DETAILED ACTION
Notice of Pre-AIA or AIA Status
The present application, filed on or after March 16, 2013, is being examined under the first inventor to file provisions of the AIA .
Information Disclosure Statement
The information disclosure statement (IDS) filed 24 January 2024 is considered, initialed, and attached hereto. Reference Cite No. 157 is lined through and was not considered because it is a duplicate of Reference Cite No. 155, which is considered.
Claim Status
Claims 1, 3, 5, 11, 15, 22, 28-29, 32-33, 45, 52-53, 79-80, 85, 94-95, and 109-110 are pending and under examination.
Claims 2, 4, 6-10, 12-14, 16-21, 23-27, 30-31, 34-44, 46-51, 54-78, 81-84, 86-93, and 96-108 are canceled.
Specification
The use of the terms Cascade blue, Texas red, EMD Millipore, TrueBlack, Biotium, Amersham, BODIPY, RHODAMINE GREEN, OREGON GREEN, ALEXA FLUOR, Marina Blue, Molecular Probes, CLARITY, Thermo Fisher, Mirus Bio, Triton X-100, Tween-20, Abcam, 9°N, New England Biolabs, Ampligase, Epicentre, SplintR, VENT, DEEPVENT, LongAmp, OneTaq, Quick-Load, KlenTaq, REDTaq, Phusion, ArrayScript, MultiScribe, ThermoScript, SuperScript, TAQMAN, SYBR, LightCycler, Cy5, ATTO-TAG, BOBO, BO-PRO, Calcium Crimson, Calcium Green-1, Calcium Orange, Calcofluor, Cascade Yellow, GeneBLAzer, Cy2, Cy3, Cy3.5, Cy5.5, Cy7, ELF, FluorX, Fluoro-Gold, FM, Fura Red, JOJO, JO-PRO, LOLO, LO-PRO, LysoSensor, LysoTracker, Magnesium Green, MitoTracker, POPO, PO-PRO, Rhodamine Red, SNAFL, SNARF, Sodium Green, SpectrumAqua, SpectrumGreen, SpectrumOrange, SpectrumRed, SYTO, SYTOX, TOTO, TO-PRO, YOYO, YO-PRO, and IRDye, which are trade names or a marks used in commerce, has been noted in this application. The terms should be accompanied by the generic terminology; furthermore the terms should be capitalized wherever they appear or, where appropriate, include a proper symbol indicating use in commerce such as ™, SM , or ® following the term.
Although the use of trade names and marks used in commerce (i.e., trademarks, service marks, certification marks, and collective marks) are permissible in patent applications, the proprietary nature of the marks should be respected and every effort made to prevent their use in any manner which might adversely affect their validity as commercial marks.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
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.
Claims 94-95 are 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 94 recites the limitation "the nucleic acid product" in line 2. There is insufficient antecedent basis for this limitation in the claim. Claim 95 is also rejected due to its dependency on claim 94.
Claim Rejections - 35 USC § 102
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 1, 5, 11, 22, 28, 45, 53, 79-80, and 85 are rejected under 35 U.S.C. 102(a)(1) and (a)(2) as being anticipated by Daugharthy et al. (US 2021/0292834, published 23 September 2021, effectively filed 10 October 2018), herein Daugharthy.
Regarding claim 1, Daugharthy teaches a method for processing a biological sample, comprising: (a) contacting the biological sample with a polymer (“A 3D matrix may be formed in situ throughout the cell and tissue sample, such as through the formation of a hydrogel matrix” [0081]), wherein: the polymer comprises one or more functional groups A (“a reactive group on the 3D matrix” [0085]); the biological sample comprises a probe or a product thereof that comprises one or more functional groups B (“The nucleic acids (e.g., RNA molecule, cDNA molecule, primer, or probe) described herein may comprise a functional moiety” [0085]); and the probe comprises a nucleic acid and is configured to directly or indirectly bind to a target analyte in the biological sample (“Identification by hybridization can comprise hybridizing a detection probe to the sequence to be identified” [0076]); (b) reacting the one or more functional groups A with the one or more functional groups B, thereby coupling the polymer to the probe or product thereof the form a polymer conjugate (“The nucleic acids can be linked to the 3D matrix by the functional moiety. The functional moiety can be reacted with a reactive group on the 3D matrix through conjugation chemistry” [0085]); removing unreacted polymer from the biological sample (“the synthetic 3D matrix may be partially or substantially cleared of certain species or classes of biomolecules, such as lipids and proteins, as by use of detergent and/or protease reagents. According to some aspects of the present disclosure, the sample can be cleared using a detergent solution, such as Triton-X or SDS. The detergent may interact with the molecules allowing the molecules to be washed out or removed” [0089], per instant specification [0098], washing is a method of removing unreacted polymer); (d) shrinking or expanding the polymer conjugate, thereby respectively shrinking or expanding the probe or product thereof (“the 3D matrix can be an expanding FISSEQ matrix” [0081]).
