DETAILED ACTION
The amendment filed on 10/28/2025 has been entered. No new matter has been added.
Claims 11, 2, 6, 7, 11, 17, 19, and 21 are amended in the claim set filed on 10/28/2025.
Claims 1-2, 4-7, 9-12, 14-17, and 19-24 in the claim set filed on 10/28/2025 are pending and currently under examination.
Response to the Arguments
Objections to the claims in the previously mailed non-final have been withdrawn in light of applicants arguments on Pg. 9.
Objections to the Specification in the previously mailed non-final are maintained in the absence of a IDS being filed.
Applicant’s arguments regarding previous rejection(s) of claim(s) 6-7, 11 and 17 under 35 U.S.C. 112 have been fully considered and are persuasive. The 35 U.S.C. 112 rejections documented in the previously mailed non-final have been withdrawn in light of applicants claim amendments and arguments on Pg. 10.
Applicant’s arguments regarding previous rejection(s) of claim(s) 21 under 35 U.S.C. 112 in the previously mailed non-final have been maintained in light of applicants’ partial argument or clarification.
Applicant’s arguments regarding previous rejection(s) of claim(s) 1-2, 4-7, 10-12, 14-17, 19-20, and 22-24 under 35 U.S.C. 103 have been fully considered and are not persuasive. Applicant’s argument on Pg. 12, states that “The cited references do not teach b) (ligating a nucleotide to the free 3' -OH moiety of at least one bound probe (by contacting the tissue sample with a first plurality of nucleotide-polymerase complexes) ... wherein at least one nucleotide-polymerase complex in the first plurality comprises the nucleotide operably linked to a polymerase via a photocleavable linker; c) illuminating at least one location of the tissue sample with light sufficient to cleave the photocleavable linker ...ligating a nucleotide to the free 3' OH moiety of the spatial barcode domain of at least one bound probe...f) collecting the probes bound to target analytes in the tissue sample.” The 35 U.S.C. 103 rejections documented in the previously mailed non-final have been maintained and revised (documented below on Pg. 4-23) in light of applicants’ arguments on Pg. 11-16.
Applicant’s arguments regarding previous rejection(s) of claim(s) 1-2, 4, 10, 12, 16, and 22-24 under Double Patenting have been fully considered and are not persuasive. The Double Patenting rejections documented in the previously mailed non-final have been maintained (documented below on Pg. 23-30) in light of the arguments and revised rejections under U.S.C. 103 documented below.
The rejections for claims 1-2, 4-7, 10-12, 14-17, 19-20, and 22-24 are documented below in this Final Office Action are maintained and revised.
Priority
This application is a U.S. National Phase application, filed under 35 U.S.C. 371, of International Application No. PCT/US2021/037772, filed June 17, 2021, which claims priority to, and the benefit of, U.S. Provisional Application No. 63/040,651, filed June 18, 2020.
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.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim 21 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 21 is/remains indefinite over the limitation “the first location of the tissue sample and the at least second location of the tissue sample are no more than about 500 nm in the x and/or y direction and no more than about 1500 nm in the z direction”. It is unclear whether the 500 nm in the x and/or y direction is in relation to itself as a starting and ending point indicating the size of location or in relation to another one or more locations.
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.
Claims 1-2, 4-7, 10-12, 14-17, 19-20 and 22-24 are/remain rejected under 35 U.S.C. 103 as being unpatentable over Beechem et al. (“Beechem”; US Patent App. Pub. No. US 20170016909 A1 Jan. 19, 2017) in view of Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012), Palluk et al. (“Palluk”; (2018). De novo DNA synthesis using polymerase-nucleotide conjugates. Nature biotechnology, 36(7), 645-650.), and Frenz et al. (“Frenz”; US Patent App. Pub. No. US 20200277664 A1, Sept. 3, 2020, filed on May 5, 2020; publication of Application No: 16/876,947, which is a CON of PCT/US2019/065072 filed on Dec. 6, 2019).
Interpretations: With regard to claim 1, “target” is considered to be any nucleic acid of interest and/or protein of interest of the sample.
With regard to claims 1 and 2, “nucleotide” is considered to be any nucleotide of interest, thus comprising reversible terminator nucleotides.
Beechem discloses probes, compositions, methods, and kits for simultaneous, multiplexed detection and quantification of protein expression in a user-defined region of a tissue, user-defined cell, and/or user-defined subcellular structure within a cell. (Abstract)
Regarding claim 1, Beechem teaches a method comprising “the method provides simultaneous spatially-resolved protein detection of a tissue sample” (Para. 15). Beechem teaches a method comprising “contacting at least one protein target in or from at least one cell in a tissue sample with at least one probe comprising a target-binding domain…” and “two or more protein targets are detected” and “detecting includes quantifying the abundance of each target” (Para. 4). Beechem teaches a method comprising “a second specific location of the tissue sample” (Para. 5).
Regarding claims 1 and 2 step (a), Beechem teaches a method comprising “contacting at least one protein target in or from at least one cell in a tissue sample with at least one probe comprising a target-binding domain” (Para. 4). Beechem also teaches a method comprising “nCounter® Molecular Barcodes. nCounter® systems and methods from NanoString Technologies®… are a preferred means for identifying target proteins and/or target nucleic acids” (Para. 134). NCounter molecular barcodes are interpreted as a target identification domain. Beechem teaches a method comprising “A set of probes includes at least one species of probes, i.e., directed to one target. A set of probes preferably includes at least two, … or more species of probes” (Para. 107).
Regarding claims 1 and 2 steps (b and d-e), Beechem teaches a method comprising “An ordered series of spatially-separable and spectrally resolvable labels of a probe is herein referred to as barcode or as a label code. The barcode or label code allows identification of a target nucleic acid or target protein that has been bound by a particular probe (Para. 89).
Regarding claims 1 and 2 step (c), Beechem teaches a method comprising “providing a force to a location of the tissue sample” (Para. 4) and “The cleavable linker may be photo-cleavable, which is cleaved by light” (Para. 7) and “the illumination of a region of interest” (Para. 10).
