DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Status of the Claims
Claims 1-4, 7-11, 13, 14, and 18-21 are pending. Claims 1, 2, 4, 7, 9, 13, 14, 18, and 19 are amended. Claims 5-6, 12, and 15-17 are canceled. Claims 20 and 21 are newly added. Claims 1-4, 7-11, 13, 14, and 18-21 are examined herein.
Priority
The present application, filed 10/12/2023, claims benefit of U.S. Provisional Patent Application 63/379,954, filed 10/18/2022. The benefit is acknowledged and the claims examined herein are treated as having an effective filing date of 10/18/2022.
Maintained Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
The rejections under 35 U.S.C. 112(b) are set forth below in view of Applicant’s amendments.
Claims 1-4, 7-11, 13, 14, and 18-21 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Regarding claim 1, it recites “wherein an aqueous two-phase system (ATPS) comprising cores and shells of a plurality of core-shell microparticles suspended in a non-aqueous liquid, and a bioassay composition results upon mixing with a test sample comprising a bioassay signal.” The limitation renders claim 1 indefinite because the grammatical and structural relationship between the recited aqueous emulsion or suspension, ATPS, bioassay composition, and test sample is unclear. Specifically, the claim recites “an aqueous two-phase system (ATPS) comprising cores and shells of a plurality of core-shell microparticles suspended in a non-aqueous liquid, and a bioassay composition results upon mixing with a test sample,” but does not clearly identify what is mixed with the test sample or what results from the mixing. It is unclear whether the ATPS results upon mixing, whether the bioassay composition results upon mixing, whether both the ATPS and bioassay composition result upon mixing, or whether the previously recited aqueous emulsion or suspension is intended to be the material that is mixed with the test sample to produce one or both thereof. Accordingly, the claim does not establish with reasonable certainty the required relationship among the aqueous emulsion or suspension, ATPS, bioassay composition, and test sample. For purposes of compact prosecution, the limitation will be interpreted to require that the previously recited aqueous emulsion or suspension is mixed with the test sample comprising a bioassay signal, resulting in the recited ATPS comprising cores and shells of a plurality of core-shell microparticles suspended in a non-aqueous liquid and the recited bioassay composition. Appropriate correction is required.
Claim 1 further recites “when the first aqueous phase and the second aqueous phase are at a sufficient to an optimal ratio for a desired bioassay.” The phrase “a sufficient to an optimal ratio for a desired bioassay” renders the claim indefinite because the claim does not provide an objective boundary for determining what constitutes a sufficient ratio, what constitutes an optimal ratio, or what range is encompassed by sufficient to an optimal. Furthermore, for a desired bioassay does not provide an objective standard for determining whether a particular ratio falls within the claimed range. Consequently, the metes and bounds of the claimed ratio cannot be determined with reasonable certainty. For purposes of compact prosecution, a sufficient to an optimal ratio for a desired bioassay will be interpreted broadly as encompassing a ratio of the first aqueous phase to the second aqueous phase that is sufficient to provide the recited intensification of the bioassay signal, including a ratio providing an optimal degree of such intensification for the bioassay being performed. No particular numerical ratio, numerical range, or degree of optimization is read into the claim. Appropriate correction is required.
Claims 2-4, 7–11, 13–14, and 18-21 are rejected under 35 U.S.C. 112(b) as being indefinite because each claim depends directly or indirectly from indefinite claim 1. Appropriate correction is required.
Regarding claim 2, it recites “wherein the first and second aqueous phases exist when the first and second water-soluble polymers are at concentrations above threshold concentrations to achieve the sufficient ratio of the aqueous phases.” The phrase “the sufficient ratio of the aqueous phases” is indefinite because claim 1 does not previously introduce a particular sufficient ratio. Rather, claim 1 recites a sufficient to an optimal ratio for a desired bioassay, which itself does not establish an objective boundary for determining the claimed ratio. It is therefore unclear whether the sufficient ratio in claim 2 refers to a particular lower endpoint of the ratio recited in claim 1, any ratio capable of providing signal intensification, an optimal ratio, or some other unidentified ratio. Additionally, the claim recites that the polymers are present at concentrations above threshold concentrations to achieve the sufficient ratio of the aqueous phases. Because the sufficient ratio itself lacks a reasonably certain boundary, the claim does not establish with reasonable certainty what relationship between the polymer concentrations and the phase ratio satisfies the limitation. For purposes of compact prosecution, the sufficient ratio of the aqueous phases will be interpreted consistently with the broadest reasonable interpretation applied to claim 1 as a ratio of the first aqueous phase to the second aqueous phase sufficient to provide the recited intensification of the bioassay signal. The phrase concentrations above threshold concentrations to achieve the sufficient ratio of the aqueous phases will be interpreted as requiring concentrations of the first and second water-soluble polymers above respective threshold concentrations sufficient to provide the first and second aqueous phases at such a ratio. No particular numerical threshold concentration or numerical phase ratio is read into the claim. Appropriate correction is required.
Regarding claim 4, it recites “an aqueous salt solution” and subsequently recites “the salt exceed threshold concentrations.” There is insufficient antecedent basis for the salt. Claim 4 introduces an aqueous salt solution, but does not separately introduce a salt before subsequently referring to the salt. Thus, the claim does not expressly establish the identity of the salt. It is unclear whether the salt refers to a salt contained in the previously recited aqueous salt solution or another salt. Accordingly, the scope of the limitation is unclear. For purposes of compact prosecution, the salt will be interpreted to mean the salt component of the previously recited aqueous salt solution. Appropriate correction is required.
