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 .
Claims 1-41 and 43-50 are pending.
Applicant’s election without traverse of the species of:
A reporting enzyme;
Protein A/G;
DAB;
Human; and
A surface of a cell
in the reply filed on 11/10/2025 is re-acknowledged. Applicant indicates in the response filed 11/10/2025 that claims 1-2, 4, 6, 25-29, 35, 37, 42-46, and 48-49 encompass the elected species.
Claim 3, 5, 7-24, 30-34, 36, 38-41, 47, and 50 remain withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected species, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 11/10/2025.
Claim 42 has been cancelled.
Claims 1-2, 4, 6, 25-29, 35, 37, 43-46, and 48-49 remain under examination on the merits.
Priority
No claim of priority has been made.
Withdrawn Objections/Rejections
The rejections of claims 1-2, 4, 6, 35, 37, 42-46, and 48-49 under 35 USC §112(a) and of claims 1-2, 4, 6, 35, 37, 42-46, and 48-49 §112(b) are withdrawn as corrected by the 04/24/2026 claim amendments removing the recited ‘variants thereof’ language. The rejections of the claims under 35 USC §103 as presented in the previous office action are withdrawn and replaced with the rejections of the claims under 35 USC §103 as presented in this Office Action to better account for the new and different claim scope resulting from the 04/24/2026 claim amendments.
Maintained-Claim Objections
Claim 28 is objected to for its dependence from rejected base claim 27, but would be allowable if rewritten in independent form.
Maintained-Claim Rejections - 35 USC § 112
35 U.S.C. 112(a)
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA 35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 25-27 and 29 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA 35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention.
The purpose of the written description requirement is to ensure that the inventor had possession, at the time the invention was made, of the specific subject matter claimed. To satisfy the written description requirement, a patent specification must describe the claimed invention in sufficient detail that one skilled in the art can reasonably conclude that the inventor had possession of the claimed invention. See, e.g., Moba, B. V. v. Dianwnd Automation, Inc., 325 F.3d 1306, 1319, 66 USPQ2d 1429, 1438 (Fed. Cir. 2003); Vas-Cath, Inc. v. Mahurkar, 935 F.2d at 1563, 19 USPQ2d at 1116.
Independent claim 1 requires incubating with a fusion comprising an immunoglobulin-binding peptide selected from Protein A/G or variants thereof. Likewise, claims 25-27 recite a Protein A/G having 80%, 90%, or 95% identity, respectively, to SEQ ID NO: 4. Additionally, claim 29 generically recites a Protein A/G without disclosed/required structure.
The specification only teaches 1 example of a Protein A immunoglobulin-binding peptide (instant SEQ ID NO: 1; see for example, pages 16-17 of the instant specification). The specification only teaches 1 example of a Protein G immunoglobulin-binding peptide (instant SEQ ID NO: 2; see for example, pages 17-18 of the instant specification). The specification only teaches 1 example of a Protein A/G immunoglobulin-binding peptide (instant SEQ ID NO: 4; see for example, pages 19-20 of the instant specification). Therefore, there is no disclosure of a representative number of species of a recombinant protein A/G, or a protein A/G having comprising a sequence having less than 100% identity to SEQ ID NO: 4 (all of which are considered to be variants of a protein A/G having 100% identity to SEQ ID NO: 4). No conserved structure is indicated as required to function in the method as claimed (there is no structure-function correlation). Therefore, in view of this disclosure, Applicant is claiming a broad genus of variant immunoglobulin-binding peptides without a representative number of species of said genus or a clearly demonstrated structure/function correlation. The specification does not provide adequate written description for the entire claimed genus of variant immunoglobulin-binding peptides encompassed by the claims because, in the absence of empirical determination, one skilled in the art would be unable to immediately envision, recognize, or distinguish at least most of the members comprised within the genus claimed, specifically, which mutants/variants encompassed by the genus would function as claimed in the method(s).