Regarding claims 1 and 2 step (e), Beechem teaches a method comprising “in a tissue sample with at least one probe comprising a target-binding domain and a signal oligonucleotide” and from a specific location of the tissue sample (Para. 4). and “repeating at least steps (2) and (3) on at least a second specific location of the tissue sample” (Para. 5). Beechem teaches a method comprising “A set of probes includes at least one species of probes, i.e., directed to one target. A set of probes preferably includes at least two, … or more species of probes” (Para. 107).
Regarding claims 1 and 2 step (f), Beechem teaches a method comprising “collecting and identifying the released signal oligonucleotide, thereby detecting the at least one protein target in or from a specific location of the tissue sample” (Para. 4). Beechem teaches a method comprising oligonucleotides that are collected only for probes that are bound to targets within region of interest (ROI), thereby permitting detection of the identities and quantities of the targets (proteins and/or nucleic acids) located within ROI (Para. 127). Beechem teaches a method comprising “ In this example, proteins in a formalin-fixed paraffin embedded (FFPE) tissue section were labeled with antibody-comprising probes that included photo-cleavable linkers and fluorescent barcodes. The probes—in a user-defined ROI of the FFPE tissue section—were subsequently exposed to focused UV light, thereby releasing the signal oligonucleotides (comprising the fluorescent barcodes) from the ROI. The released signal oligonucleotides were washed away from the FFPE sample and collected. The fluorescent barcodes from the released signal oligonucleotides were then recognized and digitally counted”(Para. 146).
Regarding claims 1 and 2 step (g), Beechem teaches a method comprising “simultaneous spatially-resolved protein detection of a tissue sample” (Para.15). Beechem teaches a method comprising “multiplexed detection and quantification of protein and/or nucleic acid expression in a user-defined region of a tissue” (Para. 81). Beechem teaches a method comprising “The plurality of target proteins and/or target nucleic acids present in each region of interest in a sample are identified in each eluate sample using… a sequencing reaction…” and “can provide spatially-resolved digital profile of protein abundance, spatially-resolved digital profile of protein and nucleic acid abundance, or spatially-resolved digital profile of nucleic acid abundance.
Regarding claim 10, Beechem teaches a method comprising Beechem teaches a method comprising “An ordered series of spatially-separable and spectrally resolvable labels of a probe is herein referred to as barcode or as a label code. The barcode or label code allows identification of a target nucleic acid or target protein that has been bound by a particular probe (Para. 89); “contacting at least one protein target in or from at least one cell in a tissue sample with at least one probe comprising a target-binding domain…” and “two or more protein targets are detected” and “detecting includes quantifying the abundance of each target” (Para. 4). Beechem teaches a method comprising “a second specific location of the tissue sample” (Para. 5); “spatially-separable and spectrally resolvable labels” is interpreted as unique labels that are specific to the location. Thus, Beechem teaches a method wherein the spatial barcode domain of at least one probe bound to a target analyte in the first location of the tissue sample is specific to the first location of the tissue sample, and the spatial barcode domain of at least one probe bound to a target analyte in the at least second location of the tissue sample is specific to the at least second location of the tissue sample.
Regarding claim 19-20, Beechem teaches a method wherein “illumination of a region of interest …comprises use of a laser scanning device” (Para. 10). The “laser scanning device” is interpreted as comprising a two-photon excitation microscope. Thus, Beechem teaches a method wherein the illumination in step (c) is provided by a light source selected from the group comprising an arc-lamp, a laser, a focused UV light source, and light emitting diode; and wherein the illumination in step (c) is provided by a two-photon excitation microscope.
Regarding claim 22, Beechem teaches a method wherein “detecting the at least one protein target in or from a specific location of the tissue sample” (Para. 4) and “repeating… on at least a second specific location of the tissue sample, the second specific location comprising at least a second cell” (Para. 5). Thus, Beechem teaches a method wherein the first location of the tissue sample and the at least second location of the tissue sample(a) are subcellular; (b) each comprise no more than one cell; or (c) each comprise no more than ten cells.
Regarding claim 24, Beechem teaches a method wherein “(5) amplifying the ligation product” and “(6) … sequencing the amplified products produced in step (5)” (Para. 5). Thus, Beechem teaches a method he method further comprising after step (f) and prior to step (g), amplifying the collected probes.
Beechem does not explicitly teach all the limitations recited in (i) claims 1 and 2 step (b) – step (e), 4-7, and 23, (ii) claims 10-12 and 14-17.
(i) Frisen discloses methods and products for the localized or spatial detection of nucleic acid in a tissue sample and in particular to a method for localized detection of nucleic acid in a tissue sample comprising: (a) providing an array comprising a substrate on which multiple species of capture probes are directly or indirectly immobilized such that each species occupies a distinct position on the array and is oriented to have a free 3' end to enable said probe to function as a primer for a primer extension or ligation reaction, wherein each species of said capture probe comprises a nucleic acid molecule with 5' to 3': (i) a positional domain that corresponds to the position of the capture probe on the array, and (ii) a capture domain; (b) contacting said array with a tissue sample such that the position of a capture probe on the array may be correlated with a position in the tissue sample and allowing nucleic acid of the tissue sample to hybridize to the capture domain in said capture probes; (c) generating DNA molecules from the captured nucleic acid molecules using said capture probes as extension or ligation primers, wherein said extended or ligated DNA molecules are tagged by virtue of the positional domain; (d) optionally generating a complementary strand of said tagged DNA and/or optionally amplifying said tagged DNA; (e) releasing at least part of the tagged DNA molecules and/or their complements or amplicons from the surface of the array, wherein said part includes the positional domain or a complement thereof; and (f) directly or indirectly analyzing the sequence of the released DNA molecules. (Abstract)
Regarding claims 1 and 2 step (a), Frisen teaches a method comprising "The positional domain (or tag) of the capture probe comprises the sequence which is unique to each feature and acts as a positional or spatial marker (the identification tag)" (Pg. 21, Para. 2, ln 5). Frisen teaches a method comprising "capture domain may be specific" (Pg. 65, Para. 2, ln 11-12). Frisen teaches a method comprising "a free 3’ end to … of said capture probe comprises a nucleic acid molecule" (Pg. 7, ln 18 and 20). The free 3’ end of a nucleotide sequence is interpreted to end in a free hydroxyl (-OH) moiety.
