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 .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on February 26, 2026 has been entered.
Status of Claims
Claims 1, 3-6, and 8-12 were pending. Claims 1 and 10 are amended.
Claims 1, 3-6, and 8-12 are examined herein.
Withdrawn Rejections
The objection to claim 1 is withdrawn in view of claim amendments.
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.
Claims 1, 3-6, and 8-12 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.
Claim 1 recites “(b) conjugating an enzyme to the target analytes bound to the probes”. The term “conjugating” in this context is confusing and misleading, because in the art of sandwich assays this term is used to indicate forming covalent bonds between assay components other than the target analytes.
For the purpose of compact prosecution and applying prior art under 35 U.S.C. 102 and 103, “conjugating” will be interpreted as “binding”.
Claim 1 recites “accumulation of the insoluble colorimetric material inside the hydrogel particles to amplify a colorimetric signal for labeling the target analytes”. The meaning of “amplify” is unclear. The specification fails to disclose any method suitable for amplifying the colorimetric signal. Accumulation of the insoluble colorimetric material cannot amplify the colorimetric signal because it is the insoluble colorimetric material that generates the colorimetric signal. Amplification of the colorimetric signal will not be given any patentable weight.
Claim 1 recites “the probes are bound to and loaded into the polymer network”. It is unclear if binding of the probes and loading them into the polymer network is one or two separate processes. If the probes are already bound to the polymer network, how can they be loaded into the polymer network?
Claim 8 recites “wherein the step of binding the enzyme comprises”. There is insufficient antecedent basis for this limitation in the claim, because claim 1 does not recite enzyme binding.
Claim 11 recites the substrate is selected from bromochloroindolyl phosphate (BCIP)/nitroblue tetrazolium (NBT), 3,3'-diaminobenzidine (DAB), 3,3',5,5'-tetramethylbenzidine (TMB), 4-chloronaphthol, 3,3'diaminobenzidine (DAB), 3-amino-9-ethylcarbazole (AEC), 6-chloro-3-indolyl-o-D galactopyranoside (Red-gal), and combinations thereof. Parent claim 1 recites a single enzyme. It is unclear how combinations of different substrates can be used with a single enzyme. For example, TMB is a substrate of a peroxidase and Red-gal is a substrate of b-galactosidase. The peroxidase cannot accept Red-gal substrate and b-galactosidase cannot accept TMB substrate.
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS. —Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
The following is a quotation of pre-AIA 35 U.S.C. 112, fourth paragraph:
Subject to the following paragraph [i.e., the fifth paragraph of pre-AIA 35 U.S.C. 112], a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claim 10 is rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends.
Claim 1 recites insoluble colorimetric material, while its dependent claim 10 recites luciferase. This enzyme does not produce an insoluble colorimetric material because instead it generates a detectable luminescent emission. Thus, claim 10 fails to include the limitation of insoluble colorimetric material of claim 1.
Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
Claim Rejections - 35 USC § 103
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 set forth in Graham v. John Deere Co., 383 U.S. 1, 148 USPQ 459 (1966), that are applied for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
Determining the scope and contents of the prior art.
Ascertaining the differences between the prior art and the claims at issue.
Resolving the level of ordinary skill in the pertinent art.
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.
Claims 1, 3, 5, 8-12 are rejected under 35 U.S.C. 103 as being unpatentable over Appleyard et al. (IDS; Nat Protoc. 2011 Oct 20; 6(11):1761-74) in view of Gerion et al. (PGPub 2008/0213814), for reasons of record which are reiterated herein below.
Regarding claim 1, Appleyard teaches assays using bar-coded hydrogel microparticles for protein detection (Title) comprising:
reacting a sample containing target analytes with hydrogel particles loaded with probes specifically binding to the target analytes. Specifically, Appleyard teaches mixing particles and unknown sample (pg. 1769, step 34). The hydrogel particles are synthesized using PEG precursors and “[b]iological entities can be mixed directly into the PEG precursor solutions, allowing 3D covalent incorporation into the resulting network” (pg. 1762, col. 2, par. 2), meeting the hydrogel particles loaded with probes (biological entities);
conjugating a label to the target analytes bound to the probes (“conjugating” is interpreted as “binding”, see 112(b) rejection above). Specifically, Appleyard teaches “(ii) a 3–4-h assay in which protein targets are captured and labeled within particles using an antibody sandwich technique” (Abstract). The reference teaches formation of a sandwich between a capture antibody bound to the particles, a target, a reporter antibody, and a fluorescent label - “assay involves assembling an antibody sandwich around the target protein by exposing particles to the sample, adding a biotinylated reporter antibody, and labeling the bound reporters with a streptavidin-phycoerythrin (SAPE) complex (Fig. 2 and pg. 1764, col. 2, par. 4). Phycoerythrin is a fluorescent label.
