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 4 December 2023 is considered, initialed, and attached hereto.
Election/Restrictions
Applicant’s election without traverse of Group I (claims 1-18), drawn to a method of linking oligonucleotides to cell nucleic acids, in the reply filed on 17 December 2025 is acknowledged.
Claims 19-21 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to nonelected inventions, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 17 December 2025.
Claim Status
Claims 1-21 are pending.
Claims 1-18 are under examination as claims 19-21 are withdrawn from consideration as laid forth above.
Specification
The disclosure is objected to because it contains an embedded hyperlink and/or other form of browser-executable code in [0058]. Applicant is required to delete the embedded hyperlink and/or other form of browser-executable code; references to websites should be limited to the top-level domain name without any prefix such as http:// or other browser-executable code. See MPEP § 608.01.
The use of the terms New England Biolabs Inc., QIAGEN, Vent, Enzymatics, Roche, Illumina, SOLiD, Applied Biosystems, Oxford Nanopore Technologies, Life Technologies, Ion Torrent, and Pacific Biosciences, which are a trade name or a mark 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 Interpretation
Claims 5, 6, and 10 recite the respective optional clauses: “the oligonucleotides […] are optionally released from the hydrogel beads after the sealing”, “the oligonucleotides […] are optionally released from the hydrogel beads before the linking”, and “the linking comprises […] and optionally performing template-dependent extension of the oligonucleotide using the nucleic acid as a template.” Because these clauses are recited as being optional, prior art does not have to teach the limitations of these clauses in order to read on the respective claims.
Claim Rejections - 35 USC § 112(b) - Indefiniteness
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 5-7, 11-12, and 18 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 5 recites the limitation "the sealing" in line 2. There is insufficient antecedent basis for this limitation in the claim. For the purpose of examination, “the sealing” will be interpreted as referring to the step of inserting one or more hydrogel bead into the openings. Claims 6 and 7 are also rejected based on their dependency on claim 5.
Claim 6 recites the limitation "the linking" in line 2. There is insufficient antecedent basis for this limitation in the claim because it is unclear whether “the linking” refers to “oligonucleotides are linked to the hydrogel beads” or the step of “linking one or more oligonucleotides to a nucleic acid” (claim 1, on which claim 6 depends via 5). For the purpose of examination, “the linking” will be interpreted as referring to the step of “linking one or more oligonucleotides to a nucleic acid”.
Claim 11 recites the limitation "the linking reagents" in lines 1-2. There is insufficient antecedent basis for this limitation in the claim.
Claim 12 recites the limitation "the linking reagents" in lines 1-2. There is insufficient antecedent basis for this limitation in the claim.
Claim 18 recites the limitation "the photocleavable cross-linker" in lines 1. There is insufficient antecedent basis for this limitation in the claim. However, claim 17 provides antecedent basis for a photocleavable cross-linker. For the purpose of examination, claim 18 is interpreted as depending on claim 17, not claim 16.
Claim Rejections - 35 USC § 112(d)
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS.—Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
The following is a quotation of pre-AIA 35 U.S.C. 112, fourth paragraph:
Subject to the following paragraph [i.e., the fifth paragraph of pre-AIA 35 U.S.C. 112], a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claim 6 is rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends. Claim 6 claims “the method of claim 5, wherein the oligonucleotides are linked to the hydrogel beads and are optionally released from the hydrogel beads before the linking”. The limitation that “the oligonucleotides are linked to the hydrogel beads” is already present in claim 5. Because the optional clause is not required by the claim, it does not limit the scope of the claim. Therefore, claim 6 fails to further limit the subject matter of claim 5 on which it depends. Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
Claim Rejections - 35 USC § 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)(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, 4-8, 10, 12, and 14-15 are rejected under 35 U.S.C. 102(a)(2) as being anticipated by De Rutte et al. (US 2025/0264477, effectively filed date 14 April 2021), herein De Rutte.