Regarding claim 8, it recites “wherein the kit is for inclusion with RT-qPCR.” There is insufficient antecedent basis for the kit. Claim 8 depends directly from amended claim 1. As presently amended, claim 1 does not previously introduce or define a kit. Although claim 7 recites a kit for inclusion with PCR, RT-PCR, NAAT, ELISA, NEAR, TMA, LAMP, HDA, CRISPR, or SDA, claim 8 does not depend from claim 7. Accordingly, it is unclear what previously recited kit is intended by the kit in claim 8. For purposes of compact prosecution, the kit will be interpreted to mean a kit included with or associated with the bioassay signal intensifier of claim 1, wherein the kit is for inclusion with RT-qPCR. No additional kit components or configuration are read into the claim. Appropriate correction is required.
Regarding claim 13, it recites “a first water-soluble polymer comprising polyethylene glycol (PEG) and a second water-soluble polymer comprising dextran as the core and the shell of the core-shell microparticles.” The recitation renders claim 13 indefinite because the relationship between the first and second water-soluble polymers and the core and the shell is unclear. Specifically, the grammatical construction a second water-soluble polymer comprising dextran as the core and the shell can reasonably be read as associating the second water-soluble polymer comprising dextran with both the core and the shell. Alternatively, the claim may be intended to require the first water-soluble polymer comprising PEG and the second water-soluble polymer comprising dextran to correspond to respective core and shell phases. However, the claim does not specify which polymer corresponds to which structural region or whether both polymers may be present in both regions. Consequently, one of ordinary skill in the art cannot determine with reasonable certainty whether the claim requires the PEG-containing polymer to form the core and the dextran-containing polymer to form the shell, the dextran-containing polymer to form the core and the PEG-containing polymer to form the shell, the dextran-containing polymer to constitute both core and shell, or another arrangement. For purposes of compact prosecution, claim 13 will be interpreted to require a first water-soluble polymer comprising PEG and a second water-soluble polymer comprising dextran, wherein the polymer-containing aqueous phases constitute the core and the shell of the core-shell microparticles. Because the claim does not specify the correspondence between the respective polymers and the core and shell, no particular PEG-core/dextran-shell or dextran-core/PEG-shell orientation is read into the claim. Appropriate correction is required.
Regarding claim 14, it recites “a first water-soluble polymer and an aqueous salt solution, and subsequently recites solutions of the first water-soluble polymer and the salt separate into the core and the shell of the core-shell microparticles.” There is insufficient antecedent basis for the salt. Claim 14 introduces an aqueous salt solution, but does not separately introduce a salt before subsequently referring to the salt. Thus, the claim does not expressly establish whether the salt refers to the salt component of the previously recited aqueous salt solution or to another salt. For purposes of compact prosecution, the salt is interpreted as the salt component of the previously recited aqueous salt solution. Appropriate correction is required.
Claim 14 further recites “solutions of the first water-soluble polymer and the salt separate into the core and the shell of the core-shell microparticles.” This recitation renders the claim indefinite because the claim does not distinctly establish the relationship between the recited solutions and the subsequently recited core and shell. In particular, the claim does not specify whether the solution of the first water-soluble polymer forms the core and the salt-containing solution forms the shell, whether the salt-containing solution forms the core and the polymer-containing solution forms the shell, or whether the resulting core and shell each comprise differing amounts of the first water-soluble polymer and salt. Accordingly, the required compositional relationship between the recited polymer/salt system and the core and shell cannot be determined with reasonable certainty. For purposes of compact prosecution, solutions of the first water-soluble polymer and the salt separate into the core and the shell is interpreted broadly to require separation of an aqueous system comprising the first water-soluble polymer and the salt into two aqueous phases constituting the core and shell of the core-shell microparticles. Because the claim does not specify which resulting phase constitutes the core and which constitutes the shell, no particular polymer-core/salt-shell or salt-core/polymer-shell configuration is read into the claim. Appropriate correction is required.
Maintained Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or
nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
The rejections under 35 U.S.C. 103 are set forth below in view of Applicant’s amendments. Accordingly, the prior art combination and supporting rationale have been updated to address the amended claim language.
Claims 1-3, 9, 13, and 18-21 are rejected under 35 U.S.C. 103 as being unpatentable over Yasukawa et al. (Monodisperse Water-in-Water-in-Oil Emulsion Droplets. ChemPhysChem. Vol. 12, No. 2, February 2011 – Non-Final dated 04/15/2026) in view of Chen et al. (Enhancing the Sensitivity of DNA and Aptamer Probes in the Dextran/PEG Aqueous Two-Phase System. Analytical Chemistry. Vol. 93, No. 24, June 2021).
Regarding claim 1, for the claimed bioassay signal intensifier comprising an aqueous emulsion or suspension comprising a first aqueous phase and a second aqueous phase, Yasukawa teaches a double emulsion prepared via an aqueous two-phase system, wherein aqueous two-phase systems are generally composed of a water solution of two particular polymers, and, at high concentrations of these components, spontaneous phase separation takes place. One phase is rich in one polymer and the second phase is rich in the other polymer (p. 263). Yasukawa further teaches that mixing two entirely aqueous solutions containing different water-soluble molecules generates a water-in-water (W/W) emulsion, comprising polymer-containing water-solvated droplets dispersed in another aqueous polymer solution (p. 263).
