing can include polyphenylene ether, polyester, polytetrafluoroethylene, glass, glass fiber, cellulose, nitrocellulose, or a combination thereof. The backing can be used to provide stability to the porous membrane.

[0029] In some examples, the self-wicking assay device can further include a flow controlling agent. The flow controlling agent may be impregnated within the porous membrane and may include buffer salts, proteins, surfactants, and the like. The flow controlling agent may increase or decrease a fluid flow rate of the sample fluid through the porous membrane.

[0030] These specific examples are provided to illustrate how the self- wicking assay device may be used, but it is noted that there are many other similar uses, particularly as these devices relate to different types of lateral flow devices, for example. The self-wicking assay devices can be used for competitive assays, sandwich assays, quantitative assays, or the like. Example sample fluids that may be tested can include urine, blood, saliva, sweat, serum, or other biological samples regardless of origin or application, e.g., environmental, disease, clinical, laboratory, hospitals, physicians, veterinary, chemical analysis, toxins, pregnancy, etc.

[0031] The sample fluid may be a fluid that can include an analyte to be detected or can exclude the analyte to be detected by the self-wicking assay device. When the sample fluid includes the analyte, a positive result may be generated by the device. When the sample fluid excludes the analyte, a negative result may be generated by the device. The analyte in the sample fluid may be selected from amino acids, peptide strands, glycans, polypeptides, antibodies, proteins, or a combination thereof. In one example, the analyte can include a protein. The analyte may be at least ten residues long. A single strand of the analyte may have a weight average molecular weight ranging from about 1 ,500 Daltons to about 250 KD, about 5,000 Daltons to about 200 KD, or from about 50 KD to about 250 KD.

Systems for Manufacturing Self-Wicking Devices

[0032] Systems for manufacturing a self-wicking devices are shown by way of Example in FIGS. 3 and 4. In FIG. 3, the system is shown in the context of an example where various fluids or “agents” can be digitally controlled by ejection from fluidjet printheads, for example. In this example, a releasing agent 160 can be applied to a complexing region 120 of a porous membrane 110. In further detail, a testing agent 170 can be applied to a first discrete location of the detecting region as well as a control agent 180 to a second discrete location of the detecting region. Application (A)-(C) can be in any order. In one example, a first fluidjet ejector 164 can be used to eject the releasing agent including a releasing compound from a first reservoir 166 when loaded therein, a second fluidjet ejector 174 can be used to eject a testing agent 170 including a testing compound from a second reservoir 176 when loaded therein, and a third fluidjet ejector 184 can be used to eject a control agent 180 including a control compound from a third reservoir 186 when loaded therein. The system can also include a hardware controller 190 to generate a command to apply releasing agent onto the complexing region of the porous membrane, apply testing agent to immobilize the testing compound at a first discrete region of the detecting region after application of the releasing agent, and apply control agent to immobilize the control compound at a second discrete region of the detecting region after application of the releasing agent. In another example, the system can include the releasing agent, the testing agent, and the control agent as part of the system components. In one example, the system further can include an additional fluidjet ejector(s) (not shown) to eject a second testing agent including a second testing compound from an additional reservoir when loaded therein, or to eject an analyte-complexing agent including an analyte-complexing compound from an additional reservoir, etc. These additional fluidjet ejector(s) can be referred to herein, arbitrarily as fourth, fifth, sixth, etc., fluid jet ejectors. In another example, the porous membrane can also be included as part of the system components. Notably, the porous membrane in this example is shown as being supported by a platform 102 that can have any configuration suitable for holding the porous membrane in place for application of the various fluid agents.

[0033] As shown in FIG. 4, an alternative system is shown that can include the same features of that shown in FIG. 3, as shown by common reference numerals outlined above, except that in this example, the analyte- complexing compound 146 can likewise be applied using a complexing agent 140 loaded in a complexing agent reservoir 146 and ejected from a complexing agent fluidjet ejector 144. In this instance, since it is the analyte-complexing compound that is being applied, it may be applied to the complexing region rather than the detecting region. Though the analyte-complexing compound is shown as being applied by a fluidjet ejector, it can likewise be applied by other methodologies, such as by the use of a pipette, blotting, soaking, or by pre- loading during manufacture of the porous membrane 110. Methods for Manufacturing Self-Wicking Devices

