DETAILED ACTION
Prosecution Reopened
In view of the appeal brief filed on 21 January 2026, PROSECUTION IS HEREBY REOPENED. A new ground of rejection is set forth below.
To avoid abandonment of the application, appellant must exercise one of the following two options:
(1) file a reply under 37 CFR 1.111 (if this Office action is non-final) or a reply under 37 CFR 1.113 (if this Office action is final); or,
(2) initiate a new appeal by filing a notice of appeal under 37 CFR 41.31 followed by an appeal brief under 37 CFR 41.37. The previously paid notice of appeal fee and appeal brief fee can be applied to the new appeal. If, however, the appeal fees set forth in 37 CFR 41.20 have been increased since they were previously paid, then appellant must pay the difference between the increased fees and the amount previously paid.
A Supervisory Patent Examiner (SPE) has approved of reopening prosecution by signing below:
/WU CHENG W SHEN/Supervisory Patent Examiner, Art Unit 1682
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 .
Priority
This application 16/770,889 is a national stage of international application no. PCT/US2018/064616 filed December 7, 2018, which claims the benefit of US provisional application no.: 62/596,337 filed December 8, 2017.
Claim status
Claims 1, 2, 7, 9, 10, 24, 25, 36, 38, and 48-53 are pending and currently under examination.
Drawings
The drawings were received on 26 January 2023. These drawings are acceptable.
Claim Interpretation
Attention is directed to MPEP 904.01 [R-08.2012].
The breadth of the claims in the application should always be carefully noted; that is, the examiner should be fully aware of what the claims do not call for, as well as what they do require. During patent examination, the claims are given the broadest reasonable interpretation consistent with the specification. See In re Morris, 127 F.3d 1048, 44 USPQ2d 1023 (Fed. Cir. 1997). See MPEP § 2111 - § 2116.01 for case law pertinent to claim analysis.
It is noted with particularity that narrowing limitations found in the specification cannot be inferred in the claims where the elements not set forth in the claims are linchpin of patentability. In re Philips Industries v. State Stove & Mfg. Co, Inc., 186 USPQ 458 (CA6 1975). While the claims are to be interpreted in light of the specification, it does not follow that limitations from the specification may be read into the claims. On the contrary, claims must be interpreted as broadly as their terms reasonably allow. See Ex parte Oetiker, 23 USPQ2d 1641 (BPAI, 1992). In added support of this position, attention is directed to MPEP 2111 [R-11.2013], where, citing In re Prater, 415 F.2d 1393, 1404-05, 162 USPQ 541, 550-51 (CCPA 1969), is stated:
The court explained that “reading a claim in light of the specification, to thereby interpret limitations explicitly recited in the claim, is a quite different thing from ‘reading limitations of the specification into a claim,’ to thereby narrow the scope of the claim by implicitly adding disclosed limitations which have no express basis in the claim.” The court found that applicant was advocating the latter, i.e., the impermissible importation of subject matter from the specification into the claim.
Additionally, attention is directed to MPEP 2111.01 [R-01.2024], wherein is stated:
II. IT IS IMPROPER TO IMPORT CLAIM LIMITATIONS FROM THE SPECIFICATION
“Though understanding the claim language may be aided by explanations contained in the written description, it is important not to import into a claim limitations that are not part of the claim. For example, a particular embodiment appearing in the written description may not be read into a claim when the claim language is broader than the embodiment.” Superguide Corp. v. DirecTV Enterprises, Inc., 358 F.3d 870, 875, 69 USPQ2d 1865, 1868 (Fed. Cir. 2004).