Regarding the claimed ATPS comprising cores and shells of a plurality of core-shell microparticles suspended in a non-aqueous liquid, Yasukawa teaches introducing the two-polymer solution into a continuous organic phase to create a stable emulsion in which phase separation occurs within each droplet, and teaches uniform W/W/O emulsion droplets, in which each drop contained a single internal droplet, creating core-shell structures (p. 264, Fig. 1). Yasukawa specifically introduces PEG/DEX W/W emulsions into a flow of hexadecane and reports monodisperse W/W/O droplets having an outer diameter of about 96 μm and inner aqueous droplets of about 67 μm (p. 264, Fig. 1). Thus, the hexadecane constitutes the claimed non-aqueous liquid. Regarding selective partitioning into one phase of the ATPS, Yasukawa further teaches adding reactive blue 180, which preferably partitions into the PEG-rich phase (p. 263), and recognizes that PEG/DEX ATPSs have established solute partitioning behavior of cells, particles and proteins and that such highly selective partitioning ability should allow effective encapsulation of solutes in the microcapsule (p. 265). These teachings would have directed a skilled artisan to investigate analytical and biological uses that exploit selective partitioning within Yasukawa’s controlled PEG/DEX core-shell compartments.
Regarding control of the claimed first and second aqueous phases at a sufficient-to-optimal ratio, Yasukawa teaches that the concentrations of each polymer in both aqueous phases can be thermodynamically determined and predicted from the phase diagrams of the aqueous two-phase systems, and that the resulting thermodynamic partition equilibrium permits precise control of the interior of the W/W/O emulsion droplet (p. 264). Yasukawa additionally teaches that by changing only the fraction of PEG-rich phase to that of the DEX-rich phase, they could control the size of the internal core, thereby controlling the middle-phase thickness (p. 264, Fig. 2).
However, Yasukawa does not expressly teach using its W/W/O core-shell ATPS as a bioassay signal intensifier wherein a bioassay composition results upon mixing with a test sample and the bioassay signal is intensified by selectively partitioning one or more signaling components into one ATPS phase, or selecting the phase ratio for the purpose of enhancing a desired bioassay signal.
Chen teaches the missing analytical application. Specifically, Chen teaches that increasing the local concentration of DNA-based probes is a convenient way to improve the sensitivity of biosensors, employs a classic aqueous two-phase system (ATPS) comprising polyethylene glycol (PEG) and dextran, and teaches that DNA oligonucleotides are selectively enriched in the dextran-rich phase (Abstract, p. 8577). Chen further teaches that a higher volume ratio of PEG-to-dextran and a higher concentration of PEG also enrich more DNA probes in the dextran-rich phase, with T15 DNA enriched approximately sevenfold at a dextran:PEG volume ratio of 1:10 (Abstract, p. 8577). Regarding a bioassay composition resulting upon mixing with a test sample and signal enhancement through selective partitioning, Chen teaches by simply mixing analytes, DNA probes, and polymers, improved detection could be achieved under optimal ATPS conditions (p. 8577), and experimentally demonstrates a molecular-beacon DNA assay in which the fluorescence intensity increased more in the ATPS and, because DNA was concentrated in the dextran phase, the final fluorescence intensity for the ATPS was around 3.8-fold higher than that in the buffer, while the limit of detection (LOD) in the ATPS improved by 3.6 times (p. 8581, Fig. 4B–C). Regarding the claimed phase ratio, Chen teaches that more DNA accumulated in the dextran phase at higher ratios and concludes that DNA enrichment can be improved by tuning the concentration, molecular weight, and volume of the PEG phase (p. 8580, Fig. 3B–C). Chen further expressly teaches that, although its experiment used a 1:1 phase ratio, by using a higher volume ratio, more enrichment in the dextran can be achieved, and that if the detection takes places in the individual droplets, even higher signal enhancement can be achieved (p. 8581).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify Yasukawa’s PEG/DEX W/W/O core-shell microparticle system according to Chen’s bioassay enrichment technique by incorporating a bioassay sample and signaling components into the PEG/DEX ATPS and selecting the relative aqueous-phase conditions so that the signaling components preferentially partition into one aqueous phase, thereby increasing their local concentration and intensifying the measured bioassay signal. Yasukawa itself provides the motivation to seek such a modification by identifying selective partitioning of biological solutes as a useful property of PEG/DEX ATPS and by teaching that such selective partitioning permits effective encapsulation, while Chen demonstrates the specific analytical advantage sought—selective enrichment of signaling DNA probes increases fluorescence and improves detection sensitivity—and further teaches that detection in individual droplets can provide even greater signal enhancement. A skilled artisan would have had a reasonable expectation of success because Yasukawa and Chen employ the same PEG/DEX ATPS partitioning principle; Yasukawa experimentally demonstrates controllable PEG/DEX core-shell droplets and selective solute partitioning, while Chen experimentally demonstrates that DNA signaling probes selectively partition in PEG/DEX ATPS, retain target-recognition/signaling functionality, and produce increased fluorescence and improved detection limits. Thus, the proposed modification uses the selective-partitioning property already recognized by Yasukawa for the bioassay signal-enhancement purpose expressly demonstrated by Chen, with the references themselves providing both the motivation and a reasonable expectation of success.
Regarding claim 2, for the additional limitation that the aqueous emulsion or suspension comprises a first water-soluble polymer and a second water-soluble polymer, as discussed above, Yasukawa teaches PEG and DEX as the two polymers of its aqueous two-phase system and prepares W/W emulsions from aqueous PEG and DEX solutions (pp. 263). Regarding wherein the first and second aqueous phases exist when the first and second water-soluble polymers are at concentrations above threshold concentrations to achieve the sufficient ratio of the aqueous phases, Yasukawa teaches that the two-polymer aqueous system spontaneously separates at sufficiently high polymer concentrations and that the concentrations of each polymer in both aqueous phases can be thermodynamically determined and predicted from the phase diagrams of the aqueous two-phase systems (p. 264). Chen further teaches a concentration-dependent PEG/DEX phase diagram, observing that phase separation indicative of the formation of the ATPS occurred only when the PEG molecular weight was 8000 or higher and that mixing different PEG and dextran concentrations produced a concentration-dependent contour line separating homogeneous and phase-separated regions (p. 8579, Fig. 1B). Chen additionally teaches that polymer concentration and phase-volume ratio affect selective DNA enrichment and therefore the resulting bioassay signal.