The written description requirement for a claimed genus may be satisfied through sufficient description of a representative number of species by actual reduction to practice, reduction to drawings, or by disclosure of relevant, identifying characteristics, i.e., structure or other physical and/or chemical properties, by functional characteristics coupled with a known or disclosed correlation between function and structure, or by a combination of such identifying characteristics, sufficient to show the applicant was in possession of the claimed genus. See Eli Lilly, 119 F.3d at 1568, 43 USPQ2d at 1406. A “representative number of species” means that the species which are adequately described are representative of the entire genus. Thus, when there is substantial variation within the genus, one must describe a sufficient variety of species to reflect the variation within the genus. Applicant has not disclosed a representative number of specific species or a conserved/consensus structure responsible for the functions required by the instant method claims. Thus, given the substantial structure variation within the genus of variant immunoglobulin-binding peptides as well as the high level of unpredictability in the art, the disclosure of 1 species is not sufficiently representative of the entire genus. The state of the art discloses Protein A/G immunoglobulin-binding peptides, but does not presently provide a sufficient number of species so as to provide the requisite degree of description for the recitation of variant immunoglobulin-binding peptides as instantly claimed.
Although screening techniques can be used to isolate variant immunoglobulin-binding peptides that possess the ability to function as claimed, Applicant is reminded that the written description requirement of 35 U.S.C. 112 is severable from the enablement provision. As stated in Vas-Cath Inc. v. Mahurkar (CA FC) 19 USPQ2d 1111, 935 F2d 1555, “The purpose of the 'written description' requirement is broader than to merely explain how to 'make and use'; the applicant must also convey with reasonable clarity to those skilled in the art that, as of the filing date sought, he or she was in possession of the invention. The invention is, for purposes of the 'written description' inquiry, whatever is now claimed.”
Applicant is further directed to In re Alonso (545 F.3d 1015 (Fed. Cir. 2008), which involved claims that were directed to methods of using antibodies wherein the court found that the claims lacked adequate written description for the recited genus of antibodies recited in the methods. (C) See p. 8, 3rd paragraph, where Applicant argues that the claims recite all essential features of the invention. Therefore, products used in methods are rightfully subject to the written description requirement.
Regarding the state of the art, Listov et al (Opportunities and challenges in design and optimization of protein function. Nat Rev Mol Cell Biol 25, 639–653 (2024)) teach that the primary amino acid sequence determines downstream structure (protein folding), which then determines function (presenting both the inverse folding problem and the inverse function problem (see for example, Figure 1 and its caption; see also Mishra et al (Inaccurate secondary structure predictions often indicate protein fold switching. Protein Sci. 2019 Aug;28(8):1487-1493. doi: 10.1002/pro.3664. Epub 2019 Jun 17)). Expanding on these problems in proteomics, Reardon (Nature 635, 246-248 (2024)) explains that the goal of designing a protein with known and predictable function, binding partners, size, location, and other traits is, for the moment, a dream. Reardon teaches that further challenges in protein design include predicting how a protein, even if it binds to target, will function upon said binding. Reardon teaches that the primary structure (amino acid sequence) of a protein is critical to function, noting that even proteins of similar shape do not execute the same functions, while those with different shapes may carry out the same tasks. Reardon goes on to teach that it is not always apparent which parts of the primary sequence are important; a seemingly useless amino-acid chain on the side of an enzyme, for instance, might affect how tightly a protein can bind to other molecules or its ability to flip between conformational states. Moreover, Reardon explains that when researchers attempt to solve the structure of a protein experimentally, they often end up seeing only the most stable conformation, which is not necessarily the form the protein takes when it is active (see for example, pages 246-247 of Reardon).
Therefore, the genus of claimed immunoglobulin-binding peptide variants represented/recited by claims 25-27 and 29 are insufficiently described through a demonstrated correlation of a conserved/identifying structure with the claimed function(s) or through a representative number of species. Therefore, claims 25-27 and 29 are deemed to fail to meet the written description requirement, as presently drafted.
Newly Necessitated-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.
Claim(s) 1, 29, and 43-46, and 48-49 is/are rejected under 35 U.S.C. 103 as being unpatentable over Al-Adhami et al (Vet Parasitol. 2014 Feb 24;200(1-2):66-73. doi: 10.1016/j.vetpar.2013.11.004. Epub 2013 Dec 1) in view of Binch et al (JOR Spine. 2020 Jun 11;3(3):e1098. doi: 10.1002/jsp2.1098) and Peuscher et al (J Immunoassay Immunochem. 2014;35(3):322-34. doi:10.1080/15321819.2013.864974).