Attention is also directed to MPEP 2111.02 II [R-07.2022]. As stated herein:
II. PREAMBLE STATEMENTS RECITING PURPOSE OR INTENDED USE
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The claim preamble must be read in the context of the entire claim. The determination of whether preamble recitations are structural limitations or mere statements of purpose or use "can be resolved only on review of the entirety of the [record] to gain an understanding of what the inventors actually invented and intended to encompass by the claim" as drafted without importing "'extraneous' limitations from the specification." Corning Glass Works, 868 F.2d at 1257, 9 USPQ2d at 1966. If the body of a claim fully and intrinsically sets forth all of the limitations of the claimed invention, and the preamble merely states, for example, the purpose or intended use of the invention, rather than any distinct definition of any of the claimed invention’s limitations, then the preamble is not considered a limitation and is of no significance to claim construction. Shoes by Firebug LLC v. Stride Rite Children’s Grp., LLC, 962 F.3d 1362, 2020 USPQ2d 10701 (Fed. Cir. 2020) (The court found that the preamble in one patent’s claim is limiting but is not in a related patent); Pitney Bowes, Inc. v. Hewlett-Packard Co., 182 F.3d 1298, 1305, 51 USPQ2d 1161, 1165 (Fed. Cir. 1999). See also Rowe v. Dror, 112 F.3d 473, 478, 42 USPQ2d 1550, 1553 (Fed. Cir. 1997) ("where a patentee defines a structurally complete invention in the claim body and uses the preamble only to state a purpose or intended use for the invention, the preamble is not a claim limitation")… (Emphasis added)
Attention is directed to MPEP 2111 [R-10.2019]. As stated therein:
During patent examination, the pending claims must be "given their broadest reasonable interpretation consistent with the specification." The Federal Circuit’s en banc decision in Phillips v. AWH Corp., 415 F.3d 1303, 1316, 75 USPQ2d 1321, 1329 (Fed. Cir. 2005) expressly recognized that the USPTO employs the "broadest reasonable interpretation" standard:
The Patent and Trademark Office ("PTO") determines the scope of claims in patent applications not solely on the basis of the claim language, but upon giving claims their broadest reasonable construction "in light of the specification as it would be interpreted by one of ordinary skill in the art." In re Am. Acad. of Sci. Tech. Ctr., 367 F.3d 1359, 1364[, 70 USPQ2d 1827, 1830] (Fed. Cir. 2004). Indeed, the rules of the PTO require that application claims must "conform to the invention as set forth in the remainder of the specification and the terms and phrases used in the claims must find clear support or antecedent basis in the description so that the meaning of the terms in the claims may be ascertainable by reference to the description." 37 CFR 1.75(d)(1). (Emphasis added).
Attention is directed to MPEP 2173.04 [R-10.2019]. As stated therein:
Breadth of a claim is not to be equated with indefiniteness. In re Miller, 441 F.2d 689, 169 USPQ 597 (CCPA 1971); In re Gardner, 427 F.2d 786, 788, 166 USPQ 138, 140 (CCPA 1970) ("Breadth is not indefiniteness."). A broad claim is not indefinite merely because it encompasses a wide scope of subject matter provided the scope is clearly defined. But a claim is indefinite when the boundaries of the protected subject matter are not clearly delineated and the scope is unclear. For example, a genus claim that covers multiple species is broad, but is not indefinite because of its breadth, which is otherwise clear. But a genus claim that could be interpreted in such a way that it is not clear which species are covered would be indefinite (e.g., because there is more than one reasonable interpretation of what species are included in the claim). (Emphasis added)
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
This application currently names joint inventors. In considering patentability of the claims the examiner presumes that the subject matter of the various claims was commonly owned as of the effective filing date of the claimed invention(s) absent any evidence to the contrary. Applicant is advised of the obligation under 37 CFR 1.56 to point out the inventor and effective filing dates of each claim that was not commonly owned as of the effective filing date of the later invention in order for the examiner to consider the applicability of 35 U.S.C. 102(b)(2)(C) for any potential 35 U.S.C. 102(a)(2) prior art against the later invention.
Holding and Rationale
Claim 1 is rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6).