Therefore, it would have been prima facie obvious to use PEG/DEX concentrations above their phase-separation thresholds and adjust those concentrations/relative phase amounts to obtain a phase ratio sufficient for Chen’s desired probe enrichment and signal enhancement, because Chen expressly identifies polymer concentration and phase-volume ratio as variables governing enrichment, while Yasukawa teaches predictable determination and control of the corresponding PEG/DEX phase composition. A skilled artisan would reasonably have expected success because both references experimentally demonstrate PEG/DEX phase separation under controlled polymer concentrations and Chen demonstrates the resulting effect of those variables on probe enrichment and signal enhancement.
Regarding claim 3, as discussed above, Yasukawa expressly teaches polyethylene glycol (PEG) and dextran (DEX) as the polymers forming its aqueous two-phase system (pp. 263-265).
Regarding claim 9, for a method of performing a bioassay, as discussed above, Chen performs fluorescence-based DNA, Hg²⁺, and ATP assays using PEG/DEX ATPS to improve detection sensitivity (pp. 8577-8578). Regarding providing a bioassay composition comprising reagents for the bioassay, Chen’s molecular-beacon assay provides the target analyte, molecular-beacon signaling probe, polymers, and assay medium, with Scheme 1 depicting the assay components and ATPS enrichment process (Scheme 1, p. 8577-8578). Regarding adding a signal intensifier according to claim 1 or substituting the signal intensifier for a reagent or solvent of the bioassay composition, Chen teaches obtaining enhanced detection by simply mixing analytes, DNA probes, and polymers in the PEG/DEX ATPS (p. 8578), while Yasukawa supplies the W/W/O core-shell form of that PEG/DEX ATPS. Regarding to partition an intensified bioassay signal within one of a core or a shell of a plurality of core-shell microparticles in a non-aqueous liquid, Chen teaches selective accumulation of the signaling probe and analyte in the dextran-rich phase and enhanced fluorescence therein, while Yasukawa teaches that its dextran-rich aqueous phase constitutes the inner phase/core of W/W/O droplets suspended in hexadecane (Fig. 1, p. 264). Yasukawa’s Figure 1 identifies W(inner): DEX-rich phase, W(middle): PEG-rich phase, and the outer oil phase as hexadecane/Span 80 (Fig. 1, p. 264). Regarding measuring the intensified bioassay signal, Chen measures fluorescence and demonstrates increased fluorescence and decreased detection limits in its ATPS assays (p. 8582).
Therefore, it would have been prima facie obvious to perform Chen’s bioassay enrichment and fluorescence-measurement method using Yasukawa’s controlled PEG/DEX W/W/O core-shell droplets because Chen expressly teaches the analytical advantage of concentrating signaling probes in an ATPS phase and further identifies individual-droplet detection as capable of providing greater signal enhancement, while Yasukawa provides stable, controllable individual PEG/DEX core-shell droplets. A skilled artisan would reasonably have expected success because Chen experimentally demonstrates probe partitioning and enhanced fluorescence in PEG/DEX droplets and Yasukawa experimentally demonstrates the same PEG/DEX phase chemistry in stable core-shell W/W/O droplets.
Regarding claim 13, interpreted broadly to require PEG- and dextran-containing aqueous phases constituting the core and shell without requiring a particular orientation, Yasukawa directly teaches this limitation and, more specifically, teaches a DEX-rich core and PEG-rich shell. Figure 1 expressly identifies W(inner): DEX-rich phase and W(middle): PEG-rich phase (Fig. 1, p. 264), and Yasukawa subsequently refers to the resultant DEX core as isolated from the oil/water interface by the PEG-rich phase (p. 265). The experimental section confirms use of aqueous PEG and DEX solutions in the W/W/O droplets (p. 265).
Regarding claim 18, Yasukawa teaches a DEX-rich phase as the inner/core aqueous phase (Fig. 1, p. 264), while Chen teaches preferential enrichment of DNA signaling probes in the dextran-rich phase, producing enhanced fluorescence and improved sensitivity (pp. 8579–8581). Therefore, applying Chen’s selective DEX-phase probe enrichment to Yasukawa’s W/W/O droplets would have predictably placed the enriched signaling components, and thus the intensified signal, in Yasukawa’s DEX-rich core. A skilled artisan would reasonably have expected this result because Yasukawa experimentally establishes a W/W/O architecture having a DEX-rich inner/core aqueous phase, while Chen experimentally demonstrates that DNA signaling probes preferentially partition into the DEX-rich phase and retain their signaling functionality with enhanced fluorescence and improved sensitivity.
Regarding claim 19, Yasukawa teaches varying the relative fractions of the PEG-rich and DEX-rich aqueous phases to control the size of the internal core and the thickness of the middle phase, and Fig. 2 depicts resulting W/W/O core-shell droplets having different relative core and shell proportions, including a configuration (Fig. 2(a)) in which the inner core occupies a smaller volume than the surrounding shell (Fig. 2(a), p. 264). Yasukawa further teaches that the volume and the thickness of the middle phase, as well as the volume and the size of inner core can be controlled by controlling the relative phase composition (p. 264).
Regarding claim 20, Yasukawa expressly identifies W(inner): DEX-rich phase in Figure 1 (p. 264) and describes the resultant DEX core (p. 265).