Regarding claims 1 and 45, Al-Adhami et al teach a new multi-host species indirect ELISA using protein A/G conjugate for detection of anti-Toxoplasma gondii IgG antibodies where microplate wells (Costar, Corning Inc., USA) were coated with 100 μl of T. gondii soluble antigen (1 mg/l). Test sera, controls and reference sera were diluted and 100 μl from each dilution was added to the wells and incubated for 1 h at 37 °C (held to read upon the first incubation step of instant claim 1 where the sera samples read upon the instantly recited first antibody and the T. gondii antigen reads upon the target moiety). After incubation, the wells were washed three times, and 100 μl of HRP-protein A/G conjugate [held to read upon the instantly recited fusion polypeptide comprising (i) a Protein A/G and (ii) a reporting species](or species-specific HRP-conjugated secondary antibody for ELISA-IgG), diluted 1:40,000 in TBS containing 1.5% BSA and 0.05% Tween-20 was added to each well. The plate was incubated for 1 h at 37 °C, then the wells were washed three times with TBS, and 100 μl of 3,3′,5,5′-tetramethylbenzidine (TMB) substrate solution was added and incubated in the dark for 20 min at room temperature (RT). The reaction was stopped by adding 50 μl of 2N H2SO4. The absorbance was read at 450 nm using a microplate reader (Spectramax plus, Molecular Devices, USA) with the aid of ProMax5 software for data recording and analysis [held to read upon the steps of visualizing the reporting species and detecting based on the location of the reporting species, the target moiety] (see for example, the Title and abstract at page 66 and section 2.10. Protocol for indirect ELISA at pages 68-69).
The difference between Al-Adhami et al’s disclosure and that of the instant claims is that Al-Adhami et al don’t teach that the antigen is contained in a mammalian species tissue section (sample) which is incubated with the primary antibody (sera comprising a first antibody against the target moiety/antigen).
However, Binch et al teach that immunohistochemistry (IHC) is commonly used on fresh frozen and paraffin embedded tissues to elucidate the expression of antigens. Binch et al discuss the principles of IHC applied to formalin fixed paraffin embedded (FFPE) intervertebral disc (IVD) tissue and supply optimized protocols (see for example, the abstract at page 1). Binch et al generally teach that IHC includes indirect ELISA (see for example, Figure 1 and its caption at page 2). Binch et al explain that FFPE enables long term storage of samples and improved morphological maintenance during sectioning of particularly large tissue samples (see column 2 of page 1 bridging column 1 of page 2). Binch et al demonstrate successful imaging/visualization of a cell-surface expressed antigen using IHC of FFPE tissues in a canine and human sample (see for example Figures 2 A and F and their respective captions at page 3).
Al Adhami et al and Binch et al do not teach that the primary antibody and the sample are from the same mammalian species.
However, Peuscher et al teach an in vitro immunofluorescence-based indirect IHC protocol for primary antibodies on homologous tissues that is four times more sensitive
than direct IHC staining and requires a much shorter exposure time. The method of Peuscher et al overcomes the background staining problem caused by the co-detection of endogenous immunoglobulins when using secondary anti-human antibodies to detect a human recombinant antibody (human primary antibody) binding to their targets in the context of human tissue samples. This approach will be useful for the detection of antigens on any tissue sample using a primary antibody from the same species (see for example, The paragraph under the Results heading at page 327 and the paragraph under the Conclusion heading at page 333).
