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 .
Information Disclosure Statement
The information disclosure statements (IDS) submitted on 01/17/2025 and 01/03/2025 are in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statement is being considered by the examiner.
Status of Claims
This office action is in response to Applicant's Response to Election / Restriction filed on October 14, 2025.
Claims 1-7, 11-12, 14, 16-23 and 27-28 are currently pending, with claims 4, 14, 22-23 and 27 withdrawn.
Claims 1-3, 5-7, 11-12 16-21, and 28 are under examination. This is the first action on the merits.
Election/Restrictions
Applicant's election without traverse of the probe detection scheme as shown in Figure 20, filed on October 14, 2025 is acknowledged 1.
Figure 20, as Illustrated below, depicts a method for detecting a protein analyte. The method involves sequential steps, including adding reagent composition to a substrate to form a complex; performing proximity ligation and Rolling Circle Amplification (RCA) for amplification; adding hairpin ECL-labeled probes for hybridization; and finally conducting ECL measurement with the application of a coreactant and voltage. (see also [039]; [0265]lines 11-18) Accordingly, claims 4, 14, 22-23 and 27 are withdrawn as they are drawn to the embodiment of Figure 20.
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Claims 4, 14, 22-23 and 27 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention. Examination on the merits commences on claims 1-3, 5-7, 11-12 16-21, and 28.
Priority
The priority date of the instant claims 1-3, 5-7, 11-12 16-21, and 28 is 12/30/2021, filling date of the US provisional application NO. 63/295,470.
Claim Objections
Claims 6 and 21 are objected to because of the following informalities:
In claim 6, line 1,"The method claim 1" should read "The method of claim 1."
In claim 21, line 8,
"ECL-labeled oligonucleotide probes to the to the oligonucleotide" should read "ECL-labeled oligonucleotide probes to the
Claim Interpretation
In evaluating the patentability of the claims presented in this application, claim terms have been given their broadest reasonable interpretation (BRI) consistent with the specification, as understood by one of ordinary skill in the art, as outlined in MPEP§ 2111.
Regarding claims 1 and 7, they both recite the term "TPA" in the recitation "an ECL co-reactant that is not TPA."
In light of the specification in [005]; [042-043]; [068]; [0303]; [0314], the term "TPA" is understood as synonymous with triethanolamine.
Claim Rejections - 35 USC § 112(b)
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.
Claims 1-3, 5-7, 11-12 16-21, and 28 are rejected under 35 U.S.C. 112(b), 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.
Claims 1 and 7 are indefinite for reciting "a binding partner or binding complex," presenting the terms “binding partner” and “binding complex” as alternatives.
This claim language is indefinite because it is unclear what distinguishes a "binding partner" from a "binding complex," and vice versa. The specification does not provide express definitions for either term. In some instances of the disclosure, a binding complex is described as comprising a binding partner ([0210] lines 19-21); while in other part of the description, a binding partner is described as comprising a binding complex ([0260] line 1).
Therefore, according to the specification, the scope of these terms appear to overlap, and it is unclear which term is broader. This renders the phrase "a binding partner or binding complex" ambiguous ꟷ what would qualify as a binding partner but not a binding complex, and vice versa? This is unclear. Without clear distinction between these terms, the metes and bounds of the claims cannot be determined with reasonable certainty.
Claims 2-3, 5-6, 11-12 16-21, and 28 are rejected for depending from claim 1 and not remedying the indefiniteness.
Claim Rejections - 35 USC § 112(d)
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.
Claim 2 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 2 recites the wherein clause below:
"wherein b) contacting the substrate with the composition comprises:
b') contacting the substrate with a composition comprising the binding partner and/or binding complex;
b") contacting the substrate with a composition comprising the plurality of ECL- labeled oligonucleotide probes; and
b"') contacting the substrate with a composition comprising the ECL co-reactant. "
This dependent claim is improper because it does not add any limitation to the method of claim 1, which already recites contacting the substrate with a single composition comprising the binding partner and/or binding complex; the plurality of ECL- labeled oligonucleotide probes; and the ECL co-reactant. Claim 2, however, describes the same contacting step as using three individual compositions, each comprising the binding partner and/or binding complex; the plurality of ECL- labeled oligonucleotide probes; and the ECL co-reactant. Given the context of the claim, the three individual compositions in claim 2 are subcomponents encompassed by the single composition in claim 1. Thus, the wherein clause in claim 2 does not further modify the method of claim 1, as there is no manipulative difference between the two.
Therefore, claim 2 fails to further limit the subject matter of claim 1, from which it depends.