Regarding claim 21, Yasukawa likewise expressly teaches the DEX-rich inner/core phase of its PEG/DEX W/W/O core-shell droplets (Fig. 1, pp. 264–265).
Claims 4 and 14 are rejected under 35 U.S.C. 103 as being unpatentable over Yasukawa et al. and Chen et al., as applied to claims 1 and 9 above, and further in view of Tan et al. (One-Step Fabrication of pH-Responsive Microcapsules with Aqueous Cargo Using Aqueous Two-Phase System. Colloids and Surfaces. A, Physicochemical and Engineering Aspects, vol. 650, No. 129506, October 5 2022).
With respect to the teachings of Yasukawa and Chen, see the discussion above, which applies equally here. These references differ from the instant claims in failing to expressly teach or specify that the aqueous emulsion or suspension comprises a first water-soluble polymer and an aqueous salt solution and the polymer and salt exceed threshold concentrations to form the two aqueous phases (claim 4), and the claimed polymer/salt core-shell alternative (claim 14).
Regarding claim 4, Tan supplies that limitation. Regarding wherein the aqueous emulsion or suspension comprises a first water-soluble polymer and an aqueous salt solution, Tan teaches an ATPS comprising an aqueous PEGDA/acrylic-acid polymer solution and ammonium sulfate. Tan explains that two immiscible aqueous phases are obtained by adding salts to a homogeneous aqueous polymer phase above a critical concentration and specifically employs an PEGDA/acrylic acid (AA)/(NH₄)₂SO₄ system (Introduction, p. 2). Regarding wherein the first and second aqueous phases exist when the first water-soluble polymer and the salt exceed threshold concentrations, Tan systematically varies ammonium-sulfate concentration and reports that the homogeneous solution separates into two immiscible aqueous layers at sufficient salt concentration, producing a salt-rich upper solution and an AP-rich bottom aqueous solution (§3.1, p. 3). Tan’s experimental preparation likewise states that ammonium sulfate was added to induce phase separation to the AA/PEGDA solutions (§2.2, p. 2). Regarding the compatibility of this polymer/salt system with the claimed/Yasukawa architecture, Tan uses the resulting salt-rich and AP-rich phases as the inner and middle phases, respectively, with mineral oil as the continuous outer phase to generate W/W/O double-emulsion droplets (§2.4, pp. 3; Fig. 1(a)–(b), p. 4). The figure on page 4 expressly depicts the salt-rich inner phase, AP-rich middle phase, and mineral-oil outer phase.
Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to further modify the Yasukawa/Chen bioassay signal-intensifier system by employing Tan’s polymer/salt ATPS as an alternative aqueous phase-forming system, using concentrations sufficient to induce separation into polymer-rich and salt-rich aqueous phases and arranging those phases as the inner and middle aqueous compartments of the W/W/O droplets. The motivation is supplied by Tan’s demonstration that the polymer/salt ATPS produces the same technically relevant W/W/O aqueous compartmentalization used by Yasukawa, while Chen establishes the analytical advantage of exploiting selective ATPS partitioning to concentrate signaling components and improve bioassay signal and detection sensitivity. The modification therefore provides a known alternative ATPS chemistry while preserving the core-shell aqueous compartmentalization needed to exploit Chen’s signal-enrichment mechanism. A skilled artisan would have had a reasonable expectation of success because Tan experimentally demonstrates both the concentration-induced polymer/salt phase separation and formation of stable W/W/O droplets from the resulting aqueous phases, while Chen demonstrates that selective ATPS partitioning can be used analytically to increase local signaling-component concentration.
Regarding claim 14, refer to the discussion above. It would have been prima facie obvious to further modify the Yasukawa/Chen system according to Tan by using Tan’s phase-separated polymer-rich and salt-rich aqueous solutions as the respective aqueous core/shell compartments because Tan expressly demonstrates that those phases are compatible with W/W/O droplet formation, thereby providing a polymer/salt alternative to Yasukawa’s polymer/polymer ATPS architecture while preserving the compartmentalization used for Chen’s selective signal enrichment. A skilled artisan would reasonably have expected success because Tan experimentally forms the claimed inner/middle aqueous configuration in mineral oil, and Chen establishes the analytical utility of concentrating signaling components through selective ATPS partitioning.
Claims 7-8 and 10-11 are rejected under 35 U.S.C. 103 as being unpatentable over Yasukawa et al. and Chen et al., as applied to claims 1 and 9 above, and further in view of Vindeirinho et al. (SARS-CoV-2 Diagnostics Based on Nucleic Acids Amplification: From Fundamental Concepts to Applications and Beyond. Frontiers in Cellular and Infection Microbiology. Vol. 12, March 2022 – Non-Final dated 04/15/2026).
With respect to the teachings of Yasukawa and Chen, see the discussion above, which applies equally here. These references differ from the instant claims in failing to expressly teach or specify the particular claimed PCR, RT-PCR, NAAT, ELISA, NEAR, TMA, LAMP, HDA, CRISPR, SDA, RT-qPCR kit implementations (claims 7, 8, 10, and 11).