It would have been prima facie obvious to the person of ordinary skill in the art to arrive at the claimed invention from the disclosures of Al-Adhami et al, Binch et al, and Peuscher et al. The artisan would have been motivated to make and use the invention as claimed because Al-Adhami et al teach that this assay has multiple advantages over some of the commercially available serological assays as it can be readily adapted for testing samples from various domestic and wild mammalian species for which commercial assays or specific conjugates are unavailable. This indirect ELISA-A/G using tachyzoite antigen of T. gondii and HRP-protein A/G as a conjugate was developed and its performance compared with those of other serological assays for toxoplasmosis. The ELISA-A/G was designed for screening serum samples from multiple mammalian species for T. gondii IgG antibodies using the same conjugate regardless of host species. Likewise, the artisan would have understood that indirect ELISA may be performed successfully on FFPE mammalian tissue to visualize/detect/localize cell-surface expressed antigens in a mammalian sample. The artisan would have found it obvious to use either an isolated antigen or an antigen in an FFPE mammalian tissue sample for analysis using a primary antibody and the Protein A/G-HRP fusion of Al-Adhami et al in order to streamline the ELISA so that it is more readily adapted to different species as taught by Al-Adhami et al. Al-Adhami et al teach that the Protein-A/G-HRP conjugate acts as a non-species-specific conjugate when used as a secondary agent in indirect ELISA (see for example, the abstract at page 66). The assay was shown to be effective and specific in detecting T. gondii antibodies in serum samples from experimentally infected pigs, cats, mice and seals (see for example paragraph 1 of the discussion at column 1 of page 71 and column 2 of page 72). The artisan would therefore understand that the secondary conjugate of Al-Adhami would be expected to work with a variety of primary antibodies from a multiplicity of species (including human primary IgG antibodies) in the commonplace ELISA formats of either Al-Adhami et al or Binch et al using a known human primary antibody applied to human tissue as taught by Peuscher et al. The artisan would have viewed using the method and secondary HRP-Protein A/G conjugate of Al-Adhami et al to examine human tissue using a human primary antibody (such as the H10 human anti-CEA antibody of Peuscher et al; see for example, Figure 3 and its caption at page 331) as obvious, yielding predictable results given that both Al-Adhami et al and Peuscher et al teach IHC methods with a species non-specific secondary agent for use across species and allowing for use of a primary antibody and sample derived from the same species (see MPEP §2143(I)(B) and (G)). Absent a convincing, evidenced showing to the contrary, the artisan would have had a reasonable expectation of success based on the cumulative disclosures of these prior art references.
Regarding claim 29, it is noted that while no special definition is provided for the term ‘recombinant’ in the instant specification, the specification does provide description of the term at page 19, providing that “Protein A/G is a recombinant fusion protein that combines IgG binding domains of both Protein A and Protein G. Protein A/G contains four Fc binding domains from Protein A and two from Protein G, yielding a final mass of approximately 50 kDa,” (see for example, lines 25-27). This would support that a Protein A/G conjugate is recombinant as claimed irrespective of whether said conjugate is produced from a single nucleotide or is chemically induced through conjugation of the protein fragments post-translation. Therefore, Al-Adhami et al’s Protein A/G conjugate must be recombinant within the scope of the instant claims, meeting and making obvious the limitation added by instant claim 29 for reasons discussed above.
Regarding claims 42-44 and 48-49, while Al-Adhami et al teach an assay using a Protein A/G-reporter construct with sera samples from pigs, cats, mice and seals, they do not explicitly teach that the antigen comes from a biological sample.
However, Binch et al teach that use of indirect ELISA to visualize a cell-surface expressed (reading on claim 49) antigen (reading on claim 48) in a FFPE (reading on claim 43) mammalian (dog or human; reading on claims 42-44) sample is known and understood to be successful in the art, as discussed above (see for example Figures 2 A and F and their respective captions at page 3).
It would have been prima facie obvious to the person of ordinary skill in the art to arrive at the claimed invention from the disclosures of Al-Adhami et al and Binch et al. The artisan would have been motivated to make and use the invention as claimed because Al-Adhami et al teach that this assay has multiple advantages over some of the commercially available serological assays as it can be readily adapted for testing samples from various domestic and wild mammalian species for which commercial assays or specific conjugates are unavailable. Likewise, the artisan would have understood that indirect ELISA may be performed successfully on FFPE mammalian tissue to visualize/detect/localize cell-surface expressed antigens in a mammalian sample. The artisan would have found it obvious to use either an isolated antigen or an antigen in an FFPE mammalian tissue sample for analysis using a primary antibody and the Protein A/G-HRP fusion of Al-Adhami et al in order to streamline the ELISA so that it is more readily adapted to different species as taught by Al-Adhami et al. The artisan would have had a reasonable expectation of success based on the cumulative disclosures of these prior art references.
