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 .
Election/Restrictions
Claims 16-20 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on January 9, 2026.
Claim Status
Claims 1-20 are currently pending in the present application. Claims 16-20 are withdrawn as directed to a nonelected invention. Claims 1-15 are currently under examination.
Priority/Effective Filing Date
It is noted that the nucleotide sequences (SEQ ID NOs: 6-9) required by claim 3 were not present in U.S. Provisional Patent Application No. 63/088,179, filed on October 6, 2020. The first disclosure of these sequences in the priority documents was in U.S. Provisional Patent Application No. 63/106,916, filed on October 29, 2020. Therefore, claims requiring these sequences enjoy the benefit of priority to the provisional application filed October 29, 2020.
Information Disclosure Statement
It is noted that no Information Disclosure Statement has been filed in the present application as of the mailing date of this action.
Nucleotide and/or Amino Acid Sequence Disclosures
REQUIREMENTS FOR PATENT APPLICATIONS CONTAINING NUCLEOTIDE AND/OR AMINO ACID SEQUENCE DISCLOSURES
Items 1) and 2) provide general guidance related to requirements for sequence disclosures.
37 CFR 1.821(c) requires that patent applications which contain disclosures of nucleotide and/or amino acid sequences that fall within the definitions of 37 CFR 1.821(a) must contain a "Sequence Listing," as a separate part of the disclosure, which presents the nucleotide and/or amino acid sequences and associated information using the symbols and format in accordance with the requirements of 37 CFR 1.821 - 1.825. This "Sequence Listing" part of the disclosure may be submitted:
In accordance with 37 CFR 1.821(c)(1) via the USPTO patent electronic filing system (see Section I.1 of the Legal Framework for Patent Electronic System (https://www.uspto.gov/PatentLegalFramework), hereinafter "Legal Framework") as an ASCII text file, together with an incorporation-by-reference of the material in the ASCII text file in a separate paragraph of the specification as required by 37 CFR 1.823(b)(1) identifying:
the name of the ASCII text file;
ii) the date of creation; and
iii) the size of the ASCII text file in bytes;
In accordance with 37 CFR 1.821(c)(1) on read-only optical disc(s) as permitted by 37 CFR 1.52(e)(1)(ii), labeled according to 37 CFR 1.52(e)(5), with an incorporation-by-reference of the material in the ASCII text file according to 37 CFR 1.52(e)(8) and 37 CFR 1.823(b)(1) in a separate paragraph of the specification identifying:
the name of the ASCII text file;
the date of creation; and
the size of the ASCII text file in bytes;
In accordance with 37 CFR 1.821(c)(2) via the USPTO patent electronic filing system as a PDF file (not recommended); or
In accordance with 37 CFR 1.821(c)(3) on physical sheets of paper (not recommended).
When a “Sequence Listing” has been submitted as a PDF file as in 1(c) above (37 CFR 1.821(c)(2)) or on physical sheets of paper as in 1(d) above (37 CFR 1.821(c)(3)), 37 CFR 1.821(e)(1) requires a computer readable form (CRF) of the “Sequence Listing” in accordance with the requirements of 37 CFR 1.824.
If the "Sequence Listing" required by 37 CFR 1.821(c) is filed via the USPTO patent electronic filing system as a PDF, then 37 CFR 1.821(e)(1)(ii) or 1.821(e)(2)(ii) requires submission of a statement that the "Sequence Listing" content of the PDF copy and the CRF copy (the ASCII text file copy) are identical.
If the "Sequence Listing" required by 37 CFR 1.821(c) is filed on paper or read-only optical disc, then 37 CFR 1.821(e)(1)(ii) or 1.821(e)(2)(ii) requires submission of a statement that the "Sequence Listing" content of the paper or read-only optical disc copy and the CRF are identical.
Specific deficiencies and the required response to this Office Action are as follows:
Specific deficiency – Nucleotide and/or amino acid sequences appearing in the drawings are not identified by sequence identifiers in accordance with 37 CFR 1.821(d). Sequence identifiers for nucleotide and/or amino acid sequences must appear either in the drawings or in the Brief Description of the Drawings.Figures 8, 9, 10, 13, and 14d contain nucleotide sequences that are not identified by sequence identifiers (i.e. “SEQ ID NO:__”).