Regarding claim 1, De Rutte teaches an example method comprising loading cells into openings in hollow nanovials (“cells are loaded into nanovials” [0511]); inserting one or more hydrogel beads into the openings, thereby occluding the openings and preventing diffusion of the cells from the openings (“blocking particles […] are seeded into the solution to cap and seal nanovials” [0511]; “the blocking particle may comprise a polymer hydrogel” [0195]; “Nanovial With Bead Blocking” FIG. 15A); lysing the cells in the hollow nanovials (“a lysis buffer is added to the solution that can diffuse through the hydrogel matrix of nanovials and lyse cells within” [0511]); linking one or more oligonucleotides to a nucleic acid from the lysed cells thereby barcoding the nucleic acid, wherein the oligonucleotides are present in the hollow nanovials before the loading, or are introduced into the hollow nanovials by the hydrogel beads (“release molecules such as mRNA from the lysed cells and capture the released molecules on the nanovial and/or blocking particle. Note that at least one of the nanovials or blocking particles comprise barcoded oligonucleotide capture molecules” [0511]).
Regarding claim 1, De Rutte also teaches methods comprising loading cells into openings in hollow nanovials (“loading an antibody-producing cell into the cavity of the nanovial” [0078, 0118]); inserting one or more hydrogel beads into the openings, thereby occluding the openings and preventing diffusion of the cells from the openings (“adding one or more blocking particles to block or reduce a size of the opening of the nanovial cavity” [0080, 0120]; FIG. 17 120 and 122; “the blocking particle may comprise a polymer hydrogel” [0195]); lysing the cells in the hollow nanovials (“lysing the antibody-producing cell” [0088, 0127]; “Lytic agent” FIG. 17); linking one or more oligonucleotides to a nucleic acid from the lysed cells thereby barcoding the nucleic acid, wherein the oligonucleotides are present in the hollow nanovials before the loading, or are introduced into the hollow nanovials by the hydrogel beads (“an oligonucleotide barcode associated with the nanovial, or nanovial label (NL) […] The NL may be located on the nanovial or a separate bead in fluid communication with the nanovial and cells therein” and “Reverse transcription is performed to convert the mRNA from the antibody producing cell […] to cDNA contiguously linked to the NL” [0094, 0129]).
Regarding claim 4, De Rutte teaches the method of claim 1, wherein a majority of the nanovials contain one or zero cells (“the methods involve performing any of the above methods on a plurality of nanovials, each nanovial comprising an antibody-producing cell. In some instances, each nanovial comprises an antibody-producing cell (e.g., a single antibody-producing cell)” emphasis added as each nanovial comprising a single cell means a majority contain one or zero cells [0090]).
Regarding claim 5, De Rutte teaches the method of claim 1, wherein the oligonucleotides are linked to hydrogel beads (“blocking particles optionally comprising barcoded oligonucleotide capture molecules” [0511]; “The NL may be located on the nanovial or a separate bead in fluid communication with the nanovial and cells therein” the hydrogel bead is one such “separate bead” [0094]). As discussed in the claim interpretation section, claim 5 does not require the art to teach releasing the oligonucleotides from the hydrogel beads after the sealing (the sealing is interpreted as the inserting step, see 35 U.S.C. 112(b) rejection of claim 5).
Regarding claim 6, De Rutte teaches the method of claim 5, wherein the oligonucleotides are linked to hydrogel beads (“blocking particles optionally comprising barcoded oligonucleotide capture molecules” [0511]; “The NL may be located on the nanovial or a separate bead in fluid communication with the nanovial and cells therein”, the hydrogel bead is one such “separate bead” [0094]). As discussed in the claim interpretation section, claim 6 does not require the art to teach releasing the oligonucleotides from the hydrogel beads before the linking (the linking is interpreted as the linking one or more oligonucleotides to a nucleic acid, see 35 U.S.C. 112(b) rejection of claim 6).
Regarding claim 7, De Rutte teaches the method of claim 5, wherein the oligonucleotides have a barcode sequence, wherein individual hydrogel beads comprise clonal copies of the oligonucleotides and wherein the barcode sequence linked to individual hydrogel beads are unique such that the barcode distinguishes the hydrogel bead from other hydrogel beads (“an oligonucleotide barcode associated with the nanovial, or nanovial label (NL). The NL may be located on the nanovial or a separate bead in fluid communication with the nanovial and cells therein” the hydrogel bead is one such “separate bead” and the barcode inherently distinguishes the hydrogel bead from other hydrogel beads because the barcode must differ from barcodes on other hydrogel beads in order to fulfill the function of being a barcode associated with the nanovial that the hydrogel bead pairs with [0094]).