Regarding claim 5, Daugharthy teaches the method of claim 1 (see above), wherein the reaction between functional group A and functional group B is a click reaction (“In some embodiments, target molecules and/or index molecules are subjected to amplification reactions” [0107]; “Nucleic acid molecules can be amplified by rolling circle amplification (RCA)” [0108]; “The amplicon may comprise functional linkage groups for tethering to the 3D matrix, such as acrylamide or click-reactive groups” [0109]).
Regarding claim 11, Daugharthy teaches the method of claim 1 (see above), wherein the target analyte is a nucleic acid target analyte, and wherein a primary probe is hybridized to a target sequence in the nucleic acid target analyte (“Methods used to determine the 3D spatial position can comprise […] hybridization” [0076]; “In some embodiments, target molecules and/or index molecules are subjected to amplification reactions” [0107]; “The target molecules may be RNA […] The target molecules may be DNA” [0140]; “The probes may be […] used for nucleic acid amplification described herein” [0152]).
Regarding claim 22, Daugharthy teaches the method of claim 1 (see above), wherein the probe or product thereof comprises a detectable label, or a region that directly or indirectly binds to a detectably labeled probe (“the 3D spatial position can be determined by hybridizing a probe having a detectable label attached thereto to an index sequence” [0076]).
Regarding claim 28, Daugharthy teaches the method of claim 22 (see above), wherein the region in the probe or product thereof comprises a barcode sequence, and wherein the barcode sequence corresponds to an analyte or a portion thereof in the biological sample (“the 3D spatial position can be determined by hybridizing a probe having a detectable label attached thereto to an index sequence co-localized with the target nucleic acid molecule” [0076] the index sequence is considered a barcode sequence). Additionally, it is noted that claim 28 is also anticipated by the quoted portion of the reference meeting the limitation “wherein the probe or product thereof comprises a detectable label” recited in claim 22, since that alternative of claim 22 was not further limited or excluded in claim 28.
Regarding claim 45, Daugharthy teaches the method of claim 1 (see above), wherein shrinking or expanding the polymer conjugate is initiated by contacting the biological sample with a solution, by changing a temperature of the biological sample, and/or by exposing the biological sample to light (“the 3D matrix can be an expanding FISSEQ matrix, such as one comprised substantially of poly(acrylate-co-acrylic acid) (PAA) or Poly(N-isopropylacrylamide) (NIPAM). The matrix comprising NIPAM may be expandable or configured to expand by a change in temperature” [0081]).
Regarding claim 53, Daugharthy teaches the method of claim 1 (see above), wherein the biological sample is a processed or cleared biological sample (“the sample can be cleared” [0089]).
Regarding claim 79, Daugharthy teaches (“a method for processing or analyzing a plurality of nucleic acid molecules of a cell or cell derivative, comprising: (a) generating in the cell or cell derivative a synthetic three-dimensional (3D) matrix” [0005]; “the method further comprises, prior to (a), providing a plurality of precursor nucleic acid molecules, and subjecting the plurality of precursor nucleic acid molecules to nucleic acid amplification to generate the plurality of nucleic acid molecules” [0014]; “probes may be used to synthesize or amplify nucleic acids” [0152] therefore, the plurality of nucleic acid molecules generated by nucleic acid amplification are the product of probes and this generation is done before contacting the sample with the polymer).