Regarding claim 46, as noted above, the protein conjugate and methods of Al-Adhami et al as modified by Binch et al are intended for and demonstrated as effective for use with multiple species. Al-Adhami et al teach that their method and conjugate were successful used for analysis on murine tissue (see for example, Table 1 and its caption and section 2.6 at page 68, section 3.2 at page 70, and pages 71-72).
It would have been prima facie obvious to the person of ordinary skill in the art to arrive at the claimed invention from the disclosures of Al-Adhami et al and Binch et al. The artisan would have been motivated to make and use the invention as claimed because Al-Adhami et al teach that this assay was successfully used to analyze murine sera samples. The artisan would have understood that indirect ELISA may be performed successfully on FFPE mammalian tissue, such as murine tissue, to visualize/detect/localize cell-surface expressed antigens in a mammalian sample. The artisan would have found it obvious to use either an isolated antigen or an antigen in an FFPE mammalian tissue sample for analysis using a primary antibody and the Protein A/G-HRP fusion of Al-Adhami et al in order to streamline the ELISA so that it is more readily adapted to different species as taught by Al-Adhami et al. The artisan would have had a reasonable expectation of success based on the cumulative disclosures of these prior art references.
Claim(s) 2, 4, 6, and 35 is/are rejected under 35 U.S.C. 103 as being unpatentable over Al-Adhami et al, Binch et al, and Peuscher et al, as applied to claims 1, 29, and 43-46, and 48-49 above, as evidenced by AAT Bioquest (Why is TMB used in conjunction with HRP in ELISA?, obtained from : https://www.aatbio.com/resources/faq-frequently-asked-questions/Why-is-TMB-used-in-conjunction-with-HRP-in-ELISA (posted online: 04/24/2020).
Regarding claim 2, Al-Adhami et al, Binch et al, and Peuscher et al teach the method of claim 1, as discussed above, where the reporter species is horseradish peroxidase (HRP).
Al-Adhami et al, Binch et al, and Peuscher et al do not explicitly state that HRP is an enzyme.
However, HRP is known in the art to be an enzyme (see for example, the 2 page AAT Bioquest reference evidencing that HRP (performing as an enzyme) reacts with TMB substrate producing an observable reaction which is stopped by an acid solution).
Regarding claim 4, Al-Adhami et al teach that prior to detection, the wells were washed three times with TBS, and 100 μl of 3,3′,5,5′-tetramethylbenzidine (TMB) substrate solution was added and incubated in the dark for 20 min at RT (see for example, column 1 of page 69).
TMB is known in the art to be a common substrate for HRP (see for example, the 2 page AAT Bioquest reference teaching that HRP reacts with TMB substrate producing an observable reaction which is stopped by an acid solution).
Regarding claim 6, Al-Adhami et al teach that, prior to detection and after the TMB substrate is incubated with the Protein/A/G-HRP bound to the first antibody bounds to the target moiety, the reaction is stopped by adding 50 μl of 2N H2SO4 (see for example, column 1 of page 69). The artisan would have understood this step to recite stopping the reaction between TMB and HRP as 2N H2SO4 is a common stop solution (sulfuric acid) for said reaction in the art (see for example, the 2 page AAT Bioquest reference teaching that HRP reacts with TMB substrate producing an observable reaction which is stopped by an acid solution such as sulfuric acid).
Regarding claim 35, as discussed above, Al-Adhami et al teach a protein A/G-reporter conjugate where the reporter is HRP, which is an enzyme (see for example, the teachings cited above).
Claims 2, 4, 6, and 35 are obvious for the same reasons that claim 1 is obvious, as discussed above.
Claim(s) 25-26 is/are rejected under 35 U.S.C. 103 as being unpatentable over Al-Adhami et al, Binch et al, and Peuscher et al as applied to claims 1, 29, and 43-46, and 48-49 above, in further view of Dalian University of Technology (CN102676562A, hereinafter referred to as ‘Dalian’; noting that the English Machine Translation of the claims and description obtained from Patent Scope are being relied upon).
As discussed above, Al-Adhami et al and Binch et al teach the method of instant claim 1.
The combined references do not teach a Protein A/G conjugate having 80%, 90%, 95%, or 100% identity to instant SEQ ID NO: 4.