Regarding steps a), b), c), e), f), and g) recited in claim 1, Shah et al. (2016) teaches “Identifying the spatial organization of tissues at cellular resolution from single-cell gene expression profiles is essential to understanding biological systems. Using an in situ 3D multiplexed imaging method, seqFISH, we identify unique transcriptional states by quantifying and clustering up to 249 genes in 16,958 cells to examine whether the hippocampus is organized into transcriptionally distinct subregions. We identified distinct layers in the dentate gyrus corresponding to the granule cell layer and the subgranular zone and, contrary to previous reports, discovered that distinct subregions within the CA1 and CA3 are composed of unique combinations of cells in different transcriptional states. In addition, we found that the dorsal CA1 is relatively homogeneous at the single cell level, while ventral CA1 is highly heterogeneous. These structures and patterns are observed using different mice and different sets of genes. Together, these results demonstrate the power of seqFISH in transcriptional profiling of complex tissues” (See Abstract).
Shah et al. (2016) teaches in Figure 1. Overview of Sequential Barcode FISH in Brain Slices.
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Figure 1. Overview of Sequential Barcode FISH in Brain Slices (A) A coronal section from a mouse brain was mounted on a slide and imaged in all boxed areas. Each image was taken at 603 magnification. (B) Example of barcoding hybridizations from one cell in a field from (A). The same points are re-probed through a sequence of 4 hybridizations (numbered). The sequence of colors at a given location provides a barcode readout for that mRNA (‘‘barcode composite’’). The genes represented by these barcodes are identified through referencing a lookup table abbreviated in (D) and quantified to obtain single-cell expression levels. In principle, the maximum number of transcripts that can be identified with this approach scales to Fn, where F is the number of fluorophores and n is the number of hybridizations. Error correction adds another round of hybridization. (C) Serial smHCR is an alternative detection method in which 5 genes are quantified in each hybridization and this process is repeated n times. Serial hybridization scales as F*n. (D) Schematic for multiplexing 125 genes in single cells. 100 genes are multiplexed in 4 hybridizations by seqFISH barcoding. This barcode scheme is tolerant to loss of any round of hybridization in the experiment. 25 genes are serially hybridized 5 genes at a time by 5 rounds of hybridization. Each number represents a color channel in smHCR. As a control, 5 genes are measured by both double rounds of smHCR as well as barcoding in the same cell. (E) SmHCR amplifies signal from individual mRNAs. After imaging, DNase strips the smHCR probes from the mRNA, enabling rehybridization on the same mRNA (step a). The ‘‘color’’ of an mRNA can be modulated by hybridizing probes that trigger HCR polymers labeled with different dyes (step b). Signals from the mRNAs are amplified following hybridization by adding the complementary hairpin pair (step c). The DNase smHCR cycle is repeated on the same mRNAs to construct a predefined barcode over time.
The aspect of performing a form of in situ hybridization on mouse brain tissue is deemed to fairly suggest limitations of claims 9 and 10.
Similar to the teachings by Shah et al. (2016), Lubeck et al (2014) teach that “the mRNAs in cells are barcoded by sequential rounds of hybridization, imaging and probe stripping (Fig. 1a and Supplementary Fig. 1). As the transcripts are fixed in cells, the corresponding fluorescent spots remain in place during multiple rounds of hybridization and can be aligned to read out a fluorophore sequence. This sequential barcode is designed to uniquely identify an mRNA.”
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Figure 1 | Sequential barcoding. (a) Schematic of sequential barcoding. In each round of hybridization, 24 probes are hybridized on each transcript, imaged and then stripped by DNase I treatment. The same probe sequences are used in different rounds of hybridization (hyb), but probes are coupled to different fluorophores. (b) Composite four-color FISH data from three rounds of hybridizations on multiple yeast cells. Twelve genes are encoded by two rounds of hybridization, with the third hybridization using the same probes as hybridization 1. The boxed regions are magnified in the bottom right corner of each image. Spots colocalizing between hybridizations are detected (as outlined in insets) and have their barcodes extracted. Spots without colocalization are due to nonspecific binding of probes in the cell as well as mishybridization. The number of instances of each barcode can be quantified to provide the abundances of the corresponding transcripts in single cells.