Regarding claim 7, wherein the bioassay composition comprises a kit for inclusion with PCR, RT-PCR, NAAT, ELISA, NEAR, TMA, LAMP, HDA, CRISPR, or SDA, Vindeirinho teaches nucleic-acid amplification diagnostic platforms including PCR/RT-qPCR and isothermal amplification technologies. For example, Vindeirinho identifies RT-LAMP and TMA as common isothermal amplification methods and further describes RT-HDA, real-time fluorescence tracking, with fluorescence-producing oligonucleotide probes, and commercial products including Lucira CHECK-IT COVID-19 Test Kit, Mobile Detect Bio BCC19 Kit, and SHERLOCK CRISPR SARS-CoV-2 Kit (p. 8; 17-20; Tables 8-9).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date to further modify the Yasukawa/Chen signal-intensifier system for use with the known nucleic-acid amplification kits taught by Vindeirinho because Chen’s signal-enhancement mechanism concentrates DNA-based signaling probes in an ATPS phase, while Vindeirinho’s nucleic-acid amplification assays use fluorescence-producing oligonucleotide probes as detection components. The references therefore provide a direct technical reason to apply Chen’s local probe-enrichment technique to Vindeirinho’s fluorescence-based nucleic-acid assays—to increase the local concentration of the signaling probe and thereby improve the measurable detection signal. A skilled artisan would reasonably have expected success because Chen experimentally demonstrates that oligonucleotide probes retain recognition and signaling functionality after PEG/DEX partitioning and produce enhanced fluorescence, while Vindeirinho establishes that fluorescent oligonucleotide probes are conventional signaling components of the recited amplification assays.
Regarding claim 8, wherein the kit is for inclusion with RT-qPCR, Vindeirinho expressly discusses commercial RT-qPCR kit technology, including a commercial multi-target RT-qPCR kit by Thermo Fisher Scientific and a RT-qPCR commercial kit produced by Roche (p. 4). Vindeirinho’s PCR-related diagnostic table further identifies RT-qPCR assays using fluorescence and oligonucleotide probes (Table 8, p. 18). Accordingly, the Yasukawa/Chen/Vindeirinho combination renders claim 8 obvious for the reasons stated for claim 7.
Regarding claims 10 and 11, refer to the discussion above. Vindeirinho teaches PCR/RT-qPCR and LAMP diagnostic compositions and explains that nucleic-acid amplification assays use primers/probes and fluorescence-based detection. Therefore, it would have been prima facie obvious to employ the conventional PCR/RT-qPCR or LAMP kit components taught by Vindeirinho in the Yasukawa/Chen signal-enhancement system because those assays employ nucleic-acid signaling components of the type Chen demonstrates can be selectively concentrated to enhance fluorescence and detection sensitivity. A skilled artisan would reasonably have expected success because Chen demonstrates preserved probe recognition and increased signal after ATPS partitioning, and Vindeirinho establishes those fluorescent oligonucleotide components as established detection components in PCR/RT-qPCR or LAMP diagnostics.
Response to Arguments
Applicant’s amendments and arguments, filed 07/14/2026, have been fully considered. The response is set forth below.
Rejection under 35 U.S.C. 112(b)
Applicant argues that claims 4, 13, and 14 have been amended to overcome the previous rejection under 35 U.S.C. 112(b), and claim 12 has been canceled. Applicant requests withdrawal of the rejection. Applicant’s arguments concerning the previously stated grounds of rejection are persuasive. In particular, the previous rejection found claims 4 and 14 indefinite because they recited the salt phase without sufficient antecedent basis; found claims 12 and 14 indefinite based upon threshold concentrations of a binodal curve; and rejected claim 13 because claim 13 depended from indefinite claim 12. Applicant has removed the previously rejected the salt phase language from claims 4 and 14, removed the threshold concentrations of a binodal curve language relevant to the prior rejection, canceled claim 12, and amended claim 13 to depend from claim 9 rather than canceled claim 12. Accordingly, the specific §112(b) grounds set forth in the previous Office action against claims 4, 12, 13, and 14 are withdrawn.
However, upon further consideration, a new ground(s) of rejection is made in view of the presently amended claim language and the additional indefiniteness issues identified in this Office action. Notwithstanding withdrawal of the previous grounds, Applicant’s assertion that amended claims 4, 13, and 14 are now definite is not persuasive as to the grounds stated in this Office action. For claim 4, Applicant changed the salt phase to the salt. Claim 4 presently introduces an aqueous salt solution and subsequently recites the salt exceed threshold concentrations. As explained in the present §112(b) rejection, the claim does not expressly introduce a particular salt before referring to the salt. Thus, although the previous the salt phase antecedent-basis rejection is withdrawn, claim 4 remains rejected for the reasons stated in the present Office action. Claim 13 no longer depends from canceled claim 12, and the previous inherited rejection is therefore withdrawn. However, amended claim 13 now recites a first water-soluble polymer comprising polyethylene glycol (PEG) and a second water-soluble polymer comprising dextran as the core and the shell of the core-shell microparticles. As explained in the present rejection, this wording does not clearly specify the structural correspondence between the first and second polymers and the recited core and shell. Similarly, amended claim 14 no longer recites the salt phase or threshold concentrations of a binodal curve, and those previous grounds are withdrawn. Claim 14 now recites a first water-soluble polymer and an aqueous salt solution, followed by solutions of the first water-soluble polymer and the salt separate into the core and the shell. The presently stated rejection therefore concerns the presently recited language—particularly the relationship of the salt to the previously recited aqueous salt solution and the unclear correspondence between the resulting aqueous phases and the claimed core and shell.
Accordingly, Applicant’s amendments overcome the specific §112(b) grounds previously stated, but claims 4, 13, and 14 remain subject to the §112(b) grounds set forth in this Office action.
Rejections under 35 U.S.C. 103
Applicant’s arguments concerning the previous §103 rejections have been fully considered. In view of the amendments and upon further consideration of the claimed subject matter, the previous §103 rejections based upon Cheung in view of Mashayekhi, alone or further in view of Kamei, Wijethunga, Atefi, and/or Yasukawa, are withdrawn and replaced by the grounds of rejection set forth in this Office action. Applicant’s arguments directed specifically to Cheung, Mashayekhi, Kamei, Wijethunga, and Atefi do not overcome the present grounds because those references are no longer relied upon for the teachings challenged by Applicant. Arguments concerning Yasukawa remain relevant because Yasukawa is applied in the present rejection and are addressed separately below.