However, Dalian teaches a Protein A/G fusion having a sequence comprising 96.8% identity to 89% of the instant SEQ ID NO: 4 (calculated to have 488 residues identical to the 540 residues of SEQ ID NO: 4, where both sequences begin with Met, such that the prior art sequence is deemed to be 90.37% identical to the instant SEQ ID NO: 4 in its totality; see the sequence alignment provided below; see for example, claim 2 reciting SEQ ID NO: 6 of Dalian).
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It would have been prima facie obvious to the person of ordinary skill in the art to arrive at the claimed invention from the disclosures of the combined references. The artisan would have been motivated to make and use the invention as claimed because where the method of Al-Adhami et al, Binch et al, and Peuscher et al make obvious the method of instant claim 1, as discussed above the artisan would have found it obvious to look for Protein A/G conjugates known in the art for use in said method. The artisan would have then found the Protein A/G fusion of Dalian which Dalian teach has broad-spectrum adsorption capacity for an antibody and an application thereof in the fields of antibody purification and blood purification (see for example, page /11 of the English translation of the description of Dalian). The artisan would have presumed any Protein A/G conjugated to a reporter would function in the method of Al-Adhami et al as modified by Binch et al in light of the degree of generality used when Al-Adhami et al discuss the Protein A/G fusion. The artisan would have had a reasonable expectation of success based on the cumulative disclosures of these prior art references.
Claim(s) 37 is/are rejected under 35 U.S.C. 103 as being unpatentable over Al-Adhami et al, Binch et al and Peuscher et al as evidenced by AAT Bioquest, as applied to claims 2, 4, 6, and 35 above, in further view of Rodig, SJ (Detecting Horseradish Peroxidase-Labeled Cells. Cold Spring Harb Protoc. 2019 Apr 1;2019(4). doi: 10.1101/pdb.prot099713; hereinafter called “Rodig”).
Regarding claim 37, as discussed above, Al-Adhami et al as evidenced by AAT Bioquest, teach the limitations of instant claim 35.
Al-Adhami et al and Binch et al as evidenced by AAT Bioquest do not explicitly teach that the substrate of the reporter enzyme is DAB.
However, Rodig teaches that a range of substrates is available for detection of cells labeled with horseradish peroxidase (HRP). Diaminobenzidine (DAB) is the most commonly used substrate and one of the most sensitive (see for example, the abstract at page 1).
It would have been prima facie obvious to the person of ordinary skill in the art to arrive at the claimed invention from the disclosures of the combined references. The artisan would have been motivated to make and use the invention as claimed because Al-Adhami et al teach that this assay has multiple advantages over some of the commercially available serological assays as it can be readily adapted for testing samples from various domestic and wild mammalian species for which commercial assays or specific conjugates are unavailable. This indirect ELISA-A/G using tachyzoite antigen of T. gondii and HRP-protein A/G as a conjugate was developed and its performance compared with those of other serological assays for toxoplasmosis. The ELISA-A/G was designed for screening serum samples from multiple mammalian species for T. gondii IgG antibodies using the same conjugate regardless of host species. The assay was shown to be effective and specific in detecting T. gondii antibodies in serum samples from experimentally infected pigs, cats, mice and seals (see for example paragraph 1 of the discussion at column 1 of page 71 and column 2 of page 72). The artisan would have had a reasonable expectation of success using DAB as the substrate of HRP based on the cumulative disclosures of these prior art references, such as the teaching that DAB is a common and sensitive substrate in the art for HRP.
Applicant’s Argument and Responses:Applicant argues for withdrawal of the rejections of the claims under 35 USC §112(a) in light of their removal of “variants thereof” language from the claims. However, claims 25-27 and 29 were specifically mentioned as problematic, representing variants in light of their percent identity language (claims 25-27) and lack of recited/required structure (claim 29) in the maintained rejection under 35 USC §112(a) as presented in the previous office action. Therefore, the problematic variants have not been substantively removed from claims 25-27 and 29, which accordingly remain rejected under 35 USC §112(a).