Required response – Applicant must provide:
Replacement and annotated drawings in accordance with 37 CFR 1.121(d) inserting the required sequence identifiers;
AND/OR
A substitute specification in compliance with 37 CFR 1.52, 1.121(b)(3) and 1.125 inserting the required sequence identifiers into the Brief Description of the Drawings, consisting of:
A copy of the previously-submitted specification, with deletions shown with strikethrough or brackets and insertions shown with underlining (marked-up version);
A copy of the amended specification without markings (clean version); and
A statement that the substitute specification contains no new matter.
Claim Interpretation
Claim 4 recites the phrase “unpaired probability”. The specification provides no special definition for this claim term. The examiner has interpreted this claim term according to the definition known in the art, described by Raden et al., “Interactive implementations of thermodynamics-based RNA structure and RNA-RNA interaction prediction approaches for example-driven teaching” PLoS Computational Biology 14(8): e1006341 (published August 30, 2018).
Raden et al. define an “unpaired probability” for a given subsequence within a polynucleotide is “the probability of all structures that show no base pairing in the single-stranded subsequence” according to the following equations and “The unpaired probability is also sometimes termed “accessibility”, as an unpaired region in an RNA is accessible for pairing to another RNA”.
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Therefore, the limitation “wherein the first anti-sense oligonucleotide has an unpaired probability for the first nucleic acid sequence of at least 0.5” has been interpreted as requiring at least a 50% chance that the first anti-sense oligonucleotide does not base-pair with itself.
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.
Claims 2 and 6 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 regards as the invention.
The term “near to” in claim 2 is a relative term which renders the claim indefinite. The term “near to” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention.
The claim recites: “a plurality of second anti-sense oligonucleotides, the sequence of which is complementary to a second nucleic acid sequence in the target gene of the biological pathogen near to the first nucleic acid sequence; wherein the first electrode is additionally connected to a first end of the plurality of second anti-sense oligonucleotides”.
It is unclear whether the relative term “near to” is meant to require that the first and second anti-sense oligonucleotides are “near to” each other on the first electrode, or that the first and second nucleic acid sequences in the target gene are “near to” each other in the target gene sequence. Furthermore, the specification and the claims do not provide a standard as to the limits of the requisite distance between a) the first and second anti-sense oligonucleotides on the first electrode (i.e. a separation in meters) or b) the first and second nucleotide sequences within the target gene (i.e. a separation in nucleotides or a separation in meters).
The term “the signal is provided additionally by binding of the plurality of second anti-sense oligonucleotides to the second nucleic acid sequence in the target gene” renders the claim indefinite because it is unclear if the claim requires: a) binding of the second anti-sense oligonucleotide (ASO2) to the second target sequence provides additional signal to that produced by the binding of the first anti-sense oligonucleotide (ASO1) to the first target sequence (i.e. signal is produced by ASO1:Target1 binding OR ASO2:Target2 binding), or b) the signal is provided by the combined binding of ASO1 AND ASO2 to their respective target sequences (i.e. the method requires both ASO1 AND ASO2 to be bound to produce signal).
Claim 6 requires that the first anti-sense oligonucleotide “has a tendency” to form a hairpin-loop structure. A “tendency” to form a hairpin-loop structure is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. It is unclear whether the claim requires that the first anti-sense oligonucleotide has a particular threshold for probability of pairing of two subsequences resulting in a hairpin-loop, or simply that the oligonucleotide comprises any sequence that may form a hairpin-loop at any probability under any conditions.
Claim Rejections - 35 USC § 102
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 1, 2, and 7-11 are rejected under 35 U.S.C. 102(a)(1) and 35 U.S.C. 102(a)(2) as being anticipated by Henkens et al., US 2007/0111202 A1 (published May 17, 2007).
Regarding claim 1, Henkens et al. teach electrochemical biosensors comprising capture probes that are attached at one end to an electrode (Henkens et al., figure 3 and paragraphs 0027 and 0041). Henkens et al. further teach the capture probe is complementary to a target sequence of a pathogen target gene (Henkens et al., paragraph 0105) and that the device comprises a first working electrode and a second reference electrode (Henkens et al., paragraph 0036).
Regarding claim 2, Henkens et al. teach biosensors comprising a second anti-sense oligonucleotide connected to the electrode that is complementary to the same target gene (Henkens et al., paragraph 0105-0106).