Regarding claims 1 and 2 step (b), Frisen teaches a method comprising "oriented to have a free 3' end to enable said probe to function as a primer for a primer extension or ligation reaction" (Pg. 7, ln 18-19). Frisen teaches a method comprising "generating DNA molecules from the captured nucleic acid molecules using said capture probes as extension or ligation primers, wherein said extended or ligated DNA molecules are tagged by virtue of the positional domain" (Pg. 7, ln 29-31). Frisen teaches a method comprising "so-called barcode sequences (or ID tags, defined herein as positional domains)" (Pg. 12, ln 34, and Pg. 13 ln 1).
Regarding claims 1 and 2 step (d), Frisen teaches a method comprising “ligation of … or extension of the nucleic acid, e.g. using an enzyme to incorporate additional nucleotides at the end of the sequence” (Pg. 32, ln 18-20.).
Regarding claim 2 steps (b-e), Frisen teaches a method comprising “the terminal nucleotide of the capture domain could be a reversible terminator nucleotide, which could be included in the capture probe during or after probe synthesis.
Frisen does not explicitly teach the limitations of claims 1 and 2 steps (b-e).
Palluk discloses an oligonucleotide synthesis strategy that uses the template-independent polymerase terminal deoxynucleotidyl transferase (TdT). Each TdT molecule is conjugated to a single deoxyribonucleoside triphosphate (dNTP) molecule that it can incorporate into a primer. After incorporation of the tethered dNTP, the 3' end of the primer remains covalently bound to TdT and is inaccessible to other TdT-dNTP molecules. Cleaving the linkage between TdT and the incorporated nucleotide releases the primer and allows subsequent extension. We demonstrate that TdT-dNTP conjugates can quantitatively extend a primer by a single nucleotide in 10-20 s, and that the scheme can be iterated to write a defined sequence. This approach may form the basis of an enzymatic oligonucleotide synthesizer. (Abstract)
Regarding claims 1 and 2 steps (b-e), Palluk teaches a method comprising “Scheme for two-step oligonucleotide extension using TdT–dNTP conjugates consisting of a TdT molecule site-specifically labeled with a dNTP via a cleavable linker. In the extension step, a DNA primer is exposed to an excess of TdT–dNTP conjugate. Upon incorporation of the tethered nucleotide into the 3′ end of the primer, the conjugate becomes covalently attached and prevents further extensions by other TdT–dNTP molecules... the linkage between the incorporated nucleotide and TdT is cleaved by addition of the cleavage reagent (e.g., …365 nm light…), thereby releasing the primer for subsequent extension. The cycle can be iterated to extend a primer by a defined sequence” (Pg. 646, Figure 1a legend; Figure 1a shown below).
Regarding claim 2 steps (b-d), Palluk teaches a method comprising “reversible terminator' dNTPs” (Pg. 645, Col. 2, Para. 2; Pg. 648, Col. 2, Para. 1).
Regarding claim 4, Palluk teaches a method wherein “Terminal deoxynucleotidyl transferase (TdT) is the only known polymerase whose predominant activity is to indiscriminately add deoxynucleotide triphosphates (dNTPs) to the 3′ end of single stranded DNA1, making it the natural candidate for use in enzymatic oligo synthesis” (Pg. 645, Col. 2, Para. 2). Thus, Palluk teaches a method wherein the polymerase is terminal deoxynucleotidyl transferase or a biologically-active portion thereof.
Regarding claim 5, Palluk teaches a method wherein “addition of … nucleotides to a primer by TdT using 3′ O-nitrobenzyl RTdNTPs” (Pg. 648, Col. 2, Para. 1). Thus, Palluk teaches a method wherein (a) the at least one reversible terminator nucleotide comprises 3'-O-(2-nitrobenzyl)- dATP, 3'-O-(2-nitrobenzyl)-dCTP, 3'-O-(2-nitrobenzyl)-dGTP or 3'-O-(2-nitrobenzyl)-dTTP; or (b) the 3' terminator moiety comprises 2-nitrobenzyl.
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Regarding claim 23, Palluk teaches a method wherein “product was purified (DCC) and tailed with ddTTP to block the strand … from further incorporations” (Pg. 652, Extension of a DNA strand by the sequences 5′-CTAGTCAGCT-3′ and 5′-CCC-3′ using TdT–dNTP conjugates. Para. 1). “tailing with ddTTP” is interpreted as a preparation treatment to prevent incorporation of nucleotides until ready to extend product or probe by nucleotide synthesis. Thus, Palluk teaches a method further comprising prior to step (a), subjecting the tissue sample to ddTTP (dideoxthymidine- triphosphate) termination, wherein subjecting the tissue sample to ddTTP termination comprises contacting the tissue sample with ddTTP and TdT.
Palluk does not explicitly teach the limitations of claims 1 and 2 steps (d-e).
Frenz discloses methods of determining a location of a biological analyte in a biological sample. (Abstract)
Regarding claims 1 and 2 (steps b-e), Frenz teaches a method comprising “a plurality of capture probes immobilized on the substrate, wherein a capture probe comprises a spatial barcode and a capture domain; (b) contacting the biological sample with the substrate” (Para. 6) and “any of the methods described herein, the biological sample comprises a tissue” (Para. 34), Frenz teaches a method comprising “comprises contacting the biological sample with an array comprising two or more analyte binding moieties” (Para. 41). Frenz teaches a method comprising “a capture probe includes an in situ synthesized oligonucleotide” and “the in situ synthesized oligonucleotide includes a barcode sequence” (Para. 416). Frenz teaches a method comprising “A barcode can be added to, for example, a fragment of a deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) sample before or during sequencing” (Para. 130). Frenz teaches a method comprising “Capture probes arrays can be prepared by in situ synthesis. … light-directed combinatorial chemical synthesis on a substrate to selectively synthesize probes … one nucleotide at a time per spot, for many spots simultaneously... linker molecules that have a protecting group on the free end that can be removed by light. UV light is directed through a photolithographic mask to deprotect and activate selected sites with hydroxyl groups that initiate coupling with incoming protected nucleotides that attach to the activated sites” (Para. 539). Frenz teaches a method comprising “A particular barcode can be unique relative to other barcodes” (Para. 129).