Regarding claim 80, Daugharthy teaches the method of claim 1 (see above), wherein the product of the probe is generated in situ in the biological sample (“The nucleic acids in the biological sample will be hybridized with a probe […] The hybridized sequence may be ligated or subjected to an extension reaction to circularize the nucleic acid molecule. Following circularization, the nucleic acid molecule can be subjected to RCA to amplify the amount of nucleic acid and increase the signal. In situ sequencing can be performed on the capture nucleic acid molecule” [0172]).
Regarding claim 85, Daugharthy teaches the method of claim 1 (see above), wherein the product of the probe is a rolling circle amplification (RCA) product (RCP) of a circular or circularizable probe or probe set that hybridizes to a DNA or RNA molecule in the biological sample (“The nucleic acids in the biological sample will be hybridized with a probe or a plurality of probes […]. The probe will be constructed such that it can be circularized, for example a molecular inversion probe or padlock probe. […] The hybridized sequence may be ligated or subjected to an extension reaction to circularize the nucleic acid molecule. Following circularization, the nucleic acid molecule can be subjected to RCA to amplify the amount of nucleic acid” [0172]; “A nucleic acid may comprise deoxyribonucleotides and/or ribonucleotides, or analogs thereof” [0034]).
Therefore, claims 1, 5, 11, 22, 28, 45, 53, 79-80, and 85 are anticipated by Daugharthy.
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.
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.
Claims 3 and 29 are rejected under 35 U.S.C. 103 as being unpatentable over Daugharthy et al. (US 2021/0292834, published 23 September 2021), herein Daugharthy, as applied to claims 1, 5, 11, 22, 28, 45, 53, 79-80, and 85 above.
Regarding claim 3, Daugharthy teaches the method of claim 1 (see 35 U.S.C. 102 rejection above), further comprising initiating polymer-polymer cross-linking to form a polymer network between the polymers (“The 3D matrix may be generated by directing precursors of the 3D matrix in to the biological specimen and subjecting the precursors to crosslinking or polymerization reactions” [0081]). Daugharthy does not explicitly teach that this cross-linking occurs between polymers of the polymer conjugate, which requires that it occurs after the polymer conjugate is formed by coupling the polymer to the probe or product thereof, as recited in claim 1.
However, it would be obvious to modify the order of Daugharthy to contact the biological specimen with a polymer (“directing precursors of the 3D matrix in to the biological specimen” [0081]), then react the functional groups A (“reactive group” [0085]) with the functional groups B (“functional moiety” [0085]) to form a polymer conjugate, then cross-link the polymers of the polymer conjugate to form a polymer network (“subjecting the precursors to crosslinking or polymerization reactions” [0081]), because “selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results” (MPEP §2144.04 IV. C.). Therefore, the invention as a whole of claim 3 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Regarding claim 29, Daugharthy teaches the method of claim 1 (see 35 U.S.C. 102 rejection above), further comprising detecting a signal associated with the probe or product thereof or the target analyte by imaging the biological sample using fluorescent microscopy (“target molecules and/or index molecules are subjected to amplification reactions” [0107]; “The target molecules may be RNA […] The target molecules may be DNA” [0140]; “probes may be […] used for nucleic acid amplification” [0152]; “the spatial origin of each index can be determined via fluorescent in situ sequencing (FISSEQ)” [0100]). Though Daugharthy also teaches expansion of the probe or product thereof (see 35 U.S.C. 102 rejection of claim 1), Daugharthy does not explicitly teach an order of steps such that the step of detecting a signal by imaging occurs after shrinking or expansion of the probe or product thereof.
However, it would be obvious to modify the order of Daugharthy to expand the probe or product thereof (“the 3D matrix can be an expanding FISSEQ matrix” [0081]), then detect a signal associated with the probe or product thereof or the target analyte by imaging the biological sample using fluorescent microscopy ([0076] and [0151] in above paragraph), because “selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results” (MPEP §2144.04 IV. C.). Therefore, the invention as a whole of claim 29 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Claims 15, 32, 94-95, and 109-110 are rejected under 35 U.S.C. 103 as being unpatentable over Daugharthy et al. (US 2021/0292834, published 23 September 2021), herein Daugharthy, as applied to claims 1, 3, 5, 11, 22, 28-29, 45, 53, 79-80, and 85 above, and further in view of Church et al. (in IDS filed 24 January 2024)(US 2019/0194709, published 27 June 2019), herein Church.