Regarding claim 7, Henkens et al. teach that the second electrode is a counter electrode or a reference electrode (Henkens et al., paragraph 0036).
Regarding claim 8, Henkens et al. teach biosensors further comprising a third electrode electrically connected to the first and second electrodes (Henkens et al., paragraph 0104).
Regarding claim 9, Henkens teaches the biosensor further comprises a substrate (a biosensor array of working and reference electrodes (Henkens et al., paragraph 0024) and a conductive film comprising gold, colloidal gold, carbon, or screen-printed conductive ink deposited on the surface of the substrate (Henkens et al., paragraph 0061).
Regarding claims 10 and 11, Henkens et al. teach the capture probes (i.e. the first anti-sense oligonucleotides) are bound at one end to colloidal gold particles (i.e. are capped with conductive nanoparticles; gold nanoparticles) (Henkens et al., figure 13).
Claims 1, 4, 5, and 7-12 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Wang et al., “Ultrasensitive electrochemical supersandwich DNA biosensor using a glassy carbon electrode modified with gold particle-decorated sheets of graphene oxide”. Microchem Acta (2014) 181:935-940 as evidenced by Raden et al., “Interactive implementations of thermodynamics-based RNA structure and RNA-RNA interaction prediction approaches for example-driven teaching” PLoS Computational Biology 14(8): e1006341 (published August 30, 2018 and as evidenced by Northwestern University “oligonucleotide properties calculator”, oligocalc.eu (2007).
Regarding claim 1, Wang et al. teach antisense oligonucleotide probes for electrochemical detection of E. coli DNA sequences (i.e. a target gene of a biological pathogen) (Wang et al., page 936, column 2, paragraph 3 and figure 1, reproduced below for convenience).
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Wang et al. teach sensing the presence of the target DNA by contacting a sample comprising the target DNA with a sensing element comprising “capture probes” (i.e. antisense oligonucleotides complementary to the target DNA) that are connected at a first end to a first electrode, wherein contact between binding of the target DNA to the capture probes provides a signal to identify the presence of the biological pathogen (i.e. E. coli) (Wang et al., figure 1). Wang et al. further teach the apparatus comprises a three electrode system consisting of a glassy carbon electrode (i.e. the first electrode), an auxiliary electrode, and a reference electrode (Wang et al., page 936, column 2, paragraph 3).
Therefore, Wang et al. teach all of the claimed structural elements, arranged as required by claim 1.
Regarding claim 4, the anti-sense oligonucleotide: “5’-CTTCCTCCCCGCTGATATTAACTTTACTCC-3’” taught by Wang et al. does not appear to comprise any sequences capable of forming stable intramolecular structures (e.g. hairpins). Furthermore, the web-based tools taught by Raden et al. do not predict that this sequence (with T substituted for U) will fold into a stable secondary structure. Averaging the predicted unpaired probabilities for all of the individual nucleotides in the sequence gives an unpaired probability of 0.62 (see “MEA_Raden_webtoolOutput_Wang” reference). The unpaired probability is an inherent property of the nucleotide sequence taught by Wang et al.
Regarding claim 5, the anti-sense oligonucleotide: “5’-CTTCCTCCCCGCTGATATTAACTTTACTCC-3’” taught by Wang et al. is estimated to exhibit a ΔG for annealing to its complement of 39.3 kcal/mol (i.e. a binding energy of -39.3 kcal/mol) (See “WangOligo_NorthwesternOligoCalc” reference for calculation details). The binding energy of the oligonucleotide taught by Wang et al. to its complement is an inherent property of its sequence.
Regarding claim 7, Wang et al. teach the apparatus comprises a second electrode that is a reference electrode (Wang et al., page 936, column 2, paragraph 3).
Regarding claim 8, Wang et al. teach the apparatus further comprises a third electrode connected to the first and second electrodes (Wang et al., page 936, column 2, paragraph 3).
Regarding claim 9, Wang et al. teach the electrochemical biosensor further comprises a substrate that is coated with a conductive film on the surface of the substrate (Wang et al., page 936, column 2, paragraph 5 and figure 1).
Regarding claims 10 and 11, Wang et al. teach the anti-sense oligonucleotides are capped with gold nanoparticles (Wang et al., figure 1).