Regarding claim 8, De Rutte teaches the method of claim 1, wherein the oligonucleotides have a barcode sequence, wherein individual nanovials comprise clonal copies of the oligonucleotides and wherein the barcode sequence linked to individual nanovials are unique such that the barcode distinguishes the nanovial from other nanovials (“an oligonucleotide barcode associated with the nanovial, or nanovial label (NL). The NL may be located on the nanovial” the barcode distinguishes the nanovial from other nanovials because the barcode must differ from barcodes on other nanovials in order to fulfill the function of being a barcode associated with the nanovial [0094]; “each nanovial may further comprise a unique nanovial-associated barcode or label such that each individual nanovial may be distinguished from other nanovials. Non-limiting examples of nanovial barcodes or labels that are suitable include: unique nucleic acid or oligonucleotide barcodes” [0053]).
Regarding claim 10, De Rutte teaches the method of claim 1, wherein the linking comprises annealing the oligonucleotides to the nucleic acids (“the oligonucleotide barcode that comprises the NL may further comprise an oligo-dT sequence (dT) to capture mRNA from the antibody producing cell” [0094]) and optionally performing template-dependent extension of the oligonucleotide using the nucleic acid as the template (“Reverse transcription is performed to convert the mRNA from the antibody producing cell […] to cDNA contiguously linked to the NL” [0094]).
Regarding claim 12, De Rutte teaches the method of claim 10, wherein the nucleic acids are RNA (“to capture mRNA from the antibody producing cell” [0094]) and the linking reagents comprise a reverse transcriptase (“reverse transcription is performed” [0094]).
Regarding claim 14, De Rutte teaches the method of claim 1, wherein the hollow nanovials comprise dextran and polyethylene glycol (PEG)(“To manufacture the nanovials, a microfluidic droplet generator device is used to form a monodisperse emulsion in an oil phase whereby the internal dispersed phase comprises an aqueous two-phase system. One part of the aqueous two-phase system is a crosslinkable hydrogel precursor such as poly(ethylene glycol) (PEG) or a derivative thereof. The other part of the aqueous two-phase system is a polymer such as dextran” [0200]).
Regarding claim 15, De Rutte teaches the method of claim 1, wherein the hydrogel beads comprise polystyrene, polymethylmethacrylate, polyethylene glycol (PEG) (“blocking particles may be comprised of a polymer selected from the group consisting of: PEG, polystyrene, poly-methylmethacrylate” [0195]).
For the reasons above, claims 1, 4-8, 10, 12, and 14-15 are anticipated by De Rutte.
Claim 9 is rejected under 35 U.S.C. 102(a)(2) as being anticipated by De Rutte et al. (US 2025/0264477, effectively filed date 14 April 2021), herein De Rutte, as applied to claims 1, 4-8, 10, 12, and 14-15 above and as evidenced by Chromium Next GEM Single Cell 5’ HT Reagent Kits v2 (Dual Index) User Guide (downloaded from https://www.10xgenomics.com/support/universal-five-prime-gene-expression/documentation/steps/library-prep/chromium-next-gem-single-cell-5-ht-reagent-kits-v-2-dual-index, last modified 20 April 2022), herein 10x Genomics User Guide.
Regarding claim 9, De Rutte teaches the method of claim 1, further comprising performing nucleotide sequencing of the barcoded nucleic acids (“if labeled with barcoded oligonucleotides (96), the nanovial (and associated cells) can be introduced into a single-cell RNA-sequencing workflow (e.g., 10x Genomics Chromium)” De Rutte [0129]). Though De Rutte does not explicitly state that the method further comprises extracting the barcoded nucleic acids from the nanovials and combining barcoded nucleic acids from different nanovials into a bulk solution prior to sequencing, these are all steps inherent to the 10x Genomics Chromium single-cell RNA-sequencing workflow that De Rutte teaches using for the sequencing step, as evidenced by 10x Genomics User Guide. To introduce this method into a single-cell RNA-sequencing workflow, nanovials are partitioned with gel beads (De Rutte FIG. 28, [0042]; “nanovials […] are directly loaded in a similar way as one would load single cells in these systems for a standard single-cell RNA-seq run, and droplets are formed normally” De Rutte [0476]) to form GEMs (“partitioning thousands of cells into nanoliter-scale Gel Beads-in-emulsion (GEMs)”, 10x Genomics User Guide page 17). 10x Genomics User Guide provides evidence that the workflow comprises extracting the barcoded nucleic acids from the nanovials and combining barcoded nucleic acids from different nanovials into a bulk solution (“GEMs are broken and pooled after GEM-RT” after which “cDNA […] is amplified”, since GEM-RT refers to the reverse transcription reaction, this occurs after the nucleic acids are barcoded by the reverse transcription reaction, 10x Genomics User Guide page 20).