Regarding claim 15, Daugharthy teaches the method of claim 11 (see 35 U.S.C. 102 rejection above), wherein the product of the primary probe is a rolling circle amplification (RCA) product (RCP) and nucleotides comprising one or more functional groups B are incorporated into the RCP during RCA (“Nucleic acid molecules can be amplified by rolling circle amplification (RCA)” [0108]; “The amplicon may comprise functional linkage groups for tethering to the 3D matrix […] the functional linkages can be incorporated during amplification using nucleotide analogs” [0109]). Daugharthy also teaches that NIPAM is one possible expandable polymer that can be used in the method (“The matrix comprising NIPAM may be expandable” [0081]). However, Daugharthy does not teach the method wherein the RCP is shrunk. This deficiency is made up for in the teachings of Church.
Regarding claim 15, Church teaches methods of coupling biomolecules to polymers and shrinking or expanding the polymers to respectively shrink or expand the coupled biomolecules (“methods related to processing of a sample so that biomolecules of interest are fixed in a 3-dimensional matrix […] the 3-dimensional matrix can expand or shrink/contract with an external stimulus” [0032]). Church also teaches that the 3-dimensional matrix may be comprised of a NIPAM gel that is able to change volume (i.e. expand or shrink/contract) in response to stimuli (“As used herein, a 3-dimensional matrix may refer to a hydrogel” [0032]; “the hydrogel may be a thermally induced hydrogel. The thermally induced hydrogel may undergo a volume transition in response to thermal stimuli. One example of a thermally induced hydrogel is a NIPAM gel” [0123]).
Regarding claim 32, Daugharthy teaches the method of claim 29 (see 35 U.S.C. 103 rejection above), wherein a signal is detected and wherein the probe or product thereof is expanded. However, Daugharthy does not teach that the expanding happens after the detecting and does not teach shrinking of the probe or product therefore before the detecting. These deficiencies are made up for in the teachings of Church.
Regarding claim 32, Church teaches methods of coupling biomolecules to polymers (as discussed with regard to claim 15) wherein the same polymer can undergo both shrinking and expanding as well as fluorescent imaging (“the hydrogel can be expanded upon an external stimulus […] and a reagent mixture may be flown through after the expansion, and then the hydrogel can be contracted subsequently with an external stimulus” [0129]; “In some embodiments, the method further comprises flowing reagents for fluorescent in situ sequencing (FISSEQ) into the three-dimensional matrix subsequent to said swelling” [0007]). Neither Daugharthy nor Church explicitly teach an order of steps such that the step of detecting occurs after the step of shrinking and the step of expanding occurs after the step of detecting. However, it would be obvious to modify the order of steps of the combination of Daugharthy and Church to shrink the probe or product thereof by shrinking the polymer, then detect the signal, then expand the probe or product thereof by expanding the polymer, because “selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results” (MPEP §2144.04 IV. C.).
Regarding claim 94, Daugharthy teaches the method of claim 85 (see 35 U.S.C. 102 rejection above), further comprising degrading the polymer conjugate after detecting a signal associated with the product of the probe in the biological sample (“Following circularization, the nucleic acid molecule can be subjected to RCA to amplify the amount of nucleic acid and increase the signal. In situ sequencing can be performed on the capture nucleic acid molecule to identify the SMI sequence. The matrix will be dissolved using enzymatic or chemical reactions to break linkages in the matrix” [0172] sentences correspond to generating the product of the probe, detecting a signal, and degrading the polymer conjugate, respectively). However, Daugharthy does not teach that the product of the probe is shrunk or that the degrading step happens after the shrinking step. These deficiencies are made up for in the teachings of Church.
Regarding claim 94, Church teaches methods of coupling biomolecules to polymers and shrinking the polymers to shrink the coupled biomolecules (“methods related to processing of a sample so that biomolecules of interest are fixed in a 3-dimensional matrix […] the 3-dimensional matrix can expand or shrink/contract with an external stimulus” [0032]). Neither Daugharthy nor Church explicitly teach an order of steps such that the step of degrading the polymer occurs after the step of shrinking. However, it would be obvious to modify the order of steps of the combination of Daugharthy and Church to shrink the product of the probe by shrinking the polymer and to detect a signal, then to degrade the polymer, because “selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results” (MPEP §2144.04 IV. C.).