Regarding claim 12, Wang et al. teach the conductive film of claim 9 comprises graphene (Wang et al., page 936, column 2, paragraph 5 and figure 1).
Claims 1, 4, 5, and 7-11 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Tripathy et al., “A miniaturized electrochemical platform with an integrated PDMS reservoir for label-free DNA hybridization detection using nanostructured Au electrodes” Analyst 2019, 144, 6953 (published September 27, 2019) as evidenced by Raden et al., “Interactive implementations of thermodynamics-based RNA structure and RNA-RNA interaction prediction approaches for example-driven teaching” PLoS Computational Biology 14(8): e1006341 (published August 30, 2018 and as evidenced by Northwestern University “oligonucleotide properties calculator”, oligocalc.eu (2007).
Regarding claim 1, Tripathy et al. teach an electrochemical biosensor for detection of Dengue virus nucleic acids (i.e. a biological pathogen in a sample) (Tripathy et al., figure 1 and page 6956, column 2-page 1, column 1 bridging paragraph).
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Tripathy et al. teach the biosensor comprises a sensing element comprising first anti-sense oligonucleotides (i.e. a probe corresponding to Dengue virus consensus primer 5’-CGGTTTCTCGCGCGTTTCAGCATATTGA-3’) that are complementary to a target nucleic acid sequence in a target gene of a pathogen (i.e. Dengue Virus 5’-TCAATATGCTGAAACGCGCGAGAAACCG-3’ (Tripathy et al., figure 1 and page 6956, column 2-page 1, column 1 bridging paragraph). Tripathy et al. teach the probe is immobilized at a first end to a first “working electrode” and the device further comprises a second “counter electrode” electrically connected to the first electrode (Tripathy et al., figure 1). Tripathy et al. teach that hybridization of the immobilized probe to a complementary pathogen nucleic acid provides a signal identifying the presence of the target nucleic acid (i.e. the biological pathogen).
Regarding claim 4, the anti-sense oligonucleotide sequence taught by Tripathy et al. does not appear to comprise any sequences capable of forming stable intramolecular structures (e.g. hairpins). Furthermore, the web-based tools taught by Raden et al. do not predict that this sequence (with T substituted for U) will fold into a stable secondary structure. Averaging the predicted unpaired probabilities for all of the individual nucleotides in the sequence gives an unpaired probability of 0.5 (see “MEA_Raden_webtoolOutput_Tripathy” reference). The unpaired probability is an inherent property of the nucleotide sequence taught by Tripathy.
Regarding claim 5, the anti-sense oligonucleotide: “5’-CTTCCTCCCCGCTGATATTAACTTTACTCC-3’” taught by Tripathy et al. is estimated to exhibit a ΔG for annealing to its complement of 41 kcal/mol (i.e. a binding energy of -41 kcal/mol) (See “Tripathy2019_Oligo_NorthwesternOligoCalc” reference for calculation details). The binding energy of the oligonucleotide taught by Tripathy et al. to its complement is an inherent property of its sequence.
Regarding claim 7, Tripathy et al. teach the second electrode is a counter electrode (Tripathy et al., figure 1).
Regarding claim 8, Tripathy et al. teach the electrochemical biosensor further comprises a third electrode (i.e. “working, counter, and reference electrodes”) connected to the first and second electrodes (Tripathy et al., page 6956, column 1 and figure 3).
Regarding claim 9, Tripathy et al. further teach the electrochemical biosensor comprises a substrate coated with a conductive film on which the first and second electrodes are deposited (Tripathy et al., figure 3).
Regarding claims 10 and 11, Tripathy et al. teach the anti-sense oligonucleotides are capped (i.e. linked at one end) with a gold nanoparticle (Tripathy et al., figure 1).
Claims 1, 7, and 8 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Heer et al., “CMOS Electro-Chemical DNA-Detection Array with On-Chip ADC” ISSCC 2008 Session 8 Medical & Displays 8.4 pg 168-170 (published 2008).
Regarding claim 1, Heer et al. teach an electrochemical biosensor for detection of HIV DNA in a sample (Heer et al., page 168, column 1, paragraph 1). Heer et al. teach the electrochemical biosensor comprises a sensing element wherein first anti-sense oligonucleotides are immobilized at one end to a first electrode and the device further comprises a second electrode electrically connected to the first electrode (Heer et al., figure 8.4.1, reproduced below).