The rejections under 35 USC §103 as previously presented have been withdrawn and replaced with the rejections presented in this Office Action to account for the new and different claim scope resulting from Applicant’s 04/24/2026 claim amendments. Therefore, the only argument presented in Applicant’s 04/24/2026 remarks which remains pertinent is Applicant’s argument for withdrawal of the rejections over Al-Adhami, Binch, AAT Bioquest, and Peuscher (see page 5 of the 04/24/2026 remarks). Applicant alleges that Peuscher et al do not teach incubation of a sample with a primary/capture antibody of the same species as the sample. This is inaccurate as Peuscher et al teach use of a human tissue sample treated (incubated) with a fully human primary antibody (see for example, page 323 of Peuscher et al). Therefore, the rejections presented in this Office Action are maintained.
Conclusion
No claim is allowed.
Notice:
As indicated in the previous office action, a search of the prior art revealed that a Protein A/G having the sequence of SEQ ID NO: 4 is free from the prior art. The closest prior art would be Dalian University of Technology (CN102676562A; noting that the English Machine Translation of the claims obtained from Patent Scope is being relied upon) which teaches a Protein A/G fusion having a sequence comprising 96.8% identity to 89% of the instant SEQ ID NO: 4 (calculated to have 488 residues identical to the 540 residues of SEQ ID NO: 4, where both sequences begin with Met, such that the prior art sequence is deemed to be 90.37% identical to the instant SEQ ID NO: 4 in its totality; see the sequence alignment provided below; see for example, claim 2 reciting SEQ ID NO: 6 of Dalian).
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As noted above, protein manipulation/generation is an unpredictable art marked by a high level of skill. Regarding the state of the art, Listov et al (Opportunities and challenges in design and optimization of protein function. Nat Rev Mol Cell Biol 25, 639–653 (2024)) teach that the primary amino acid sequence determines downstream structure (protein folding), which then determines function (presenting both the inverse folding problem and the inverse function problem (see for example, Figure 1 and its caption; see also Mishra et al (Inaccurate secondary structure predictions often indicate protein fold switching. Protein Sci. 2019 Aug;28(8):1487-1493. doi: 10.1002/pro.3664. Epub 2019 Jun 17)). Expanding on these problems in proteomics, Reardon (Nature 635, 246-248 (2024)) explains that the goal of designing a protein with known and predictable function, binding partners, size, location, and other traits is, for the moment, a dream. Reardon teaches that further challenges in protein design include predicting how a protein, even if it binds to target, will function upon said binding. Reardon teaches that the primary structure (amino acid sequence) of a protein is critical to function, noting that even proteins of similar shape do not execute the same functions, while those with different shapes may carry out the same tasks. Reardon goes on to teach that it is not always apparent which parts of the primary sequence are important; a seemingly useless amino-acid chain on the side of an enzyme, for instance, might affect how tightly a protein can bind to other molecules or its ability to flip between conformational states. Moreover, Reardon explains that when researchers attempt to solve the structure of a protein experimentally, they often end up seeing only the most stable conformation, which is not necessarily the form the protein takes when it is active (see for example, pages 246-247 of Reardon). Therefore, the instant, novel protein sequence is deemed to be free of the art and allowable.
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure.
Patmawati et al (Biotechnol J. 2018 Jun;13(6):e1700624. doi: 10.1002/biot.201700624. Epub 2018 Apr 25) teach that recombinant protein production can create artificial proteins with desired functions by introducing genetic modifications to the target proteins. Horseradish peroxidase (HRP) has been used extensively as a reporter enzyme in biotechnological applications; however, recombinant production of HRP has not been very successful, hampering the utilization of HRP with genetic modifications. A fusion protein comprising an antibody binding protein and HRP will be an ideal bio-probe for high-quality HRP-based diagnostic systems. A HRP-protein A/G fusion protein (HRP-pAG) is designed and its production in silkworm (Bombyx mori) is evaluated for the first time. HRP-pAG is expressed in a soluble apo form, and is activated successfully by incubating with hemin. The activated HRP-pAG is used directly for ELISA experiments and retains its activity over 20 days at 4 degrees C. Moreover, HRP-pAG is modified with biotin by the microbial transglutaminase (MTG) reaction. The biotinylated HRP-pAG is conjugated with streptavidin to form a HRPpAG multimer and the multimeric HRP-pAG produced higher signals in the ELISA system than monomeric HRP-pAG. The successful production of recombinant HRP in silkworm will contribute to creating novel HRP-based bioconjugates as well as further functionalization of HRP by applying enzymatic post-translational modifications.