Regarding claim 95, the combination of Daugharthy and Church teach the method of claim 94 (see above). Daugharthy further teaches that the degrading comprises contacting the biological sample with a stripping buffer, changing the temperature of the biological sample, and/or exposing the biological sample to light (“In this example, the proteinase K will be used to dissolve the gel […] For hydrogel embedded matrices, sodium periodate may be used to digest acrylate gels containing DHEBA crosslinker” [0172]; “the nucleic acid material may be linked to the 3D matrix via cleavable linkers and the nucleic acid material can be released from the 3D matrix by cleaving the linkers. The cleavable linkers can be chemically cleavable linkers or photo-cleavable linkers” [0144]).
Regarding claim 109, Daugharthy teaches a method, comprising: (a) generating a rolling circle amplification product (RCP) at a location in a biological sample (“Nucleic acid molecules, such as SMI molecules or endogenous molecules may be amplified using various methods for nucleic acid amplification […] Nucleic acid molecules can be amplified by rolling circle amplification (RCA) […] The resulting population of clonal, or substantially clonal indexing molecules is referred to as the amplicon” [0108]); (b) contacting the biological sample with a polymer (“A 3D matrix may be formed in situ throughout the cell and tissue sample, such as through the formation of a hydrogel matrix” [0081]), wherein: the polymer comprises one or more functional groups A (“a reactive group on the 3D matrix” [0085]), and the RCP comprises one or more functional groups B (“The amplicon may comprise functional linkage groups for tethering to the 3D matrix” [0109]); (c) reacting the one or more functional groups A with the one or more functional groups B, thereby coupling the polymer to the RCP to form a polymer conjugate (“The nucleic acids […] described herein may comprise a functional moiety. The nucleic acids can be linked to the 3D matrix by the functional moiety. The functional moiety can be reacted with a reactive group on the 3D matrix through conjugation chemistry” [0085]); (d) removing unreacted polymer from the biological sample (“the synthetic 3D matrix may be partially or substantially cleared of certain species or classes of biomolecules, such as lipids and proteins, as by use of detergent and/or protease reagents. According to some aspects of the present disclosure, the sample can be cleared using a detergent solution, such as Triton-X or SDS. The detergent may interact with the molecules allowing the molecules to be washed out or removed” [0089], per instant specification [0098], washing is a method of removing unreacted polymer); and (f) detecting a signal associated with the RCP by imaging the biological sample using fluorescent microscopy, thereby detecting the RCP at the location in the biological sample (“target molecules and/or index molecules are subjected to amplification reactions” [0107]; “The target molecules may be RNA […] The target molecules may be DNA” [0140]; “probes may be […] used for nucleic acid amplification” [0152]; “the spatial origin of each index can be determined via fluorescent in situ sequencing (FISSEQ)” [0100]; “FISSEQ library construction […] library construction may involve […] amplifying the original molecule, such as […] rolling circle amplification” [0118]). However, Daugharthy does not teach shrinking the polymer conjugate, thereby shrinking the RCP. This deficiency is made up for in the teachings of Church.
Regarding claim 109, Church teaches methods of coupling biomolecules to polymers and shrinking the polymers to shrink the coupled biomolecules (“methods related to processing of a sample so that biomolecules of interest are fixed in a 3-dimensional matrix […] the 3-dimensional matrix can expand or shrink/contract with an external stimulus” [0032]).
Regarding claim 110, the combination of Daugharthy and Church teach the method of claim 109, and both teach expanding the polymer conjugate, thereby expanding the RCP (“the 3D matrix can be an expanding FISSEQ matrix” Daugharthy [0081]; “the 3-dimensional matrix can expand or shrink/contract with an external stimulus” Church [0032]). Neither Daugharthy nor Church explicitly teach an order of steps such that the step of expanding occurs after the step of detecting. However, it would be obvious to modify the order of steps of the combination of Daugharthy and Church to detect the signal, then expand the probe or product thereof by expanding the polymer, because “selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results” (MPEP §2144.04 IV. C.).