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
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.
Claims 1-3, 5-7, 11-12 16-21, and 28 are rejected under 35 U.S.C. 103 as being unpatentable over Aghvanyan (Aghvanyan et al. US20190391140A1 - Assay methods ; published 2019-12-26), in view of
Richter (Richter et al. US20010023063A1 - Assays employing electrochemiluminescent labels and electrochemiluminescence quenchers ; Published 2001-09-20), as evidenced by
Schweitzer (Schweitzer et al. , Immunoassays with rolling circle DNA amplification: a versatile platform for ultrasensitive antigen detection. Proc Natl Acad Sci U S A. 2000 Aug 29;97(18):10113-9. doi: 10.1073/pnas.170237197. PMID: 10954739; PMCID: PMC27732) and
Li (Li et al. Aptamer-based fluorescent assay for sensitive detection of cardiac Troponin I. J. Beijing Inst. Technol. 2020, 29, 45).
A) Regarding claims 1 and 7, while they are separate independent claims, they will be addressed together below as the claims' scopes largely overlap. Claim 1 is narrower in scope than claim 7 and further recites a binding reagent on a substrate, which immobilizes binding partner and/or binding complex via binding interaction.
Aghvanyan teaches methods and systems of performing immunoassays for analyte detection, comprising Proximity Ligation and Rolling Circle Amplification using ECL-labeled probes for electrochemiluminescence detection (see examples 1 and example 9; see also Fig. 19).
Regarding claims 1 and 7, Aghvanyan teaches an electrochemiluminescence (ECL) detection method comprising:
providing a substrate comprising an electrode and having a binding reagent (Fig. 19, capture oligo) immobilized on a surface of the substrate([0006]; [0012]; Fig. 19);
contacting the substrate with a composition (Fig. 19; [0329]), the composition comprising:
i) a binding partner and/or binding complex comprising an oligonucleotide (Fig. 19, 1905-2019-1902),
wherein the binding reagent binds the binding partner and/or binding complex;
ii) a plurality of ECL-labeled oligonucleotide probes (Fig. 19, Detection oligonucleotide; [0013] detection probe having ECL label; [0275], detection probe comprising electrochemiluminescence label SULFOTAG) comprising an oligonucleotide sequence that is complementary to an oligonucleotide sequence of the oligonucleotide of the binding partner and/or binding complex;
allowing a portion of the plurality of ECL-labeled oligonucleotide probes to hybridize to the oligonucleotide of the binding partner and/or binding complex (Fig. 19),
wherein the binding partner and/or binding complex is bound by the binding reagent(Fig. 19), and
wherein another portion of the plurality of ECL-labeled oligonucleotide probes is not hybridized to the oligonucleotide of the binding partner and/or binding complex bound by the binding reagent
(Fig. 19; [0329]; [0275], lines 16-19; [0258] lines 15-22, unbound probes are removed by washing);
applying a voltage to the electrode to generate ECL ([0258] lines 22-24; [0012]); and
measuring the ECL ([0258] lines 22-24; [0012-0013]).
In summary, Aghvanyan discloses an electrochemiluminescence (ECL) detection method using ECL labeled probes for detection of analytes. The method involves forming a binding complex attached to a surface comprising an electrode for ELC measurement (Fig 19; [0006]; [0012-0013]), allowing a portion of the ECL-labeled probes to hybridize to the target extension product (e.g., RCA extension product), and removing the portion of excess probes that are not hybridized by through a washing step (Fig. 19; [0329]; [0275], lines 16-19; [0258] lines 15-22, unbound probes are removed by washing).
While Aghvanyan does not specifically teach selectively dequenching and measuring ECL without removing the probes not hybridized to their targets. Richter’s teaching fills this gap.
Richter describes an improved non-separation ECL assaying method and system utilizing an ECL quencher and an ECL label (see Abstract; [0061]). Particularly relevant to Aghvanyan’s teaching of detection probes labeled with ECL label, Richter provides further advancements by specifically teaching self-hybridizing, hairpin detection probes that possesses both an ECL label and an ECL quenching moiety. In the absence of the target DNA, the probe self-hybridizes (typically forming a hairpin or hairpin-loop structure), bringing the ECL label and the ECL quenching moiety into quenching contact. In the presence of the target DNA, the probe preferentially anneals to the target DNA, and in doing so, separates the ECL label from the ECL quenching moiety so that they are no longer in quenching contact, and the ECL emission increases. Thereby allowing for measurement of ECL emission as indicator for amount of target DNA, without the need for a washing step to remove excess detection probes because those do not hybridize to target will stay quenched and not emit ECL ([0167]).