Applicant argues that Cheung in view of Mashayekhi fails to teach or suggest amended claims 1 and 9 because the claimed signal intensifier allegedly exists as a suspension of one aqueous phase in a second aqueous phase before inclusion of the sample, whereas the ATPS of Cheung and Mashayekhi is generated by addition of surfactant to the sample. Applicant further argues that neither reference teaches concentrations providing a 1:1 phase-volume ratio or teaches modifying phase volume by changing surfactant concentration, and that neither teaches a core-shell particle suspended in a non-aqueous liquid. Applicant’s arguments with respect to claim(s) 1 and 9 have been considered but are moot because the new ground of rejection does not rely on any reference applied in the prior rejection of record for any teaching or matter specifically challenged in the argument. The present rejection instead relies principally on Yasukawa for the claimed physical W/W/O core-shell architecture and Chen for the bioassay signal-enrichment functionality.
Applicant’s core-shell/non-aqueous-liquid argument therefore does not identify a deficiency in the present rejection. Yasukawa expressly teaches introducing the PEG/DEX two-polymer aqueous system into a continuous organic phase, producing uniform W/W/O emulsion droplets, in which each drop contained a single internal droplet, creating core-shell structures. Yasukawa further identifies the outer non-aqueous phase as hexadecane containing Span 80. Thus, unlike the previous Cheung/Mashayekhi ground challenged by Applicant, the present rejection expressly accounts for the claimed core-shell microparticles suspended in a non-aqueous liquid.
Applicant’s argument concerning the failure of Cheung and Mashayekhi to identify a 1:1 volume ratio likewise does not overcome the present rejection. First, amended claim 1 does not require a 1:1 ratio. Claim 1 requires the phases to be at a sufficient to an optimal ratio for a desired bioassay. Under the compact-prosecution interpretation set forth in the §112(b) rejection, this encompasses a phase ratio sufficient to provide the recited signal intensification and does not import a 1:1 numerical ratio from the specification. Second, Chen directly investigates phase-volume ratio as a parameter affecting enrichment of signaling DNA probes and teaches that increasing the relative PEG-phase volume increases enrichment in the dextran-rich phase. Chen further teaches that detection in individual droplets can produce even higher signal enhancement. Yasukawa correspondingly teaches controlling the relative fractions of its PEG-rich and DEX-rich aqueous phases and thereby controlling the volume and size of the internal core and the thickness of the surrounding phase. The present rejection therefore does not depend upon Cheung’s or Mashayekhi’s surfactant concentration teachings to satisfy the phase-ratio limitation.
Applicant’s argument that the previous art forms its ATPS only upon mixing with the sample likewise does not establish patentability over the present combination. Claim 1, as presently drafted, itself recites that an ATPS and bioassay composition results upon mixing with a test sample. To the extent Applicant intends to require a fully formed core-shell suspension to exist before contact with the sample, that temporal sequence is not stated with such clarity in the claim and is not imported from the specification. The present rejection applies the claim according to the compact-prosecution interpretation stated under §112(b). Accordingly, Applicant’s arguments directed to Cheung and Mashayekhi do not overcome the presently stated rejection of claims 1 and 9 over Yasukawa in view of Chen.
For claims 2-4 and 13-14, Applicant argues that Kamei and Wijethunga fail to cure the deficiencies asserted against Cheung and Mashayekhi because Kamei and Wijethunga teach two aqueous polymers or an aqueous polymer/salt system but do not teach a core-shell structure in a non-aqueous liquid. Applicant’s arguments with respect to claim(s) 2–4, 13, and 14 have been considered but are moot because the new ground of rejection does not rely on any reference applied in the prior rejection of record for any teaching or matter specifically challenged in the argument. The present rejection does not rely upon Kamei or Wijethunga. For claims 2, 3, and 13, Yasukawa itself supplies the PEG/DEX polymer/polymer core-shell architecture suspended in a non-aqueous phase, while Chen supplies the bioassay signal-enrichment teaching. Yasukawa expressly identifies W(inner): DEX-rich phase and W(middle): PEG-rich phase in Fig. 1 and describes the resultant DEX core. Thus, the presently applied art directly addresses the core-shell limitation Applicant identified as absent from Kamei and Wijethunga.
For claims 4 and 14, the present rejection additionally relies on Tan rather than Kamei or Wijethunga. Tan teaches an aqueous polymer/salt ATPS comprising an AA/PEGDA polymer-containing phase and ammonium sulfate, teaches salt-induced separation into two aqueous phases, and then uses the resulting salt-rich and AP-rich phases as the inner and middle phases, respectively, of W/W/O double-emulsion droplets in mineral oil. Accordingly, the precise core-shell/non-aqueous-liquid feature that Applicant argues was absent from Kamei and Wijethunga is expressly accounted for in the presently stated combination. Applicant’s arguments therefore do not overcome the grounds of rejection of claims 2–4, 13, and 14.
Applicant canceled claims 5–6, 12, and 15–17. Accordingly, the previous rejections of these claims are withdrawn as moot, and no further substantive response concerning their patentability is necessary.
Applicant argues that Yasukawa does not cure the deficiencies of Cheung and Mashayekhi because Yasukawa concerns preparation of monodisperse W/W/O droplets primarily for microcapsule formation and is allegedly not directed to an assay, analytical device, or analytical use. Applicant additionally asserts, with respect to new claims 20 and 21, that Yasukawa teaches that the core is always PEG rich and not the dextran phase. Applicant’s arguments have been fully considered but they are not persuasive. Applicant’s characterization of Yasukawa does not account for Yasukawa’s express disclosure of the DEX-rich inner/core phase, its selective-partitioning teachings, or the combined teachings of Yasukawa and Chen, as explained below.