Regarding step d) recited in claim 1, Lawson et al. (2012) teach that “The urea wash was tested at 0, 1, 2, 4 and 8 mol -1 with NaCl at 0.9 mol -1 (Kourilsky et al. 1970; Simard et al. 2001; Soe et al. 2011)”. (See bridging paragraph, pages 265-266).
In view of the above presentation, it would have been quite obvious to one of ordinary skill in the art at the time of the invention, to have adapted the sequencing method of mRNAs to employ either formamide or urea as a denaturing agent and wherein the target nucleic acid is within the cell. In view of the well-developed state of the art, said ordinary artisan would have been amply motivated and would have had a most reasonable expectation of success.
In view of the above presentation and in the absence of convincing evidence to the contrary, claim 1 is rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6).
Claim(s) 2, 7, 24, 25, and 53 are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 2016/0369329 A1 (Cai et al.).
See above for the basis of the rejection as it pertains to the disclosure of Shah et al., Lubeck et al., and Lawson et al.
Neither Shah et al., Lubeck et al., nor Lawson et al., have been found to explicitly teach the use of a bridge probe, to what the bridge probe binds, nor specify the number of targets.
Cai et al., at paragraphs [0039] and [0041], teach:
[0039] In one aspect, disclosed herein is a composition comprising a plurality of primary probes, a first plurality of bridge probes, and first plurality of readout probes. (Emphasis added)
[0041] In some embodiments, each bridge probe in the first plurality of bridge probes comprises a binding sequence that specifically binds to all or a part of the first overhang sequence of a primary probe of the plurality of primary probes, and one or more readout binding targets connected in series and linked to the binding sequence. (Emphasis added)
Cai et al., at paragraphs [0006] and [0036], teach:
In some embodiments, the present invention provides methods for detecting multiple targets, e.g., transcripts or DNA loci, in a cell through a sequential barcoding scheme that permits multiplexing of different targets.
[0036] In some embodiments, a plurality of detectably labeled oligonucleotides target two or more nucleic acids (“targets”).
The aspect of there being any values that is “more” than 1 is deemed to fairly encompass “three or more readout binding targets” as is specified in claim 25.
Cai et al., in paragraphs [0042] and [0043], teach of using readout probes and how they can be linked to a “signal moiety”.
Cai et al., at paragraphs [0320] and [0348] teach of using a single type of fluorophore for labeling a set of probes. As stated at paragraph [0348]:
During a round of hybridization, each transcript is targeted by a set of multiple, for example, 24 FISH probes, labeled with a single type of fluorophore.
The above showing is deemed to fairly suggest limitations of claim 53.
In view of the above presentation, it would have been quite obvious to have employed the combination of primary probes and bridge probes for as disclosed, such probes, in combination of readout probes, enable one to detect the presence of a target nucleic acid via sequential hybridization.
In view of the above presentation and in the absence of convincing evidence to the contrary, claims 2, 7, 24, 25, and 53 are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 2016/0369329 A1 (Cai et al.).
Claims 9, 10, and 51 are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 2015/0099269 A1 (Ma), US 2004/0091880 A1 (Wiebusch et al.), and US 2002/0102543A1 (Friedrick et al.).
See above for the basis of the rejection as it pertains to the disclosure of Shah et al., Lubeck et al., and Lawson et al.
Neither Shah et al., Lubeck et al., nor Lawson et al., have been found to explicitly teach detection of a target nucleic acid in an intact cell as recited in claims 9, 10 and 51.