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Regarding claim 7, Heer et al. further teach the second electrode is a reference electrode or a counter electrode (Heer et al., figure 8.4.1).
Regarding claim 8, Heer et al. further teach the biosensor comprises a third electrode connected to the first and second electrodes (Heer et al., figure 8.4.1).
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.
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 and 4-15 are rejected under 35 U.S.C. 103 as being unpatentable over Wang et al., “Ultrasensitive electrochemical supersandwich DNA biosensor using a glassy carbon electrode modified with gold particle-decorated sheets of graphene oxide”. Microchem Acta (2014) 181:935-940 as evidenced by Raden et al., “Interactive implementations of thermodynamics-based RNA structure and RNA-RNA interaction prediction approaches for example-driven teaching” PLoS Computational Biology 14(8): e1006341 (published August 30, 2018 and as evidenced by Northwestern University “oligonucleotide properties calculator”, oligocalc.eu (2007) in view of Kékedy-Nagy et al., “Effect of a Dual Charge on the DNA-conjugated Redox Probe on DNA Sensing by Short Hairpin Beacons Tethered to Gold Electrodes” Analytical Chemistry, 2016, 88, 7984-7990.
Regarding claim 1, Wang et al. teach antisense oligonucleotide probes for electrochemical detection of E. coli DNA sequences (i.e. a target gene of a biological pathogen) (Wang et al., page 936, column 2, paragraph 3 and figure 1, reproduced below for convenience).
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Wang et al. teach sensing the presence of the target DNA by contacting a sample comprising the target DNA with a sensing element comprising “capture probes” (i.e. antisense oligonucleotides complementary to the target DNA) that are connected at a first end to a first electrode, wherein contact between binding of the target DNA to the capture probes provides a signal to identify the presence of the biological pathogen (i.e. E. coli) (Wang et al., figure 1). Wang et al. further teach the apparatus comprises a three electrode system consisting of a glassy carbon electrode (i.e. the first electrode), an auxiliary electrode, and a reference electrode (Wang et al., page 936, column 2, paragraph 3).
Regarding claim 4, the anti-sense oligonucleotide: “5’-CTTCCTCCCCGCTGATATTAACTTTACTCC-3’” taught by Wang et al. does not appear to comprise any sequences capable of forming stable intramolecular structures (e.g. hairpins). Furthermore, the web-based tools taught by Raden et al. do not predict that this sequence (with T substituted for U) will fold into a stable secondary structure. Averaging the predicted unpaired probabilities for all of the individual nucleotides in the sequence gives an unpaired probability of 0.62 (see “MEA_Raden_webtoolOutput_Wang” reference). The unpaired probability is an inherent property of the nucleotide sequence taught by Wang et al.
Regarding claim 5, the anti-sense oligonucleotide: “5’-CTTCCTCCCCGCTGATATTAACTTTACTCC-3’” taught by Wang et al. is estimated to exhibit a ΔG for annealing to its complement of 39.3 kcal/mol (i.e. a binding energy of -39.3 kcal/mol) (See “WangOligo_NorthwesternOligoCalc” reference for calculation details). The binding energy of the oligonucleotide taught by Wang et al. to its complement is an inherent property of its sequence.
Regarding claim 6, Wang et al. do not teach a first anti-sense oligonucleotide that has a tendency to form a hairpin-loop structure.
However, Kékedy-Nagy et al. teach electrochemical biosensors for detection of nucleic acids in a sample wherein the first anti-sense oligonucleotide has a tendency to form a hairpin loop (Kékedy-Nagy et al., figure 1 and abstract).
Kékedy-Nagy et al. teach that hairpin-loop forming electrochemical probes advantageously provides for robust “off-on” nanomolar DNA sensing (Kékedy-Nagy et al., abstract). Kékedy-Nagy et al. further teach that one of the simplest off-on “genosensor” platforms relies on relatively short one-loop one-step hairpins immobilized on gold on one end and the other end coupled to methylene blue (i.e. a redox sensor) for highly sensitive detection of a complementary sequence that is easy to regenerate (Kékedy-Nagy et al., page 7984, column 1, paragraph 3).