For the reasons above, claim 9 is anticipated by De Rutte as evidenced by 10x Genomics User Guide.
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.
Claim 2 is rejected under 35 U.S.C. 103 as being unpatentable over De Rutte et al. (US 2025/0264477, effectively filed date 14 April 2021), herein De Rutte, as applied to claims 1, 4-8, 10, 12, and 14-15 above, in view of Tkachenko et al. (US 2019/0137481, published 9 May 2019), herein Tkachenko.
Regarding claim 2, De Rutte teaches the method of claim 1 as described in the 35 U.S.C. 102 rejection above. However, De Rutte does not teach that the occluding comprises inducing swelling of the one or more hydrogel bead inserted into the nanovials. This deficiency is made up for in the teachings of Tkachenko.
Regarding claim 2, Tkachenko teaches a method of occluding the openings of microwells by inducing swelling of a hydrogel bead inserted into the opening of the microwell (“one bead can enter each microwell, and once in a microwell, a bead rests on the conical sidewalls, effectively sealing the microwell” [0054], “beads can be made from a variety of materials, including hydrogels” [0057], “a minor reduction in the ionic strength of the medium (by medium exchange), when beads are in wells, will cause beads to swell, tightly sealing the microwells” [0064]).
It would have been obvious to one of ordinary skill in the art to improve the method of De Rutte with the teachings of Tkachenko prior to the effective filing date of the claimed invention. De Rutte teaches that a purpose of the hydrogel beads in the method is to reduce the size of the opening of the nanovial cavity (“adding one or more blocking particles to block or reduce a size of the opening of the nanovial cavity” De Rutte [0080, 0120]), so one of ordinary skill in the art would recognize that De Rutte presents a base method that can be improved by techniques that improve the reduction of the size of the opening of the nanovial cavity that has a bead inserted into the opening. Tkachenko presents a teaching that the size of an opening of being occluded by a hydrogel bead can be further reduced (“tightly sealing the microwells” Tkachenko [0064]) by inducing swelling of hydrogel beads inserted in the openings (“cause the beads to swell” Tkachenko [0064]), thereby representing an improvement that could be applied to the base method of De Rutte. One of ordinary skill in the art would recognize that applying the swelling technique of Tkachenko to the base method of De Rutte would yield predictable results because both methods involve using hydrogel beads, the object of the swelling modification taught by Tkachenko, to block the openings of individual reaction chambers (microwells and nanovials). Therefore, the invention as a whole of claim 2 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 3 and 11 are rejected under 35 U.S.C. 103 as being unpatentable over De Rutte et al. (US 2025/0264477, effectively filed date 14 April 2021), herein De Rutte, as applied to claims 1, 4-8, 10, 12, and 14-15 above, in view of Di Carlo et al. (cited in IDS dated 4 December 2023)(US 2021/0268465, published 2 September 2021), herein Di Carlo.
Regarding claim 3, De Rutte teaches the method of claim 1 as described in the 35 U.S.C. 102 rejection above. However, De Rutte does not teach that the occluding comprises inducing swelling of the nanovials. This deficiency is made up for in the teachings of Di Carlo.
Regarding claim 3, Di Carlo teaches that the swelling of nanovials (dropicles or drop-carrier particles) can be induced (“the matrix of the drop-carrier particle 12 is formed from a material that swells upon a change in environmental conditions” [0149]). Additionally, Di Carlo teaches that inducing the swelling of the nanovials is advantageous because it enables “triggered release of encapsulated reagents stored in the matrix” ([0149]).
Regarding claim 11, De Rutte teaches the method of claim 10 as described in the 35 U.S.C. 102 rejection above and Di Carlo teaches the use of nanovials (“assays such as PCR […] can be carried out in dropicles 10) to link oligonucleotides (Drop-carrier particles 12 functionalized with nucleic acids) to nucleic acids wherein the nucleic acids are DNA (“are mixed in aqueous solution with the appropriate assay reagents (e.g., target DNA)”) and the linking reagents comprise a DNA polymerase (“appropriate assay reagents (e.g., […] polymerases” [0078; FIG. 13A]).