Regarding claim 6, Frenz teaches a method wherein “each of the plurality of capture probes comprises a region selected from the group consisting of…a unique molecular identifier” (Para. 12) and “the length of a UMI sequence can be at least about …14…nucleotides or longer” (Para. 405). Thus, Frenz teaches a method wherein the probes further comprise a unique molecular identifier, wherein the unique molecular identifier is at least about 14 nucleotides in length.
Regarding claim 7, Frenz teaches a method wherein “an amplification primer binding site” (Para. 1266; Para. 1269) and “The oligonucleotide primers are of sufficient length to provide for hybridization to complementary genetic material under annealing conditions. The length of the primers generally depends on the length of the amplification domains but will typically be … at least 20 bp… or longer, where the length of the primers will generally range from 18 to 50 bp” (Para. 157). At least 24 nucleotides is interpreted as being comprised within “at least 20 bp … or longer”. “hybridization to complementary genetic material” is interpreted as primer binding site. Thus, Frenz teaches a method wherein the probes further comprise an amplification primer binding site, wherein the amplification primer binding site is at least about 24 nucleotides in length.
(ii) Regarding claim 10, Frenz teaches a method wherein “a spatial barcode (e.g., a nucleic acid sequence that provides information as to the position of the capture probe within a cell or a tissue sample” and “the spatial barcode can be a nucleic acid that has a unique sequence” (Para. 121). Frenz teaches a method wherein “The barcode sequence can act as a spatial barcode and/or as a unique molecular identifier” (Para. 186). Frenz teaches a method wherein “spatial barcode subsequences” (Para. 395) and “a barcode includes two or more sub-barcodes that together function as a single barcode” (Para. 131). “Spatial barcode subsequences” and “sub-barcodes” are interpreted as unique spatial identifier sequences comprised within the spatial barcode. Thus, Frenz teaches a method wherein the spatial barcode domain of a probe comprises a unique spatial identifier sequence.
Regarding claim 11, Frenz teaches a method wherein “spatial barcode subsequence can be about … 16 nucleotides or longer” (Para. 395). “at least about 20 nucleotides” is interpreted as comprising “about 16 nucleotides or longer”. Thus, Frenz teaches a method wherein the spatial identifier sequence comprises at least about 20 nucleotides.
Regarding claim 12, Frenz teaches a method wherein “Barcodes can spatially-resolve molecular components found in biological samples, for example, at single-cell resolution (e.g., a barcode can be or can include a “spatial barcode” and “a barcode includes two or more sub-barcodes that together function as a single barcode. For example, a polynucleotide barcode can include two or more polynucleotide sequences (e.g., sub-barcodes)” (Para. 131). “Two or more sub-barcodes” are interpreted as comprising “four spatial identification domains”. Thus, Frenz teaches a method wherein the spatial identifier sequence comprises at least four spatial identification domains, preferably wherein:(a) each of the at least four spatial identification domains comprise the same number of nucleotides; or (b) at least one of the at least four spatial identifications domains comprise a different number of nucleotides as compared to another spatial identification domain within the same spatial barcode.
Regarding claim 14, Frenz teaches a method wherein “a blocking domain can be incorporated into the capture probe when it is synthesized, or after its synthesis. The terminal nucleotide of the capture domain is a reversible terminator nucleotide (e.g., 3′-O-blocked reversible terminator and 3′-unblocked reversible terminator), and can be included in the capture probe during or after probe synthesis” (Para.422). Thus, Frenz teaches a method wherein the method further comprises, after step (e) and prior to step (f), repeating steps (c) and (d) to extend the spatial barcode domain in each location of the tissue sample such that the spatial barcode domain comprises, at the 3' end, a delimiting domain.
Regarding claim 15, Frenz teaches a method wherein “the feature includes the spatial barcode 802 in combination with a poly(T) capture domain 803” (Para.397; Figure 8). The composition on the bead as shown in Figure 8 is interpreted as having been extended to include a Poly (T) sequence prior to sequencing. Thus, Frenz teaches a method wherein the method further comprises, after step (e) and prior to step (f), extending the spatial barcode domain in each location of the tissue sample such that the spatial barcode domain comprises a polyT domain.
Regarding claim 16-17, Frenz teaches a method wherein “probes… can be prepared by in situ synthesis” and “The process can be repeated, a new mask is applied activating different sets of sites and coupling different bases, allowing different oligonucleotides to be constructed at each site. This process can be used to synthesize hundreds of thousands of different oligonucleotides” (Para. 539); “activating different sets of sites and coupling different bases, allowing different oligonucleotides to be constructed at each site” is interpreted as being able to further extend a probe to any different desired sequences of any desired length such as a 4-6 nucleotide delimiting domain and/or polyT domain prior to collection and sequencing. Thus, Frenz teaches a method wherein the method further comprises, after step (e) and prior to step (f):(i) repeating steps (c) and (d) to extend the spatial barcode domain in each location of the tissue sample such that the spatial barcode domain comprises a delimiting domain; and (ii) extending the spatial barcode domain in each location of the tissue sample such that the spatial barcode domain comprises a polyT domain.
Beechem, Frisen, Palluk and Frenz are considered to be analogous to the claimed invention because they are in the same field of synthetic oligonucleotide and/or spatial analyte detection using synthetic oligonucleotides. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method comprising multiplexed detection and quantification of two or more targets in defined region(s) of a tissue; contacting a sample with at least two probes comprising a target and identification domain; each probe directed to one target; a barcode label that allows for identification; a photocleavable linker where cleavage is inducible at a region interest; collection of a probe domain correlated to a target bound probe; and detection and quantification of target analyte by sequencing as taught by Beechem to incorporate:
the method comprising a probe with a capture domain, identification domain and free 3’-OH; generation of DNA molecules using extension or ligation to the 3’ OH of the capture probe with an enzyme to incorporate additional nucleotides to the 3’ end; also comprising a barcode domain as taught by Frisen and provide a probe with a free 3’ end to be ligate one or more nucleotides to thus extended the probe.
the method comprising a nucleotide-polymerase complex; a nucleotide operably linked to polymerase by a photocleavable linker; incorporation of nucleotide to free 3’ end nucleotide sequence; light inducible cleavage of photocleavable linker and release of polymerase for subsequent extension of the sequence as taught by Palluk and provide a method of extending the free 3’ end of the probe to include a barcode domain one nucleotide at time using nucleotides that are operably linked by a photocleavable linker.
method comprising two or more species of capture probes prepared by light directed combinatorial chemical synthesis selectively ligating a nucleotide one at time at one or more locations utilizing photocleavable linked nucleotides; a spatial barcode and capture domain; in situ synthesis of a barcode; subsequences/sub-barcodes (unique spatial identifier sequences) of a spatial barcode; and a unique barcode can be added before or during sequencing as taught by Frenz and provide a method of spatially profiling target analyte at least two different locations by extending the free 3’ end of each probe to include an in situ synthesized barcode domain with unique spatial identifier sequences followed by quantification by sequencing of the probe to help indicate the abundance of each detected analyte at one or more locations.