Ma, at paragraph [0002], teaches:
[0002] The presence or localization of specific DNA sequences in human chromosomes can be detected in chemically fixed cells by In Situ Hybridization (ISH), a method based on complementary base-pairing between the target sequence and an oligonucleotide probe that carries a detectable tag (e.g., a fluorescent dye). (Emphasis added)
While Ma teaches performing in situ hybridization of fixed cells, it is not clear if the cells are intact.
Weibusch et al., in paragraph [0070, teach:
[0070] The term hybridisation refers to the phenomenon that single strand nucleic acids or parts thereof are forming pairs with complementary or partly complementary single nucleic acid strands. In situ hybridisation refers to hybridisation under conditions maintaining the cell substantially intact. (Emphasis added)
While Weibusch et al. do teach performing in situ hybridization, they have not been found to teach detecting sequences in an intact mouse embryonic stem cell.
Friedrich et al., at paragraph [0019], teach:
[0019] The current invention relates to novel polynucleotides which are expressed in mouse embryonic stem cells ("ES cells") and which provide unique tools for gene discovery, diagnostic gene expression analysis, cross species hybridization analysis, and for genetic manipulations using a variety of techniques known to those skilled in the art, like, for example, antisense inhibition, gene targeting, etc.
Friedrich et al., at paragraph [0098], teach:
[0098] The level of expression of genes can also be assayed by detecting and measuring the transcription of such genes. For example, RNA from a cell type or tissue known, or suspected to express any of the genes of the current invention can be isolated and tested utilizing hybridization or PCR techniques (e.g., northern or RT PCR) such as those described, above. Such analyses may reveal both quantitative and qualitative aspects of the expression pattern of the respective gene, including activation or inactivation of gene expression. In situ hybridization using suitably radioactively or enzymatically labeled forms of the described polynucleotide sequences can also be used to assess expression patterns in vivo. (Emphasis added)
In view of the above presentation, it would have been quite obvious to one of ordinary skill in the art at the time of the invention to have modified the methods of Shah et al., Lubeck et al., and Lawson et al., whereby one is detecting target nucleic acids in intact human cells as well as a mouse embryonic stem cell (Friedrich et al.).
In view of the above presentation and in the absence of convincing evidence to the contrary, claims 9, 10, and 51 are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 2015/0099269 A1 (Ma), US 2004/0091880 A1 (Wiebusch et al.), and US 2002/0102543 A1 (Friedrich et al.).
Claim 36 and 38 are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 6,020,135 (Levine et al., Date of Patent 2/1/2000).
See above for the basis of the rejection as it pertains to the disclosures of Shah et al., Lubeck et al., and Lawson et al.
Neither Shah et al., Lubeck et al., nor Lawson et al., have been found to teach the length of the probes.
Levine et al., at column 4, first full paragraph, teach:
(5) Probes according to the present invention may be labeled or unlabeled, tethered to another substance or in solution, synthetically made or isolated from nature. Probes can be nucleic acids, either RNA or DNA, which contain naturally occurring nucleotide bases or modified bases. The probes may contain normal nucleotide bonds or peptide bonds. Oligonucleotide probes may be of any length which provides meaningful specificity of hybridization. Thus probes may be as small as 10 nucleotides, and preferably they are between 12 and 30 nucleotides in length. However, oligonucleotide probes may be significantly longer, in the range of 30 to 100 nucleotides, 100 to 500 nucleotides or 500 to 2000 nucleotides. (Emphasis added)
In view of the above presentation and in the absence of convincing evidence to the contrary, it would have been quite obvious to one of ordinary skill in the art at the time of the method to have selected probes that comprise 10 to 17 nucleotides as such are well within the ranges of lengths of probes disclosed by Levine et al.
In view of the well-developed state of the art, said ordinary artisan would have been quite motivated and would have also had a most reasonable expectation of success.
In view of the above presentation and in the absence of convincing evidence to the contrary, claims 36 and 38 are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 6,020,135 (Levine et al.).
Claims 48-50 are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 2003/0073150 A1 (Woerner et al.).