The reference teaches using cross-linking agent (PEG DA) in the synthesis of the hydrogel particles (pg. 1763, col. 2, par. 3). Appleyard does not specifically teach three-dimensionally crosslinked polymer network, but since the synthesis of the hydrogel particles produces “porous, 3D gel matrix” (pg. 1762, col. 2, par. 2), one would expect the polymer network to have three-dimensionally crosslinked polymer chains.
Appleyard does not specifically teach an enzyme used to produce an insoluble colorimetric material.
Regarding claim 1, Gerion teaches a surface plasmon resonance detection method in sandwich immunoassays (Abstract, Fig. 1-2). Gerion also teaches that sandwich assays can use enzymes for analyte detection ([0053]) among many other methods ([0052]), specifically, enzymes “that convert a soluble compound into an insoluble compound” ([0054]).
It would have been obvious to one having ordinary skill in the art before the effective filing date of the claimed invention to have modified the sandwich assay method of Appleyard by employing colorimetric detection as taught by Gerion in order to provide detection of the target analytes, as an obvious matter of simple substitution of one known element (colorimetric detection) for another (fluorescent detection) to obtain predictable results.
One having ordinary skill in the art would have had a reasonable expectation of success in combining the prior art references because both Appleyard and Gerion teach sandwich format immunoassays, which are known in the art for accepting wide variety of different labels (Gerion [0052]). The substitution of the colorimetric detection for fluorescent detection is routine and the results would have been predictable.
Appleyard and Gerion do not specifically teach a three-dimensionally crosslinked polymer network that restricts diffusion of the insoluble colorimetric material, and the insoluble colorimetric material is generated and immobilized inside the hydrogel particles such that outward diffusion of the insoluble colorimetric material is suppressed by the polymer network, thereby causing accumulation of the insoluble colorimetric material inside the hydrogel particles. However, the references teach all necessary components of the method: hydrogel particles with capture antibodies immobilized to the particles throughout the particles volume (Appleyard, Fig. 2), formation of the sandwich between the capture antibodies, the targets (Fig. 2a), and the reporter antibodies (Fig. 2b). The reporter antibodies can be conjugated to reporter enzymes, such as, alkaline phosphatase and horseradish peroxidase (Gerion, [0054]), which can convert their substrates into an insoluble colorimetric material. When all the assay elements are present, the accumulation of the insoluble colorimetric material necessarily follows from the method and its mechanism.
Regarding claim 3, Appleyard teaches “[b]iological entities can be mixed directly into the PEG precursor solutions, allowing 3D covalent incorporation into the resulting network” (pg. 1762, col. 2, par. 2), meeting the limitation of claim 3 reciting the probes are loaded during
synthesis of the hydrogel particles. The biological entities of Appleyard are the probes specifically binding to the target analytes of the instant invention.
Regarding claim 5, Appleyard teaches the probes are antibodies. Specifically, Appleyard teaches capture antibodies (pg. 1762, col. 2, par. 2 and Fig. 2a).
Regarding claim 8, Appleyard in view of Gerion teaches a reporter antibody (Appleyard, Fig. 2a) as a secondary binding material specifically binding to the target analytes and adding the enzyme for binding to the secondary binding materials. The reporter antibody of Appleyard is bound to a fluorescent label and Gerion teaches that alkaline phosphatase and horseradish peroxidase can be used instead ([0054]).
Regarding claim 9, Appleyard teaches the secondary binding materials are antibodies. Specifically, Appleyard teaches reporter antibody that binds to separate epitopes of the target (Fig. 2b).
Regarding claim 10, Appleyard in view of Gerion teaches the enzyme is alkaline phosphatase or horseradish peroxidase ([0054]).
Regarding claim 11, Appleyard in view of Gerion teaches the substrate is 3,3',5,5'-tetramethylbenzidine (TMB), 4-chloronaphthol, or 3,3'-diaminobenzidine (DAB) ([0055]).
Regarding claim 12, Appleyard teaches the target analyte is protein, specifically, IL-2 (pg. 1766, col. 1, par. 3, “Reagents”).