Furthermore. the MPEP states "Where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation." (MPEP 2144.05). Thus, it would also be obvious to optimize cleaving the photocleavable linker of the nucleotide-polymerase complexes in claim 1 to cleaving the photocleavable linker of the reversible terminator nucleotides in claim 2 with the same expected result of releasing the reversible 3' moieties and exposing a free 3'-OH moiety on the spatial barcode domain at the at least one location of the tissue sample.
Response to Arguments
Applicant' s arguments filed 10/28/2025 (Pg.11-15) with respect to claim 1-2, 4-7, 10-12, 14-17, 19-20 and 22-24 have been considered but are not persuasive. To clarify some instances argued in the response filed 10/28/2025 see responses to each argument made by Applicant below:
Applicants’ argument: “The cited references do not teach all of the claim limitations” (Pg. 12)… “Beecham does not disclose collecting the probe region containing the Examiner's alleged "spatial barcode domain" (see annotations i and iv illustrating the Examiner's alleged "target-binding domain" and also illustrating the Examiner's site for the claimed 3'OH ligation/ the Examiner's "spatial barcode domain", which Beecham does not disclose collecting. Instead, Beecham only discloses collecting the signal oligonucleotide for analysis.”(Pg. 13)
Response: Applicant's argument filed 10/28/2025 has been fully considered but is not persuasive because, as stated in the revised rejection under 35 U.S.C. 103 (documented above) towards claims 1 and 2, Beechem does teach collecting the probes bound to target analytes in the tissue sample. Also, Beechem teaches collecting a probe comprising a spatial barcode domain (Para. 146). Furthermore, in response to applicant's arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986).
Applicants’ argument: “A person having ordinary skill in the art would not combine the references” (Pg. 14)
Response: Applicant's argument filed 10/28/2025 has been fully considered but is not persuasive. In response to applicant’s argument that there is no teaching, suggestion, or motivation to combine the references, the examiner recognizes that obviousness may be established by combining or modifying the teachings of the prior art to produce the claimed invention where there is some teaching, suggestion, or motivation to do so found either in the references themselves or in the knowledge generally available to one of ordinary skill in the art. See In re Fine, 837 F.2d 1071, 5 USPQ2d 1596 (Fed. Cir. 1988), In re Jones, 958 F.2d 347, 21 USPQ2d 1941 (Fed. Cir. 1992), and KSR International Co. v. Teleflex, Inc., 550 U.S. 398, 82 USPQ2d 1385 (2007). In this case, Beechem, Frisen, Palluk and Frenz are considered to be analogous to the claimed invention because they are in the same field of synthetic oligonucleotide and/or spatial analyte detection using synthetic oligonucleotides. Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method comprising multiplexed detection and quantification of two or more targets in defined region(s) of a tissue; contacting a sample with at least two probes comprising a target and identification domain; each probe directed to one target; a barcode label that allows for identification; a photocleavable linker where cleavage is inducible at a region interest; collection of a probe domain correlated to a target bound probe; and detection and quantification of target analyte by sequencing as taught by Beechem to incorporate:
the method comprising a probe with a capture domain, identification domain and free 3’-OH; generation of DNA molecules using extension or ligation to the 3’ OH of the capture probe with an enzyme to incorporate additional nucleotides to the 3’ end; also comprising a barcode domain as taught by Frisen and provide a probe with a free 3’ end to be ligate one or more nucleotides to thus extended the probe.
the method comprising a nucleotide-polymerase complex; a nucleotide operably linked to polymerase by a photocleavable linker; incorporation of nucleotide to free 3’ end nucleotide sequence; light inducible cleavage of photocleavable linker and release of polymerase for subsequent extension of the sequence as taught by Palluk and provide a method of extending the free 3’ end of the probe to include a barcode domain one nucleotide at time using nucleotides that are operably linked by a photocleavable linker.
method comprising two or more species of capture probes prepared by light directed combinatorial chemical synthesis selectively ligating a nucleotide one at time at one or more locations utilizing photocleavable linked nucleotides; a spatial barcode and capture domain; in situ synthesis of a barcode; subsequences/sub-barcodes (unique spatial identifier sequences) of a spatial barcode; and a unique barcode can be added before or during sequencing as taught by Frenz and provide a method of spatially profiling target analyte at least two different locations by extending the free 3’ end of each probe to include an in situ synthesized barcode domain with unique spatial identifier sequences followed by quantification by sequencing of the probe to help indicate the abundance of each detected analyte at one or more locations.
Claim 9 is/remains rejected under 35 U.S.C. 103 as being unpatentable over Beechem et al. (“Beechem”; US Patent App. Pub. No. US 20170016909 A1 Jan. 19, 2017) in view of Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012), Palluk et al. (“Palluk”; (2018). De novo DNA synthesis using polymerase-nucleotide conjugates. Nature biotechnology, 36(7), 645-650.), and Frenz et al. (“Frenz”; US Patent App. Pub. No. US 20200277664 A1, Sept. 3, 2020, filed on May 5, 2020) as applied to claim 1, and further in view of Beechem et al. (“Beechem ‘19”; US Patent App. Pub. No. US 20190249248 A1, Aug. 15, 2019, filed on Feb. 11, 2019).