Therefore, it would have been prima facie obvious prior to the effective filing date of the claimed invention for one of ordinary skill in the art to have modified the non-hairpin forming anti-sense oligonucleotides taught by Wang et al. for electrochemical detection of hybridization between the anti-sense probe and a complementary target pathogen gene sequence with the teachings of Kékedy-Nagy et al. comprising first anti-sense oligonucleotides with self-complementary 3’ and 5’ ends, capable of forming a hairpin-loop structure (Kékedy-Nagy et al., figure 1).
The ordinary artisan would have been motivated to modify the electrochemical biosensor taught by Wang et al. with the hairpin-folding anti-sense oligonucleotide features taught by Kékedy-Nagy et al. because Kékedy-Nagy et al. teach that hairpin-loop probes are one of the simplest implementations of an off-on genosensor with robust, easily interpretable off-on discrimination between the folded beacon and the open (hybridized) dsDNA states (Kékedy-Nagy et al., page 7989, column 2).
Regarding claim 7, Wang et al. teach the apparatus comprises a second electrode that is a reference electrode (Wang et al., page 936, column 2, paragraph 3).
Regarding claim 8, Wang et al. teach the apparatus further comprises a third electrode connected to the first and second electrodes (Wang et al., page 936, column 2, paragraph 3).
Regarding claim 9, Wang et al. teach the electrochemical biosensor further comprises a substrate that is coated with a conductive film on the surface of the substrate (Wang et al., page 936, column 2, paragraph 5 and figure 1).
Regarding claims 10 and 11, Wang et al. teach the anti-sense oligonucleotides are capped with gold nanoparticles (Wang et al., figure 1).
Regarding claim 12, Wang et al. teach the conductive film of claim 9 comprises graphene (Wang et al., page 936, column 2, paragraph 5 and figure 1).
Regarding claim 13, Kékedy-Nagy et al. teach a first anti-sense oligonucleotide forms hairpin-loop structures in the absence of target nucleic acids and unfolds in the presence of the target nucleic acids, wherein the unfolding provides the detectable signal (Kékedy-Nagy et al., Abstract (see figure below)).
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Regarding claim 14, Kékedy-Nagy et al. teach the second end (the end that is not bound to the electrode) of the anti-sense oligonucleotide is bound to a redox reporter (i.e. the red circle in the figure above) molecule such that the redox reporter is in proximity to the electrode in the hairpin (off) state, and upon binding to the target nucleic acid, the redox reporter is moved away from the first electrode, providing the signal.
Regarding claim 15, Kékedy-Nagy et al. teach that the redox reporter is methylene blue (Kékedy-Nagy et al., abstract).
Claims 1 and 3 are rejected under 35 U.S.C. 103 as being unpatentable over Wang et al., “Ultrasensitive electrochemical supersandwich DNA biosensor using a glassy carbon electrode modified with gold particle-decorated sheets of graphene oxide”. Microchem Acta (2014) 181:935-940 as evidenced by Raden et al., “Interactive implementations of thermodynamics-based RNA structure and RNA-RNA interaction prediction approaches for example-driven teaching” PLoS Computational Biology 14(8): e1006341 (published August 30, 2018 and as evidenced by Northwestern University “oligonucleotide properties calculator”, oligocalc.eu (2007) in view of Tripathy et al., “Label-Free Electrochemical Detection of DNA Hybridization: A method for COVID-19 Diagnosis” Transactions of the Indian National Academy of Engineering (2020) 5:205-209 (published May 25, 2020), Genbank: MT246667, Thornburg et al. (published July 2, 2020), Metsky et al., US 2018/0340215 A1 (published November 29, 2018), Tanner et al., US 10,968,493 B1 (Granted April 6, 2021 and filed July 24, 2020), and Zhang et al., US 2021/0292823 A1 (Published September 23, 2021 and filed June 5, 2020).
It is noted that Tanner et al. and Zhang et al. are prior art under U.S.C. 102(a)(2), as they are U.S. Patents or Published U.S. Patent Applications with filing dates prior to the effective filing date of the present application.
Regarding claim 1, Wang et al. teach antisense oligonucleotide probes for electrochemical detection of E. coli DNA sequences (i.e. a target gene of a biological pathogen) (Wang et al., page 936, column 2, paragraph 3 and figure 1, reproduced below for convenience).