Richter explains that the use of both an ECL label and ECL quencher enables more desirable non-separation assays([0046]; [0050-0051]; [0061]), eliminating the need for physical separation of unused assay reagents, such as washing steps, which are required in the method disclosed in Aghvanyan:
“ [0050] Chemical and biological assays may often be conveniently classified as "separation assays" or "non-separation assays." Generally, in separation assays, the detection product(s) is physically separated from other products and/or unreacted analyte of interest and unreacted assay reagents. (For example, it is often necessary to physically separate the detection product so that only those labels which are part of the detection product are detected, and not those of the excess labeling reagent.) The amount of analyte may then be determined either directly from the amount of labeled detection product, or indirectly from the amount of unused labeling reagent. Separation may often be achieved by exploiting a selective binding reaction between members of a binding pair (e.g., biotin-avidin, antibody-antigen, oligonucleotide hybridization probe-oligonucleotide ). For example, a labeled detection product having one member of a binding pair may be first formed in a fluid phase (e.g., in solution), and separation may then be effected, for example, by capture of the ECL labeled detection product by a solid phase reagent having the other member of the binding pair; the detection product may then be recovered by washing the solid phase free of unreacted analyte and reagents. Many other separation strategies employing binding pairs are well known in the art.
[0051] Assays which do not require a separation step are highly desirable, as they typically require less sample manipulation and are often readily adapted to "real time" assays. Such assays may often be conveniently classified as "non-separation assays." In non-separation assays, the detection product is typically not physically separated from unused assay reagents and unused analyte. Instead, the presence of the detection product is typically detected by a property which at least one of the assay reactants acquires or loses only as a result of contacting the analyte of interest. A number of such non-separation assays have been developed.
…
[0061] Using the efficient ECL quenchers disclosed herein, assays may be developed which employ an ECL label and an ECL quencher and which permit, inter alia, assays such as non-separation assays which offer many, if not all, of the advantages offered by ECL detection methods over other detection methods.” [emphasis added]
Therefore, Richter's teaching of self-hybridizing, hairpin detection probes that possesses both an ECL label and an ECL quenching moiety meet the limitations of selectively dequenching and measuring ECL without removing the probes not hybridized to their targets.
Additionally, Richter teaches a composition for ECL measurement, comprising an ECL co-reactant that is not TPA ([0195]; [0199]; [0200], peroxodisulfate, oxalate; [0030] Ru(bpy)32+ FOR ECL generation with coreactants peroxodisulfate (i e., S2O/-,persulfate) and oxalate; [0042] “ECL of Ru(bpy)3 2+ has been employed in the determination of a wide range of coreactants. For example, Ru(bpy)/+ ECL has been effectively used to determine oxalate and persulfate to levels as low as 10-13 moles/liter.”).
In view of the above, it would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention to modify the method in Aghvanyan by incorporating the hairpin probes comprising both an ECL label and an ECL quencher, as disclosed in Richter, as the detection probe in Aghvanyan's ECL immunoassay.
A skilled artisan would have been motivated to make this modification because the probes disclosed in Richter enable a more desirable assay approach that obviates the need for washing away unbound detection probes. This approach offers potential benefits, such as reduced sample manipulation and greater adaptability to "real-time" assays, as suggested by Richter ([0051]).
Furthermore, a skilled artisan would have found it obvious to incorporate the dual-labeled probes from Richter into Aghvanyan' immunoassay method, because the use of molecular beacon-type hairpin probes in immunoassays with RCA, such as the one depicted in Fig. 19 of Aghvanyan, is well-established in the art, as evidenced by Schweitzer and Li (Fig. 1).
Schweitzer is a review article on the topic of Immunoassays with rolling circle DNA amplification, and teaches: "Incorporation of molecular beacon type detector probes into the immunoRCA assay could also eliminate post hybridization washing steps because unhybridized beacons are nonfluorescent." (page 10118, right-hand col, para 3, lines 22-24).
The person of ordinary skill would have had a reasonable expectation of success in combining these teachings because both Aghvanyan and Richter pertain to the field of analyte detection using ECL assays with ECL-labeled probes. Their teachings overlap and are technically compatible. A skilled artisan would possess the knowledge and skill to apply hairpin detection probes to an immunoassay using RCA, as this approach is well-known in the art.