The teachings of Beechem, Frisen, Palluk, and Frenz are documented above in the rejection of claims 1-2, 4-7, 10-12, 14-17, 19-20 and 22-24 under 35 U.S.C. 103. Claim 9 depends on claim 1. The combined teachings of Beechem, Frisen, Palluk, and Frenz do not explicitly teach a method wherein the probes comprise, from 5' to 3', the target binding domain, followed by the amplification primer binding site, followed by the unique molecular identifier, followed by the target identification domain, followed by the constant region.
Beechem ‘19 discloses probes, compositions, methods, and kits for simultaneous, multiplexed detection and quantification of protein and/or nucleic acid expression in a user-defined region of a tissue, user-defined cell, and/or user-defined subcellular structure within a cell that are adaptable for use with existing sequencing technologies. (Abstract)
Regarding claim 9, Beechem ‘19 teaches a method wherein “the probe can comprise a target binding domain … The probe further comprises an identifier oligonucleotide. The identifier oligonucleotide can comprise a unique nucleic acid sequence which identifies the target analyte bound to the target binding domain. The identifier oligonucleotide can also comprise a first amplification primer binding site, a second amplification primer binding site, or a unique molecular identifier. The identifier oligonucleotide can comprise any combination of these features. Any of these features can also be flanked by regions comprising constant nucleic acid sequences” (Para. 199). Beechem ‘19 also teaches probes comprising from 5' to 3', the target binding domain, followed by the amplification primer binding site, followed by the unique molecular identifier, followed by the target identification domain (Figure 26 - see figure below).
The constant domain is interpreted as being flanked to the 3’ of the target identification domain. Thus, Beechem ‘19 teaches a method wherein the probes comprise, from 5' to 3', the target binding domain, followed by the amplification primer binding site, followed by the unique molecular identifier, followed by the target identification domain, followed by the constant region.
Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of producing a spatially resolved profile of at least a first target analyte at a location and a at least another target analyte at a another location of a tissue sample using in situ synthesis to extend the probe to comprise a unique barcode based on the spatial location of the analyte detected as taught by Beechem,
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Frisen, Palluk, and Frenz to incorporate the method of a probe comprising 5' to 3', the target binding domain, followed by the amplification primer binding site, followed by the unique molecular identifier, followed by the target identification domain and flanked by a constant region on the 3’ end of the target identification domain as taught by Beechem ‘19 and provide a probe for detecting the abundance and location of analyte in a tissue sample. Doing so would help highlight the distinct domains incorporated into the probe.
Response to Arguments
Applicant's arguments filed 10/28/2025 have been fully considered but they are not
persuasive. Arguments against Beechem, Frisen, Palluk and Frenz on Pg. 16 are not persuasive as discussed above.
Claim 21 is/remains rejected under 35 U.S.C. 103 as being unpatentable over Beechem et al. (“Beechem”; US Patent App. Pub. No. US 20170016909 A1 Jan. 19, 2017) in view of Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012), Palluk et al. (“Palluk”; (2018). De novo DNA synthesis using polymerase-nucleotide conjugates. Nature biotechnology, 36(7), 645-650.), and Frenz et al. (“Frenz”; US Patent App. Pub. No. US 20200277664 A1, Sept. 3, 2020, filed on May 5, 2020) as applied to claim 1, and further in view of Webster et al. (“Webster”; US Patent App. Pub. No. US 20140371088 A1, Dec. 18, 2018).
The teachings of Beechem, Frisen, Palluk, and Frenz are documented above in the rejection of claims 1-2, 4-7, 10-12, 14-17, 19-20 and 22-24 under 35 U.S.C. 103. Claim 21 depends on claim 1. The combined teachings of Beechem, Frisen, Palluk, and Frenz do not explicitly teach wherein the first location of the tissue sample and the at least second location of the tissue sample are no more than about 500 nm in the x and/or y direction and no more than about 1500 nm in the z direction.
Webster discloses compositions and methods for the detection and quantification of individual target molecules in biomolecular samples. In particular, the invention relates to coded, labeled compositions comprising at least two probes hybridized to each other that are capable of binding to and identifying target molecules based on the probes' label codes. Methods of making and using such compositions are also provided. The compositions can be used in diagnostic, prognostic, quality control and screening applications. (Abstract)
Regarding claim 21, Webster teaches a method wherein “The label attachment regions of a reporter or capture probe will vary in size … In various embodiments, a label attachment region can have a length anywhere from 10 nm to 10,000 nm, … more preferably from 100 nm to 1,000 nm. … the label attachment region corresponds closely to the size of a diffraction-limited spot” (Para. 96). The size is interpreted as the length in any direction. No more than about 500 nm or 1500 nm is interpreted as being comprised in the ranges of 10 nm to 10,000 nm and 100 nm to 1,000 nm. Thus, Webster teaches a method wherein the first location of the tissue sample and the at least second location of the tissue sample are no more than about 500 nm in the x and/or y direction and no more than about 1500 nm in the z direction.
Therefore, it would have been obvious to someone of ordinary skill in the art before the effective filing date of the claimed invention to have modified the method of producing a spatially resolved profile of at least a first target analyte at a location and a at least another target analyte at a another location of a tissue sample using in situ synthesis to extend the probe to comprise a unique barcode based on the spatial location of the analyte detected as taught by Beechem, Frisen, Palluk, and Frenz to incorporate the method of a location being no more than 500 nm or 1500 nm in a direction as taught by Webster and provide a probe for detecting the abundance and location of analyte in a tissue sample. Doing so would help highlight the distinct domains incorporated into the probe.
Response to Arguments
Applicant's arguments filed 10/28/2025 have been fully considered but they are not
persuasive. Arguments against Beechem, Frisen, Palluk and Frenz on Pg. 16 are not persuasive as discussed above.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
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Claim 1 is/remains rejected on the ground of nonstatutory double patenting as being unpatentable over claim 1 of U.S. Patent No. 11377689 (“U.S. Patent No. ‘689”; U.S. App. No. 17/476,707) in view of Beechem et al. (“Beechem”; US Patent App. Pub. No. US 20170016909 A1 Jan. 19, 2017), Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012), Palluk et al. (“Palluk”; (2018). De novo DNA synthesis using polymerase-nucleotide conjugates. Nature biotechnology, 36(7), 645-650.), and Frenz et al. (“Frenz”; US Patent App. Pub. No. US 20200277664 A1, Sept. 3, 2020, filed on May 5, 2020). Although the claims at issue are not identical, they are not patentably distinct from each other because the instantly claimed invention is made obvious over the claims of U.S. Patent No. ‘689.