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Wang et al. teach sensing the presence of the target DNA by contacting a sample comprising the target DNA with a sensing element comprising “capture probes” (i.e. antisense oligonucleotides complementary to the target DNA) that are connected at a first end to a first electrode, wherein contact between binding of the target DNA to the capture probes provides a signal to identify the presence of the biological pathogen (i.e. E. coli) (Wang et al., figure 1). Wang et al. further teach the apparatus comprises a three electrode system consisting of a glassy carbon electrode (i.e. the first electrode), an auxiliary electrode, and a reference electrode (Wang et al., page 936, column 2, paragraph 3).
Regarding claim 3, Wang et al. do not teach that the biological pathogen is SARS-CoV-2 and wherein the sequence of the first anti-sense oligonucleotide comprises one of SEQ ID NO: 6-9.
However, Tripathy et al. teach miniature electrochemical platforms comprising a gold working electrode, a platinum counter electrode, and a platinum reference electrode wherein the working electrode is coated with anti-sense oligonucleotide probes complementary to SARS-CoV-2 nucleic acids coupled to gold nanoparticles coating the working electrode (Tripathy et al., figures 1, 2, and page 207, column 1-2 bridging paragraph). Tripathy et al. teach that the single-stranded probe can be designed against any particular SARS-CoV-2 virus RNA or corresponding cDNA (Tripathy et al., page 207, column 1, paragraph 2).
Neither Wang et al. nor Tripathy et al. teach that the particular SARS-CoV-2-specific anti-sense oligonucleotide for SARS-CoV-2 detection is one of SEQ ID No. 6-9.
However, it is noted that each of these sequences are the naturally occurring complementary sequence to a portion of the SARS-CoV-2 N gene comprised within the published genome of SARS-CoV-2 (Genbank).
Furthermore, sequence 27 (an oligonucleotide for detection of Coronavirus) taught by Tanner et al. comprises a nucleic acid sequence that is 100% identical to the presently claimed SEQ ID NO: 7 (see alignment below).
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Furthermore, sequence 318054 (a probe for the detection of Coronavirus) taught by Metsky et al. comprises a sequence that is 100% identical to the presently claimed SEQ ID NO: 8 (see alignment below).
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Furthermore, sequence 61985 (a probe for the detection of Coronavirus) taught by Zhang et al. comprises a sequence that is 100% identical to the presently claimed SEQ ID NO: 9 (see alignment below).
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Therefore, it would have been prima facie obvious prior to the effective filing date of the claimed invention for one of ordinary skill in the art to have modified the electrochemical biosensor taught by Wang et al. by selecting any SARS-CoV-2 genomic fragment (or cDNA fragment thereof) as taught by Tripathy et al. as the first anti-sense oligonucleotide to which a SARS-CoV-2 RNA or cDNA molecule hybridizes, thus generating a detectable signal in the biosensor taught by Wang et al. The ordinary artisan would have been motivated to modify the electrochemical biosensor taught by Wang et al. with SARS-CoV-2-hybridizing first anti-sense oligonucleotides because of the explicit teaching of Tripathy et al. that “the target nucleotide can be the COVID-19 specific viral RNA, or the corresponding cDNA, or any unique sequence specific to them. A complementary single-stranded probe can then be designed, with respect to the target sequence, with thiol modification at one end. The thiol modified probe can be attached onto the gold sensing electrodes via gold-thiol self-assembly… when the target… is introduced onto the sensor… it hybridizes with the complementary probe… recorded using electrochemical techniques…[for] diagnosis” (Tripathy et al., page 207, column 1-2 bridging paragraph).
Regarding the specific sequences of SEQ ID NO: 6-9, as described above, probes for the detection of Coronaviruses comprising sequences 7-9 were known in the art prior to the effective filing date of the claimed invention. Furthermore, SEQ ID NO: 6 is a fragment of the naturally occurring SARS-CoV-2 genome taught by Genbank prior to the effective filing date of the claimed invention.
Following the teachings of Tripathy et al., the ordinary artisan would have been motivated to utilize “any unique sequences specific to [the COVID-19 specific viral RNA or the corresponding cDNA” (Tripathy, page 207, column 1, paragraph 2) as a complementary probe in an electrochemical biosensor such as those taught by Tripathy et al. and Wang et. al.
Conclusion
No claim is allowed.
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/Z.M.T./Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682