[0034] A method 300 of manufacturing a self-wicking assay device can include applying 310 releasing agent including a releasing compound to a porous membrane to leave the releasing compound at a complexing region of the porous membrane. The releasing agent in this example is not applied to a separate detecting region that is positioned downstream along the porous membrane relative to the complexing region. The method can further include applying 320 a testing agent including a testing compound to the porous membrane at a first discrete location of the detecting region to provide an immobilized testing compound at the first discrete location, and applying 330 a control agent including a control compound to the porous membrane at a second discrete location of the detecting region to provide an immobilized control compound at the second discrete location. In one example, applying the releasing agent can be via a first fluidjet ejector, applying the testing agent can be via a second fluidjet ejector, applying the control agent can be via a third fluidjet ejector, or a combination thereof. The method can further include applying a complexing agent including an analyte-complexing compound to the complexing region. In another example, the method can include applying a second testing agent including a second testing compound to the releasing compound at a third discrete location of the detecting region to provide a second immobilized testing compound at the third discrete location. The immobilized testing compound can include a molecule to bind an analyte that is complexed with the analyte-complexing compound, the immobilized second testing compound can include a second molecule to bind a second analyte that is complexed with the analyte-complexing compound, and the immobilized control compound can include a third molecule to bind the analyte-complexing compound that is unbound to the analyte, or to some other compound that is not bound to the analyte. Releasing Compounds

[0035] Releasing compounds can be applied to the complexing region from a releasing agent, which can be a fluid that carries and contains the releasing compound. The releasing agent can be, for example, formulated to be reliably ejected from fluidjet ejectors, e.g., thermal or piezo jetting architecture. There are a few different classes of compounds that can be used to modify the complexing region to provide a suitable level of release of the analyte- complexing agent and the analyte bonded to the analyte-complexing agent when a sample fluid is introduced (typically with the analyte) to flow through the self-wicking assay devices described herein. Examples of releasing compounds that may be carried to the complexing region with a releasing agent may include membrane-modifying compounds, solute-interacting compounds, or the like.

[0036] A membrane-modifying compound can be described as a compound (or compounds that interact in situ) which includes chemistry for bonding with the porous membrane. The membrane-modifying compound may also include moieties that are suitable for releasing analyte-bound and/or unbound analyte-complexing compound. In this example, the surface of the porous membrane can be changed to be more hydrophilic, for example, allowing for quicker wetting and resolubilization of compounds that are introduced to this complexing region. As porous membranes of the present disclosure may be hydrophobic in nature, they often include a considerable amount of nonspecific binding that may otherwise hold onto biomolecules when introduced. Once dried, the hydrophobic membranes can be harder to wet, which means that any unbound biomolecules may have difficulty resolubilizing and may not be able to be released from the porous membrane while the sample fluid is being passed therethrough. Treatment with a membrane- modifying compound can provide a decrease in nonspecific binding of biomolecules to the porous membrane, particularly hydrophobic porous membranes.

[0037] As one example, a membrane-modifying compound can be delivered in a fluid and can include a dual functioning copolymer with a modifiable functional portion that serves to block binding of the biomolecule, while another portion may have a stronger affinity to the hydrophobic substrate. The modifiable portion can include functionalities or moieties that can provide steric or electrostatic hindrance to binding, e.g., PEG moieties, primary amines to form amide linkages, etc., that may allow for binding of BSA, casein, or other biomolecules that act as blocking agents. As another example, a Pluronic block copolymer surfactant with terminal alcohols can be used to noncovalently bind to the surface of the hydrophobic membrane, while leaving hydrophilic portions available to change the membrane surface energy. A Pluronic block copolymer may include hydrophilic ethylene oxide (EO) and hydrophobic propylene oxide (PO) blocks arranged block copolymer structure, e.g., A-B-A structure. The terminal alcohols may also be transformed to primary amines/carboxylic acids, which then can attach further blocking agents. Thus, a portion with affinity to the porous substrate may be hydrophobic to enable noncovalent hydrophobic binding. Stated another way, one example of a membrane-modifying compound may be a dual functioning copolymer to non-covalent bind to the porous substrate and block biomolecule binding. The molecular weight of the copolymer can be dependent on the polymer composition, which may be dictated by feed ratio and reactivities of the monomers applied, but in some examples, the molecular weight may range from about 2,000 Daltons to about 1 ,000,000 Daltons, from about 2,000 Daltons to about 500,000 Daltons, or from about 4,000 Daltons to about 500,000 Daltons. In further detail, though hydrophobic non-covalent attraction is mentioned above, other mechanisms of attraction can likewise be used, depending on the porous membrane. In some examples, the burial of hydrophobic surfaces in protic-polar solvents may dominate the interaction, especially with hydrophobic surfaces. Multivalent hydrogen bonding may also aid in association of complimentary donor/acceptor pairs existing between the polymer and surface. In situ polymerization or polymerization followed by attraction or attachment can likewise be used for providing releasing compound at the complexing region. These compounds for polymerization can be applied by fluidjet ejection in some examples. [0038] Polymerizable free-radical or condensation monomers (including some which may act as crosslinking monomers) that can be used to prepare such polymers as releasing compounds may include the following: and/or where R represents a moiety that may provide functionality to the polymer system, with example structures including aliphatic or aromatic chemical groups to impart hydrophobicity; oxygen-, nitrogen-, or sulfur-containing moieties that generate polar bonds and impart hydrophilicity; pH sensitive moieties that form anionic or cationic species; surface bound crosslinking sites; etc. Notably, the surface of the porous membrane may also be chemically modified so as to provide acryloyl, condensable, or other functional groups that may incorporate into the growing polymer chain or initiate polymerization of monomers in solution, thereby providing a covalent link to the membrane surface, depending on the chemistry of the porous substrate being used.