The teachings of Beechem, Frisen, Palluk, and Frenz are documented above in the rejection of claims 1-2, 4-7, 10-12, 14-17, 19-20 and 22-24 under 35 U.S.C. 103.
Claim 1 of U.S. Patent No. ‘689 is drawn to:
1. A method for spatially detecting at least one target analyte in a first location and a second location of a tissue sample comprising: a) contacting the tissue sample with a plurality of nucleic acid probes, wherein each of the nucleic acid probes comprise a target binding domain that binds to the at least one target analyte, wherein the tissue sample [has been] is treated to facilitate binding of the nucleic acid probes to the target analyte; b) collecting the nucleic acid probes, or portions thereof, bound to the at least one target analyte in a first location of the tissue sample under conditions that release the nucleic acid probes, or portions thereof, from the first location of the tissue sample; c) collecting the nucleic acid probes, or portions thereof, bound to the at least one target analyte in a second location of the tissue sample under conditions that release the nucleic acid probes, or portions thereof, from the second location of the tissue sample; d) performing an extension reaction that incorporates at least one nucleic acid sequence that identifies the first location of the tissue sample into each of the nucleic acid probes, or portions thereof, collected in step (b), thereby forming a first plurality of extension products that comprise the nucleic acid probes, or portions thereof, collected in step (b) and the at least one nucleic acid sequence that identifies the first location of the tissue sample; e) performing an extension reaction that incorporates at least one nucleic acid sequence that identifies the second location of the tissue sample into each of the nucleic acid probes, or portions thereof, collected in step (c), thereby forming a second plurality extension products that comprise the nucleic acid probes, or portions thereof, collected in step (c) and the at least one nucleic acid sequence that identifies the second location of the tissue sample; and f) identifying the first plurality of extension products and the second plurality of extension products by sequencing the first plurality of extension products and the second plurality of extension products, thereby spatially detecting the at least one target analyte in the first location of the tissue sample and the second location of the tissue sample.
Claim 1 of U.S. Patent No. ‘689 is made obvious over the instantly claimed invention. One of ordinary skill in the art would have had a reasonable expectation of success given the lack of novelty. It would have been obvious to produce a spatially resolved profile of at least a first target analyte at a location and at least another target analyte at another location of a tissue sample using in situ synthesis to extend the probe to comprise a unique barcode based on the spatial location of the analyte detected according to the limitations of the instant application based on U.S. Patent No. ‘689 in view of Beechem, Frisen, Palluk, and Frenz.
Claim 1 is/remains rejected on the ground of nonstatutory double patenting as being unpatentable over claim 1 of U.S. Patent No. 11473142 (“U.S. Patent No. ‘142”; U.S. App. No. 17/476,712) in view of Beechem et al. (“Beechem”; US Patent App. Pub. No. US 20170016909 A1 Jan. 19, 2017), Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012), Palluk et al. (“Palluk”; (2018). De novo DNA synthesis using polymerase-nucleotide conjugates. Nature biotechnology, 36(7), 645-650.), and Frenz et al. (“Frenz”; US Patent App. Pub. No. US 20200277664 A1, Sept. 3, 2020, filed on May 5, 2020). Although the claims at issue are not identical, they are not patentably distinct from each other because the instantly claimed invention is made obvious over the claims of U.S. Patent No. ‘142.
The teachings of Beechem, Frisen, Palluk, and Frenz are documented above in the rejection of claims 1-2, 4-7, 10-12, 14-17, 19-20 and 22-24 under 35 U.S.C. 103.
Claim 1 of U.S. Patent No. ‘142 is drawn to:
A method comprising: a) collecting a plurality of oligonucleotides from a first location of a tissue sample under conditions that release the plurality of oligonucleotides from the first location of the tissue sample; b) collecting a plurality of oligonucleotides from a second location of the tissue sample under conditions that release the plurality of oligonucleotides from the second location of the tissue sample; c) synthesizing a first plurality of DNA products by performing a synthesis reaction that uses the plurality of oligonucleotides collected in step (a) as templates and incorporates at least one nucleic acid sequence that identifies the first location of the tissue sample into each of the first plurality of DNA products; d) synthesizing a second plurality of DNA products by performing a synthesis reaction that uses the plurality of oligonucleotides collected in step (b) as templates and incorporates at least one nucleic acid sequence that identifies the second location of the tissue sample into each of the second plurality of DNA products; and e) identifying the first plurality of DNA products and the second plurality of DNA products synthesized in step (c) and step (d) by sequencing the first plurality of DNA products and the second plurality of DNA products, thereby spatially detecting the plurality of oligonucleotides collected from the first location of the tissue sample and the plurality of oligonucleotides collected from the second location of the tissue sample.
Claim 1 of U.S. Patent No. ‘142 is made obvious over the instantly claimed invention. One of ordinary skill in the art would have had a reasonable expectation of success given the lack of novelty. It would have been obvious to produce a spatially resolved profile of at least a first target analyte at a location and a at least another target analyte at another location of a tissue sample using in situ synthesis to extend the probe to comprise a unique barcode based on the spatial location of the analyte detected according to the limitations of the instant application based on U.S. Patent No. ‘142 in view of Beechem, Frisen, Palluk, and Frenz.
Claims 1-2, 4, 10, 12, 16 and 22-24 are provisionally rejected on the ground of nonstatutory double patenting as being unpatentable over claims 1-2, 4-6, 12-13, 15, 17-18, 22 and 24-25 of copending Application No. 18/570,190 (U.S. Patent App. Pub. No. 20240279723) claims filed on Aug. 09, 2024 in view of Beechem et al. (“Beechem”; US Patent App. Pub. No. US 20170016909 A1 Jan. 19, 2017), Frisen et al. (“Frisen”; Patent App. Pub. No. WO 2012140224 A1, Oct. 18, 2012), Palluk et al. (“Palluk”; (2018). De novo DNA synthesis using polymerase-nucleotide conjugates. Nature biotechnology, 36(7), 645-650.), and Frenz et al. (“Frenz”; US Patent App. Pub. No. US 20200277664 A1, Sept. 3, 2020, filed on May 5, 2020). Although the claims at issue are not identical, they are not patentably distinct from each other because the instantly claimed invention is made obvious over the claims of copending Application No. 18/570,190.