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to perform the simple substitution of shrinking in the method taught by Church for expansion in the method taught by Daugharthy. One of ordinary skill in the art would expect that the results of the substitution would be predictable because Daugharthy and Church both teach methods of analyzing biomolecules by coupling the biomolecules to hydrogels, such as a NIPAM gel, that have their volume changed. Therefore, the invention as a whole of claims 15, 32, 94-95, and 109-110 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Claim 33 is rejected under 35 U.S.C. 103 as being unpatentable over Daugharthy et al. (US 2021/0292834, published 23 September 2021), herein Daugharthy, in view of Church et al. (in IDS filed 24 January 2024)(US 2019/0194709, published 27 June 2019), herein Church, as applied to claims 15, 32, 94-95, and 109-110 above, and further in view of Ishidao et al. ("Solvent concentrations of dimethylsulfoxide-water and 1-propanol-water solutions inside and outside poly (N-isopropylacrylamide) gel" Colloid Polym Sci 272, pages 1313–1316 (1994)), herein Ishidao, and Asano et al. ("Expansion Microscopy: Protocols for Imaging Proteins and RNA in Cells and Tissues" Current Protocols in Cell Biology 80, e56 (2018)), herein Asano.
Regarding claim 33, the combination of Daugharthy and Church teach the method of claim 32 (see 35 U.S.C. 103 rejection above) wherein the polymer conjugate expanded comprises poly(N-isopropylacrylamide) (NIPA or NIPAM) (“the 3D matrix can be an expanding FISSEQ matrix, such as one comprised substantially of poly(acrylate-co-acrylic acid) (PAA) or Poly(N-isopropylacrylamide) (NIPAM)” Daugharthy [0081]; “the hydrogel may be a thermally induced hydrogel. The thermally induced hydrogel may undergo a volume transition in response to thermal stimuli. One example of a thermally induced hydrogel is a NIPAM gel” Church [0123]). However, neither Daugharthy nor Church teaches that the polymer conjugate is expanded in a buffer composition comprising at least or about 50% DMSO, a detergent or surfactant, and a salt. These deficiencies are made up for in the combination of the teachings of Ishidao and Asano.
Regarding claim 33, Ishidao teaches that NIPA gels expand in buffer compositions of >90% DMSO (Fig. 1, the filled circles represent DMSO-water solutions where the DMSO concentration ranges from 0% to 100% on the y-axis, when the swelling ratio V/V0 on the x-axis exceeds 100 swelling occurs, at the highest tick mark on the y-axis corresponding to 90% DMSO the swelling ratio is clearly >100). However, neither Daugharthy, Church, nor Ishidao teach a polymer conjugate being expanded in a buffer composition comprising (1) a detergent or surfactant and (2) a salt. This deficiency is made up for in the teachings of Asano.
Regarding claim 33, Asano teaches a method of expanding a gel for expansion microscopy of a nucleic acid target wherein the expansion buffer comprises a detergent or surfactant and a salt (“To expand the samples, wash three times, each time for 10 min with 0.05× SSCT. The expansion factor can be tuned by altering the salt concentration” page 25, step 25; SSCT, 0.05× recipe, page 35, demonstrates that 0.05× SSCT contains Tween-20, a surfactant and detergent, and SSC (saline-sodium citrate), a salt, and that together these are 0.75% volume of a 0.05× SSCT solution).
In view of Asano’s teaching that the expansion factor can be tuned by altering the salt concentration, one of ordinary skill in the art would be motivated to combine the expansion buffer for expanding gels coupled with nucleic acids for expansion microscopy taught by Asano with the method of analyzing samples by coupling nucleic acids with an expandable gel taught by the combination of Daugharthy and Church (the combination of Daugharthy and Church is obvious as discussed in the 35 U.S.C. 103 rejection of claims 15, 32, 94-95, and 109-110 above). This would improve the method of the combination of Daugharthy and Church by allowing the expansion factor to be tuned by the salt concentration, thereby providing motivation to use the buffer of Asano. One of ordinary skill in the art would have a reasonable expectation of success, because Daugharthy, Church, and Asano teach methods of coupling nucleic acids with expandable gels. Therefore, the combination of Daugharthy, Church, and Asano would be obvious to one of ordinary skill in art prior to the effective filing date of the claimed invention.