Claims 4 and 6 are rejected under 35 U.S.C. 103 as being unpatentable over Appleyard in view of Gerion, as applied to claims 1 and 3 above, and further in view of Hermanson (Chapter 15, Editor(s): Greg T. Hermanson, Bioconjugate Techniques (Third Edition), Academic Press, 2013, Pages 589-740), for reasons of record which are reiterated herein below.
The teachings of Appleyard and Gerion have been set forth above. Additionally, Appleyard teaches incorporation of the probes can be performed during the hydrogel particles synthesis using acrylate-modified reactive species (pg. 1762, col. 1, par. 1) and post-synthesis, by using attachment of biotinylated capture antibodies or the direct attachment to amine or carboxyl groups (pg. 1764, col. 2, par. 1).
Appleyard and Gerion fail to teach the probes loaded after synthesis of the hydrogel particles comprises a capture portion specifically binding to the corresponding target analyte and a functional group connected to the capture portion and bonded to an unreacted end in the form of a carbon-carbon double bond connected to the polymer network.
Regarding claim 4, Hermanson teaches various methods for immobilization of ligands on chromatography supports (Chapter 15, Title). Chromatography support of Hermanson is equivalent to the hydrogel particles of the instant invention. Hermanson teaches that the immobilized ligands can be used for “analyti-cal separations involving the assay of a targeted mol-ecule” (pg. 589. Col. 1, par. 2 and Fig. 15.1), which is analogous art for the instant disclosure.
Chapter 15 teaches large variety of coupling methods. Specifically, section 2.2 teaches immobilization of thiols (pg. 670, col. 1). Thiol groups can be generated on the surface of antibodies using a reducing agent (Fig. 15.67(A)). Antibodies with thiol groups can be coupled to maleimide-activated support (Fig. 15.69).
The antibody of Hermanson comprises a capture portion specifically binding to the corresponding target analyte, where the functional group connected to the capture portion is the thiol group, which bonds with unreacted end in the form of a carbon-carbon double bond (the double bond in maleimide moiety) connected to the polymer network.
It would have been obvious to one having ordinary skill in the art before the effective filing date of the claimed invention to have modified the assay method of Appleyard and Gerion by employing coupling chemistry as taught by Hermanson in order to load the probes after synthesis of the hydrogel particles, as an obvious matter of simple substitution of one known element maleimide-activated linker) for another (attachment of biotinylated capture antibodies or the direct attachment to amine or carboxyl groups of Appleyard (pg. 1764, col. 2, par. 1) to obtain predictable results.
One having ordinary skill in the art would have had a reasonable expectation of success in combining the prior art references because Gerion is generic on specific chemistry for coupling the reporter enzyme to the reporter antibody and Hermanson teaches coupling chemistry approaches successfully practiced in the art for many years, so the results of the substitution would have been predictable.
Regarding claim 6, Hermanson teaches the thiol groups generated on the surface of antibodies using a reducing agent (Fig. 15.67(A)), meeting the limitation of claim 6 reciting the functional groups are thiol groups.
Response to Arguments
Applicant's arguments filed February 26, 2026 have been fully considered.
Applicant argues that “The Examiner cites Appleyard as teaching hydrogel particles, Gerion as teaching generation of an insoluble colorimetric product and Hermanson as teaching methods for immobilization of ligands on chromatography supports. However, none of the references-alone or in combination-teaches or suggests a diffusion-restricting polymer network that traps the insoluble product inside the particle to achieve internal accumulation and amplification” (pg. 7, par. 4-5).
The argument is not persuasive because the "diffusion-restricting polymer network that traps the insoluble product inside the particle" is an inherent property of the hydrogel particles taught by Appleyard. The diffusion-restricting property of porous particles is well-known and routinely used in size exclusion chromatography where smaller molecules migrate deeper inside the particles, while penetration of larger molecules is limited by reduced diffusion caused by the polymer network. As such, the insoluble product taught by Gerion gets trapped inside the particles because of the reduced diffusion and accumulates there.
Amplification of the insoluble product is not known in the art and applicant fails to disclose the nature of this amplification (see 112(b) rejection above).
Conclusion
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Alexander Volkov whose telephone number is (571) 272-1899. The examiner can normally be reached M-F 9:00AM-5:00PM (EST).
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Bao-Thuy Nguyen can be reached on (571) 272-0824. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/ALEXANDER ALEXANDROVIC VOLKOV/Examiner, Art Unit 1677
/REBECCA M GIERE/Primary Examiner, Art Unit 1677