[0039] A solute-interacting compound is another type of releasing compound that may not bind with the porous membrane perse, but rather interacts with various components that may be introduced to the complexing region, e.g., analyte-complexing compound (either bound or bound to the analyte from the sample fluid). The term “solute-interacting compound” in this instances does not infer that components added to the complexing region to interact with the solute-interacting compound are necessarily dissolved in solution, for example, as such “solute" components introduced could be in the form of a fine dispersed compound, for example. The solute-interacting compound may include simple components that protect biomolecules or other components from denaturation during drying steps, e.g., sugars, or may be more complex copolymers with functionalities to enable selective binding and protection. The solute-interacting compound can thus be prepared to treat the porous membrane to protect analyte, analyte-complexing compound, or a combination thereof from drying, provide rapid wetting, and assist with resolubilization and release from the porous membrane upon sample flow.

[0040] An example solute-interacting compound that can be suitable for use includes polymer-based hydrogel particles. These functional copolymer particles can be used to protect the biomolecules during fluidjet dispensing and drying. The polymer-based hydrogel particles can either be dispensed alone, or dispensed with components such as the analyte-complexing compound. If dispensed separately, dispensing may be carried out before dispensing a complexing agent that contains the analyte-complexing compound within a time interval so that both are still undried when they are contacted at the complexing region. Either way, if dispensed together or in series while both remain wet, e.g., applied wet-on-wet, such applications may allow for the various types of compounds, e.g., solute and solute-interacting compound, a chance to more immediately interact without going through a drying and re-wetting step. Notably, the functional groups or moieties of the polymer-based hydrogel particles can dictate the interaction chemistry with solutes or sample fluids, e.g., proteins, colloids, etc.

[0041] In another example, a solute-interacting compound may provide polymer interaction with subsequently applied compounds via a combination of complimentary hydrophobic, ionic, and/or hydrogen bonding interactions. For example, exposed hydrophobic groups of a polymer may associate with hydrophobic moieties of solutes, e.g., analyte, analyte-complexing compound, or a combination thereof, in an entropically driven association, e.g., release of water. Anionic and/or cationic groups of a polymer may associate with their complimentary compounds or solutes to form salt bridges. Hydrogen bonding pairs between the polymer and the solute, e.g., analyte, analyte-complexing compound, or a combination thereof, may also contribute to a paired association. Hydrophilic aprotic groups may in some cases disrupt polymer associations of other applied compounds or solutes.

[0042] In another example, the solute-interacting compound may be able to capture and then release the biomolecules in a controlled manner. Example polymers used in this manner may include those that exhibit temperature-, pH-, salt-, etc., dependent phase change. A release of bound solute may be obtained by triggering a reduction in the density of complimentary interactions.

[0043] In further detail, Pluronic block copolymers can be used, which can form micelles to capture, protect, and release biomolecules. Another type of polymer that can be used may include self-immolative polymers, which can break down upon the introduction of a triggering element. The triggering element may vary depending on the chemistry chosen, e.g., pH, specific small molecule, etc., and may cause a spontaneous depolymerization and release of captured solute, such as the analyte, the analyte-complexing compound, or a combination thereof, etc.

Definitions

[0044] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the content clearly dictates otherwise.

[0045] As used herein, a plurality of items, structural elements, compositional elements, and/or materials may be presented in a common list for convenience. However, these lists should be construed as though individual members of the list are individually identified as a separate and unique member. Thus, no individual member of such list should be construed as a de facto equivalent of any other member of the same list solely based on presentation in a common group without indications to the contrary.

[0046] Concentrations, dimensions, amounts, and other numerical data may be presented herein in a range format. A range format is used merely for convenience and brevity and should be interpreted flexibly to include the numerical values explicitly recited as the limits of the range, and also to include all the individual numerical values or sub-ranges encompassed within that range as if individual numerical values and sub-ranges are explicitly recited. For example, a numeric range that ranges from about 10 to about 500 should be interpreted to include the explicitly recited sub-range of about 10 to about 500 as well as sub-ranges thereof such as about 50 and about 300, as well as sub- ranges such as from about 100 to about 400, from about 150 to about 450, from about 25 to about 250, etc.

[0047] The terms, descriptions, and figures used herein are set forth by way of illustration and are not meant as limitations. Many variations are possible within the disclosure, which is intended to be defined by the following claims and their equivalents in which all terms are meant in the broadest reasonable sense unless otherwise indicated.