See above for the basis of the rejection as it pertains to the disclosures of Shah et al., Lubeck et al., and Lawson et al.
Neither Shah et al., Lubeck et al., nor Lawson et al., have been found to teach using formamide, much less concentrations of same.
Werner et al., at paragraph [0103], teach:
[0103] The DNA in the tissues and the DNA probes are denatured either independently prior to hybridization or simultaneously. In general the denaturation is carried out by placing the tissues or DNA probes into a hybridization solution preferably containing a denaturing agent such as formamide and heating from 70.degree. C. to 95.degree. C. Preferably the concentration of formamide is from about 30% to about 70%, more preferably about 40% to about 60%. (Emphasis added)
In view of the above presentation it would have been quite obvious to one of ordinary skill in the art at the time of the invention to have used formamide as a denaturant, and to have used same in concentrations ranging from about 30% to about 60%.
In view of the well-developed state of the art, said ordinary artisan would have been quite motivated and would have also had a most reasonable expectation of success.
In view of the above presentation and in the absence of convincing evidence to the contrary, claims 48-50 are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 2003/0073150 A1 (Woerner et al.).
Claim(s) 52 is/are rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 2006/0110735 A1 (Heinzel et al.).
See above for the basis of the rejection as it pertains to the disclosures of Shah et al., Lubeck et al., and Lawson et al.
Neither Shah et al., Lubeck et al., nor Lawson et al., have been found to teach of Rlim mRNA being a target for detection.
Heinzel et al., in claim 20, claim a kit for the detection of RLIM mRNA. The fact that RLIM mRNA is to be detected speaks directly to RLIM mRNA being a target nucleic acid.
In view of the above presentation it would have been quite obvious to one of ordinary skill in the art at the time of the invention to have designed the method of sequential hybridization to have as its target a known nucleic acid, including that of RLIM mRNA.
In view of the well-developed state of the art said ordinary artisan would have been amply motivated ad would have had a most reasonable expectation of success.
In view of the above presentation and in the absence of convincing evidence to the contrary, claim 52 is rejected under 35 U.S.C. 103 as being unpatentable over Shah et al. (2016) (In Situ Transcription Profiling of Single Cells Reveals Spatial Organization of Cells in the Mouse Hippocampus, Neuron, 2016 Oct 19;92(2):342-357. doi: 10.1016/j.neuron.2016.10.001) in view of Lubeck et al (2014) (Single-cell in situ RNA profiling by sequential hybridization, Nat Methods., 2014 Apr;11(4):360-1. doi: 10.1038/nmeth.2892) and Lawson et al. (2012) (Dimethyl formamide-free, urea-NaCl fluorescence in situ hybridization assay for Staphylococcus aureus, Lett Appl Microbiol., 2012 Mar;54(3):263-6. doi: 10.1111/j.1472-765X.2011.03197.x. Epub 2012 Jan 6) as applied to claim 1 above, and further in view of US 2006/0110735 A1 (Heinzel et al.).
Conclusion
Objections and/or rejections which appeared in the prior Office action and which have not been repeated hereinabove have been withdrawn.
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure.
US 2002/0107640 A1 (Ideker et al.), in paragraph [0109] teaches performing hybridization of mRNA with a hybridization buffer that comprises 50% formamide.
US 2002/0042095 A1 (Adler et al.), teaches preforming hybridization reactions with mRNA where the formamide concentration is “less than 50%”.
US 2003/0087279 A1 (Shao et al.), at paragraph [0081], teaches using either formamide or urea in a hybridization buffer where one is detecting mRNA.
US 2002/0081668 A1 (Friedrich et al.), in paragraph [0019] teaches of gene expression analysis of mousse embryonic stem cells.
US 2017/0220733 A1 (Zhuang et al.), in paragraphs [0175], [0193] teach of using readout probes.
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/Bradley L. Sisson/Primary Examiner, Art Unit 1682
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