Liyanage, TD (Research Gate, posted 04/14/2022, obtained from: https://www.researchgate.net/post/I_am_trying_to_develop_an_inhouse_ELISA_to_detect_total_antibody_response_for_T_gondii_in_marsupials_Has_anyone_used_Protein_L_in_an_ELISA) teaches that in ELISA, Protein A/G is a good non-species-specific reagent but it only binds with the Fc region of IgG (see for example, the first post by Liyanage, TD on page 1/6).
US 20190285624 A1, US 20150204868 A1, and US 20140121125 A1 are deemed relevant.
Bhide et al (Comp Immunol Microbiol Infect Dis. 2004 May;27(3):191-9. doi: 10.1016/j.cimid.2003.10.001) teach that Protein A/G conjugates were known in the art for use in immunoassays (see for example, Bhide et al at the Title and abstract at page 191 and paragraph 1 of the Discussion section at page 196).
Vashist et al (Analyst. 2015 Jul 7;140(13):4445-52. doi: 10.1039/c5an00690b) teach a rapid and highly-sensitive surface plasmon resonance (SPR)-based immunoassay (IA) has been developed and validated for detecting human C-reactive protein (CRP), a specific biomarker for inflammatory and metabolic disorders, and infections. The 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide hydrochloride (EDC)-activated protein A/G (Pr A/G) was diluted in 1% (v/v) 3-aminopropyltriethoxysilane (APTES), dispensed on a KOH-treated gold (Au)-coated SPR chip, and incubated for 30 min. The Pr A/G functionalized Au SPR chip was then bound to anti-human CRP capture antibody (Ab), blocked with bovine serum albumin, and subsequently used for the detection of CRP. The highly-simplified oriented Ab immobilization strategy enabled the leach-proof binding of capture Ab in 5-fold shorter time than conventional procedures. The developed IA detected 1.2-80 ng mL(-1) of CRP with a limit of detection (LOD) and a limit of quantification (LOQ) of 1.2 ng mL(-1) and 4.6 ng mL(-1), respectively. It detected CRP spiked in diluted human whole blood, serum and plasma as well as the CRP levels in the ethylenediaminetetraacetic acid (EDTA) plasma samples of patients with the same precision as the clinically-accredited analyzer-based IA and conventional CRP sandwich ELISA. The Ab-bound SPR chips stored at 4 °C retained their functional activity for 10 weeks, resulting in significant reduction in the overall analysis time.
Chow et al (PLoS One. 2017 Apr 5;12(4):e0175081. doi: 10.1371/journal.pone.0175081) teach that large serological studies on animals could provide important information on the geographic distribution of Coccidioides. To facilitate such studies, we used protein A/G, a recombinant protein that binds IgG antibodies from a variety of mammalian species, to develop an enzyme immunoassay (EIA) that detects IgG antibodies against Coccidioides in a highly sensitive and high-throughput manner. We showed the potential of this assay to be adapted to multiple animal species by testing a collection of serum and/or plasma samples from dogs, mice, and humans with or without confirmed coccidioidomycosis.
WO2015036956 A1 claim and teach an in vitro method for quantifying a target-specific test antibody in a test sample, comprising the steps of: a) performing an immunoassay using a target immobilized on a support which is brought into contact with the test sample, the immunoassay comprising a step of measuring the binding of the target-specific test antibody to the immobilized target by using a detectable non-antibody ligand that binds to the Fc region or to a light chain of an antibody, whereby a concentration-related value of the target-specific test antibody in the test sample is obtained, and b) comparing the concentration-related value obtained at step a) with a reference value obtained by performing an immunoassay using the target immobilized on a support which is brought into contact with a calibration sample comprising a known concentration of a target-specific calibration antibody, the immunoassay comprising a step of measuring the binding of the target-specific calibration antibody to the immobilized target by using the detectable non-antibody ligand of step a), and wherein : (i) the target-specific test antibody of step a) and the target-specific calibration antibody of step b) are identical, or (ii) the target-specific test antibody of step a) and the target-specific calibration antibody of step b) are distinct.
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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/Ashley Gao/
Examiner, Art Unit 1678
/GREGORY S EMCH/Supervisory Patent Examiner, Art Unit 1678
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