Applicant is correct to the limited extent that Yasukawa does not itself expressly perform the bioassays presently recited. The rejection, however, does not rely upon Yasukawa alone for that teaching. Chen supplies the bioassay signal-enrichment functionality. In response to applicant’s arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). Yasukawa is relied upon for the PEG/DEX W/W/O physical architecture and the known selective-partitioning capability of that architecture. Yasukawa teaches that PEG/DEX ATPSs had been developed for separation of macromolecules, cells, and organelles and discusses solute partitioning of cells, particles, and proteins. Yasukawa further concludes that such highly selective partitioning ability should allow effective encapsulation of solutes in the microcapsule. Chen supplies the analytical use that Applicant contends is absent from Yasukawa. Chen expressly teaches that increasing the local concentration of DNA probes improves biosensor sensitivity; experimentally demonstrates selective enrichment of DNA probes in the dextran-rich phase of a PEG/DEX ATPS; applies that enrichment to fluorescent molecular-beacon detection; obtains increased fluorescence and improved detection limits; and expressly teaches that detection in individual droplets can yield still greater signal enhancement. Thus, the rejection does not require Yasukawa itself to teach every feature of the claimed bioassay.
Yasukawa supplies a controlled individual-droplet PEG/DEX core-shell architecture possessing the selective-partitioning behavior, while Chen demonstrates why a skilled artisan would exploit that same PEG/DEX partitioning behavior for bioassay signal enhancement. Yasukawa identifies selective biological-solute partitioning as useful and expressly contemplates effective encapsulation resulting from that partitioning, while Chen demonstrates the particular analytical benefit of using PEG/DEX selective partitioning to concentrate signaling probes and enhance fluorescence and sensitivity. Chen’s express statement concerning greater signal enhancement through individual-droplet detection provides an additional technical reason to employ its assay-enrichment principle in Yasukawa’s controlled individual droplets. Accordingly, Yasukawa’s focus on W/W/O droplets and microcapsules does not defeat the obviousness rejection when the combined teachings of Yasukawa and Chen are considered.
Applicant’s assertion that Yasukawa teaches that the core is always PEG rich and not the dextran phase is inconsistent with Yasukawa’s express disclosure. Yasukawa Figure 1 expressly identifies: W(inner): DEX-rich phase and W(middle): PEG-rich phase (Fig. 1, p. 264). Yasukawa further explains that the resultant DEX core is isolated from the oil/water interface by the surrounding PEG-rich phase (p. 265). Thus, in the disclosed W/W/O architecture, the inner/core aqueous phase is DEX-rich and the surrounding middle/shell aqueous phase is PEG-rich—not the reverse. This teaching directly addresses new claims 20 and 21, which require that the core comprises dextran. Yasukawa expressly discloses that configuration. Applicant’s argument is therefore not persuasive.
Claim 18 requires the aqueous phase containing the intensified signal to be the core, and new claim 20 further requires the core to comprise dextran. Yasukawa establishes that its core is the DEX-rich aqueous phase. Chen establishes that DNA signaling probes preferentially enrich in the dextran-rich phase and that this enrichment increases fluorescence and improves detection sensitivity. Applying Chen’s selective DEX-phase signaling-probe enrichment to Yasukawa’s DEX-rich core therefore predictably places the enriched signaling component—and consequently the intensified signal—in the core. Applicant’s characterization of Yasukawa as lacking an assay does not rebut this combination because Chen, rather than Yasukawa, supplies the bioassay signal-enrichment teaching.
Applicant amended claim 19 to require wherein the core has a smaller volume than the shell. Yasukawa expressly teaches varying the relative fractions of the PEG-rich and DEX-rich aqueous phases to control the size of the internal core and the thickness of the middle phase. Yasukawa’s Figure 2 depicts resulting W/W/O core-shell droplets having different relative core and shell proportions, including a configuration (Fig. 2a) in which the inner core occupies a smaller volume than the surrounding shell (Fig. 2a, p. 264). Yasukawa further explains that the volume and the thickness of the middle phase, as well as the volume and the size of inner core can be controlled through the relative phase composition (p. 264). Accordingly, the amendment to claim 19 does not distinguish the claim from the presently applied Yasukawa/Chen combination.
Claim 21 depends from claim 1 and requires wherein the core comprises dextran. As explained above, Yasukawa expressly identifies the inner phase as DEX-rich in Figure 1 and expressly describes the resulting structure as having a DEX core. The additional limitation of claim 21 is therefore directly taught by Yasukawa.
Applicant concludes that all pending claims are in condition for allowance. That conclusion is not persuasive for the reasons stated above and in the rejections set forth in this Office action. The amendments overcome certain grounds of the previous Office action, but the presently pending claims remain subject to the §112(b) and §103 grounds stated herein. The previous §103 rejections relying upon Cheung, Mashayekhi, Kamei, Wijethunga, and Atefi are withdrawn. The previous §112(b) grounds concerning the salt phase, threshold concentrations of a binodal curve, and claim 13’s dependency from canceled claim 12 are likewise withdrawn. Withdrawal of those particular grounds does not constitute allowance because the presently amended claims have been reconsidered and are rejected on the grounds fully set forth in this Office action. Claims 1–4, 7–11, 13–14, and 18–21 therefore remain rejected for the reasons stated above.
Conclusion
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/E.O./Examiner, Art Unit 1677
/BAO-THUY L NGUYEN/Supervisory Patent Examiner, Art Unit 1677
September 24, 2026