The teachings of Beechem, Frisen, Palluk, and Frenz are documented above in the rejection of claims 1-2, 4-7, 10-12, 14-17, 19-20 and 22-24 under 35 U.S.C. 103.
Claims 1-2, 4-6, 12-13, 15, 17-18, 22 and 24-25 of copending Application No. 18/570,190 is drawn to:
1. (Original) A method for in situ synthesis of a nucleic acid sequence in a tissue sample, the method comprising: a) contacting the tissue sample with at least one probe, wherein the probe comprises a target-binding domain and a target identification domain, wherein the probe comprises a free 3'-OH moiety, and wherein the target-binding domain binds to at least one target molecule located at a first location of the tissue sample; b) contacting the tissue sample with at least one reversible terminator nucleotide, at least one polymerase, at least one caged chelator-cofactor complex, and at least one unbound caged chelator, wherein the at least one caged chelator-cofactor complex comprises at least one cofactor bound to a caged chelator; wherein the at least one reversible terminator nucleotide comprises the nucleotide operably linked to a cleavable 3' terminator moiety; c) illuminating the first location of the tissue sample with light sufficient to uncage the at least one caged chelator-cofactor complex, thereby releasing the at least one cofactor, thereby activating the at least one polymerase, thereby ligating the at least one reversible terminator nucleotide to the free 3'-OH moiety of the at least one bound probe; d) washing the tissue sample; e) treating the tissue sample under conditions sufficient to cleave the 3' terminator moiety of the at least one reversible terminator nucleotide ligated in step (c), thereby exposing a free 3'- OH moiety on the extended at least one bound probe; and f) repeating steps (b) - (e) until the nucleic acid sequence has been synthesized.
2. (Currently Amended) The method of claim 1, wherein the target-binding domain binds to at least one target molecule at an at least second location of the tissue sample, and wherein the method further comprises repeating steps (b) - (f) at the at least second location, wherein the nucleic acid sequence synthesized at the first location of the tissue sample is different than the nucleic acid sequence synthesized at the at least second location of the tissue sample, wherein the nucleic acid sequence synthesized in the first location of the tissue sample comprises a first spatial barcode domain, and wherein the nucleic acid sequence synthesized at the at least second location of the tissue sample comprises a second spatial barcode domain.
4. (Currently Amended) 'The method of claim 2, wherein the method further comprises: g) collecting the probes bound to target analytes in the tissue sample.
5. (Currently Amended) The method of claim 4, the method further comprising: h) quantifying via sequencing the probes collected in step (g), thereby determining the abundance of the at least two target analytes in the first and the at least second location of the tissue sample, thereby producing a spatially-resolved profile of the abundance of the at least two target analytes.
6. (Currently Amended) The method of claim 1, wherein the polymerase is terminal deoxynucleotidyl transferase or a biologically active fragment thereof.
12. (Currently Amended) The method of claim1, wherein the probes further comprise: i) a unique molecular identifier; ii) an amplification primer binding site; iii) a constant region; or iv) any combination of the preceding.
13. (Currently Amended) The method of claim 2, wherein the first spatial barcode domain comprises a unique spatial identifier sequence specific to the first location of the tissue sample, and/or the second spatial barcode domain comprises a unique spatial identifier sequence specific to the at least second location of the tissue sample.
15. (Currently Amended) The method of claim 13, wherein the spatial identifier sequence comprises at least four spatial identification domains, wherein: i) each of the at least four spatial identification domains comprise the same number of nucleotides; or ii) at least one of the at least four spatial identifications domains comprise a different number of nucleotides as compared to another spatial identification domain within the same spatial barcode.
17. (Currently Amended) The method of claim 15, wherein each of the at least four spatial identification domains comprise the same nucleotide at the 3' terminus.
18. (Currently Amended) The method of claim 2,wherein the method further comprises, after step (f) _i)repeating steps (b) - (e) to extend the spatial barcode domain in each location of the tissue sample such that the spatial barcode domain comprises, at the 3' end, a delimiting domain; and/or ii) extending the spatial barcode domain in each location of the tissue sample such that the spatial barcode domain comprises a polyT domain.
22. (Currently Amended) The method of claim 2, wherein the first location of the tissue sample and the at least second location of the tissue sample are; ii subcellular iii) individually comprise no more than one cell; or iii) individually comprise no more than ten cells.
24. (Currently Amended) The method of claim 1, the method further comprising prior to step (a), subjecting the tissue sample to ddTTP (dideoxthymidine-triphosphate) termination, wherein subjecting the tissue sample to ddTTP termination comprises contacting the tissue sample with ddTTP and TdT.
25. (Currently Amended) The method of claim5, the method further comprising after step (g) and prior to step (h), amplifying the collected probes.
Claims 1-2, 4-6, 12-13, 15, 17-18, 22 and 24-25 of copending Application No. 18/570,190 are made obvious over the instantly claimed invention. One of ordinary skill in the art would have had a reasonable expectation of success given the lack of novelty. It would have been obvious to produce a spatially resolved profile of at least a first target analyte at a location and a at least another target analyte at another location of a tissue sample using in situ synthesis to extend the probe to comprise a unique barcode based on the spatial location of the analyte detected according to the limitations of the instant application based on copending Application No. 18/570,190 in view of Beechem, Frisen, Palluk, and Frenz.
This is a provisional nonstatutory double patenting rejection.
Response to Arguments
Applicant's arguments filed 10/28/2025 have been fully considered but they are not
persuasive. Arguments against Beechem, Frisen, Palluk and Frenz on Pg. 16 are not persuasive as discussed above.
Conclusion of Response to Arguments
In view of the amendments, documented above in this Final Office Action are revised rejections as well as responses to arguments. No claims are in condition for allowance.
Conclusion
THIS ACTION IS MADE FINAL. 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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/KENDRA R VANN-OJUEKAIYE/Examiner, Art Unit 1682
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682