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to perform the simple substitution of DMSO in the swelling solution of Ishidao for the nuclease-free water in the buffer taught by Asano in the combination of Daugharthy, Church, and Asano. In Figure 1, Ishidao teaches that both components, DMSO and water, are known to have the same function of causing a NIPA gel to swell when the buffer is primarily water (see Figure 1 when the DMSO concentration of the DMSO-water mixture is <10%) and when the buffer is primarily DMSO (see Figure 1 when the DMSO concentration of the DMSO-water mixture is >90%), so both elements and their function were known in the prior art. One of ordinary skill in the art would expect that the results of the substitution of DMSO for water in the buffer would be predictable in that it would maintain the function of the gel being able to expand in the buffer because Daugharthy and Church both teach the gel being a NIPA gel and Ishidao teaches NIPA gels being expanded in both mixtures that are primarily water or primarily DMSO. Furthermore, the salt and detergent taught by Asano only makes up 0.75% of the solution, so one of ordinary skill in the art would be able combine the salt and detergent with the >90% DMSO taught by Ishidao. Therefore, the invention as a whole of claim 33 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Claim 52 is rejected under 35 U.S.C. 103 as being unpatentable over Daugharthy et al. (US 2021/0292834, published 23 September 2021), herein Daugharthy, as applied to claims 1, 3, 5, 11, 22, 28-29, 45, 53, 79-80, and 85 above, and further in view of Dewal (in IDS filed 24 January 2024)(WO 2020/142490, published 9 July 2020).
Regarding claim 52, Daugharthy teaches the method of claim 1 (see 35 U.S.C. 102 rejection above), wherein the biological sample is a tissue (“A biological sample may be solid matter (e.g., biological tissue” [0045]). However, Daugharthy does not explicitly teach that the tissue is a tissue section. This deficiency is made up for in the teachings of Dewal.
Regarding claim 52, Dewal teaches a method of expansion microscopy (ExM) wherein a tissue section is coupled with a polymer and expanded by expanding the polymer (“The present invention is an expansion upon a method for optical imaging of biological specimens with resolution better than the classical microscopy diffraction limit, based on physically expanding the specimen itself” [057]; “In this method, cultured cells, fixed tissue, or in principle any other type of sample of interest, including biological materials, are infused with a composition […] that results in it becoming embedded in the sample material, and then the composition can be expanded isotropically” [057]; “In an embodiment of this ExM concept, the composition comprises a polyelectrolyte hydrogel” [058]; “The sample may be a thin section of a frozen or paraffin embedded tissue or other biological specimen section” [063]).
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to perform the simple substitution of the tissue section taught by Dewal for the tissue in the method taught by Daugharthy. One of ordinary skill in the art would expect that the results of the substitution would be predictable because Daugharthy and Dewal both teach methods of analyzing biomolecules by coupling the biomolecules to expandable hydrogels. Therefore, the invention as a whole of claim 52 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Conclusion
Claims 1, 3, 5, 11, 15, 22, 28-29, 32-33, 45, 52-53, 79-80, 85, 94-95, and 109-110 are rejected. Claims 2, 4, 6-10, 12-14, 16-21, 23-27, 30-31, 34-44, 46-51, 54-78, 81-84, 86-93, and 96-108 are canceled.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Jeffrey Lawrence Bellah whose telephone number is (571)272-1024. The examiner can normally be reached M-Th, 7:30-5 ET.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Anne Gussow can be reached at (571)272-6047. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
Information regarding the status of published or unpublished applications may be obtained from Patent Center. Unpublished application information in Patent Center is available to registered users. To file and manage patent submissions in Patent Center, visit: https://patentcenter.uspto.gov. Visit https://www.uspto.gov/patents/apply/patent-center for more information about Patent Center and https://www.uspto.gov/patents/docx for information about filing in DOCX format. For additional questions, contact the Electronic Business Center (EBC) at 866-217-9197 (toll-free). If you would like assistance from a USPTO Customer Service Representative, call 800-786-9199 (IN USA OR CANADA) or 571-272-1000.
/JEFFREY BELLAH/Examiner, Art Unit 1683
/ANNE M. GUSSOW/Supervisory Patent Examiner, Art Unit 1683
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