Additionally, this combination would have been obvious as it represents the KSR principle of predictable use of prior art elements (i.e., the dual-labeled hairpin probes from Richter) according to a known method (i.e., the ECL assay method in Aghvanyan) to yield predictable results (i.e., eliminate the need for post hybridization washing steps). (See MPEP §2143).
B) Regarding claim 2, it is obvious in view of the combined teachings of Aghvanyan and Richter because it does not further limit the claimed method. See detailed discussion in
"Claim Rejections - 35 USC § 112(d)" section above.
Regarding claim 3, Aghvanyan teaches sequential assaying steps ([0329]; [0274-0275]).
Regarding claim 5, Aghvanyan teaches b') contacting the substrate with a first composition comprising the binding partner and/or binding complex and allowing the binding partner and/or binding complex to immobilize on the surface by binding to the binding reagent ([0329]; [0274] lines 14-16); and b") contacting the substrate comprising the immobilized binding partner and/or binding complex with a second composition comprising the plurality of ECL-labeled oligonucleotide probes and allowing a portion of the plurality of ECL-labeled oligonucleotide probes to hybridize to the oligonucleotide of the immobilized binding partner and/or binding complex ([0329]; [0275] lines 6-8); and
Richter teaches contacting the substrate with a third composition comprising an ECL co-reactant ([0195]; [0199]; [0200] ECL assay media is added to sample prior to performing ECL measurement and comprises co-reactants, which "permit the use of simpler means for generating ECL").
Regarding claim 6, Aghvanyan teaches washing the substrate following the contacting the substrate with the binding partner and/or binding complex to remove binding partner and/or binding complex not bound by the binding reagent, wherein the washing is prior to contacting the substrate with the composition comprising the plurality of ECL-labeled oligonucleotide probes ([0275] lines 1-3).
Regarding claim 11, Aghvanyan teaches wherein the binding partner and/or binding complex comprises an analyte (Fig. 19; [0329]).
Regarding claim 12, Aghvanyan teaches analyte comprises a peptide (Fig. 19, autoantibody comprises a peptide).
Regarding claim 16, Aghvanyan teaches wherein the analyte is labeled with the oligonucleotide by binding the analyte with a detection reagent comprising the oligonucleotide(Fig. 19, autoantibody binds to anti-human Ig antibody conjugated to oligo probe).
Regarding claim 17, Aghvanyan teaches the oligonucleotide of the binding partner and/or binding complex comprises multiple copies of the sequence complementary to the oligonucleotide sequence of the plurality of the ECL-labeled oligonucleotide probes (Fig. 19, RCA extends the oligo of 1902, thereby creating multiple binding sites for detection probes; [0142]; [0176-0177]).
Regarding claim 18, Aghvanyan teaches prior to contacting the substrate with the plurality of the ECL-labeled oligonucleotide probes, performing an amplification reaction to generate the multiple copies of the sequence complementary to the oligonucleotide sequence of the plurality of the ECL-labeled oligonucleotide probes (Fig. 19, RCA; [0275]).
Regarding claim 19, Aghvanyan teaches the analyte is labeled with the oligonucleotide by binding the analyte with a detection reagent comprising an oligonucleotide primer, and wherein the oligonucleotide primer is extended by a polymerase to generate the oligonucleotide that comprises the multiple copies of the sequence complementary to the oligonucleotide sequence of the ECL-labeled oligonucleotide probes (Fig. 19).
Regarding claim 20, Aghvanyan teaches rolling circle amplification reaction(Fig. 19).
Regarding claim 21, Richter teaches the ECL-labeled oligonucleotide probes include a stem-loop or hairpin structure, an ECL label, and a quenching moiety, wherein the quenching moiety is in proximity to the ECL label and quenches the ECL label when the oligonucleotide probe is in a stem-loop or hairpin configuration, but does not quench the ECL label when the stem-loop or hairpin structure is in an open configuration, and wherein the selectively dequenching comprises hybridizing the portion of the plurality of ECL-labeled oligonucleotide probes to the to the oligonucleotide of the binding partner and/or binding complex in the open configuration (Richter, [0167]).
Regarding claim 28, Richter teaches piperazine-N,N' -bis(2-ethanesulfonic acid) (PIPES) ([0201] lines 7-8).
Conclusion
Claims 6 and 21 are objected to; claims 1-3, 5-7, 11-12 16-21, and 28 are rejected. No claims are allowed.
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/TIAN NMN YU/Examiner , Art Unit 1681 /AARON A PRIEST/Primary Examiner, Art Unit 1681
1 The probe detection schemes from Figures 14, 15, and 21 are drawn to non-elected inventions.