In view of Di Carlo’s teaching that inducing the swelling of nanovials provides the advantageous property of enabling a triggered release of a reagent, it would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to combine the teachings of De Rutte with the teachings of Di Carlo to obtain the claimed invention. Because De Rutte’s method of conducting a biological reaction in hollow nanovials includes lysis and linking steps that utilize various reagents, such as lysis reagents (see 35 U.S.C. 102 rejection of claim 1) and reverse transcriptase (see 35 U.S.C. 102 rejection of claim 12), one of ordinary skill in the art would be motivated to combine the method with Di Carlo’s teaching that reagents for reactions in hollow nanovials can have their release triggered by inducing swelling of the nanovials. One of ordinary skill in the art would have a reasonable expectation of success because the methods of De Rutte and Di Carlo both act on similar nanovials used for isolated biological reactions. Therefore, the invention as a whole of claims 3 and 11 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 13 is rejected under 35 U.S.C. 103 as being unpatentable over De Rutte et al. (US 2025/0264477, effectively filed date 14 April 2021), herein De Rutte, as applied to claims 1, 4-8, 10, 12, and 14-15 above, in view of El-Sagheer et al. (“New strategy for the synthesis of chemically modified RNA constructs exemplified by hairpin and hammerhead ribozymes” Proc Natl Acad Sci U S A, 107(35) 15329-15334 (2010)), herein El-Sagheer.
Regarding claim 13, De Rutte teaches the method of claim 1 wherein linking comprises linking the oligonucleotides to the nucleic acids as described in the 35 U.S.C. 102 rejection above. However, De Rutte does not teach that the linking is done via a click chemistry reaction. This deficiency is made up for in the teachings of El-Sagheer.
Regarding claim 13, El-Sagheer teaches linking oligonucleotides to nucleic acids (other oligonucleotides) via a click chemistry reaction (“individual RNA oligonucleotides are assembled by automated solid-phase synthesis, purified by HPLC then chemically ligated by click chemistry to produce much larger molecules” page 1, column 2, paragraph 2).
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 click chemistry reaction taught by El-Sagheer for the capture of released mRNA molecules from lysed cells by oligonucleotide capture molecules in the method of De Rutte (see 35 U.S.C. 102 rejection of claim 1). One of ordinary skill in the art would be able to substitute the click chemistry reaction of El-Sagheer for the capture reaction of De Rutte and would expect that the results of the substitution would be predictable because the click chemistry reaction is used for the same purpose of connecting two oligonucleotides together as El-Sagheer teaches and because the click chemistry reaction is compatible with aqueous media (“The CuAAC reaction was chosen for RNA ligation owing to its very high speed, efficiency, orthogonality with functional groups present in nucleic acids and compatibility with aqueous media” El-Sagheer page 1, column 2, paragraph 2) and the cavity of the nanovial may be filled with aqueous media (“in some cases, the cavity of the nanovial comprises an aqueous fluid disposed therein” De Rutte [0005]). Therefore, the invention as a whole of claim 13 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 and 17-18 are rejected under 35 U.S.C. 103 as being unpatentable over De Rutte et al. (US 2025/0264477, effectively filed date 14 April 2021), herein De Rutte, as applied to claims 1, 4-8, 10, 12, and 14-15 above, in view of Norris et al. (cited in IDS dated 4 December 2023)(“Photodegradable Polyacrylamide Gels for Dynamic Control of Cell Functions”, ACS Appl. Mater. Interfaces 13, 5929-5944 (2021)), herein Norris.
Regarding claim 15, De Rutte teaches the method of claim 1, wherein the hydrogel beads comprise polystyrene, polymethylmethacrylate, polyethylene glycol (PEG) (“blocking particles may be comprised of a polymer selected from the group consisting of: PEG, polystyrene, poly-methylmethacrylate” [0195]). However, De Rutte does not teach hydrogel beads comprising polyacrylamide, silica, alginate, nylon, or agarose.
Regarding claim 15, Norris teaches a “polyacrylamide hydrogel” (Abstract) and teaches that “cross-linked polyacrylamide hydrogels are commonly used in biotechnology and cell culture applications due to advantageous properties, such as the precise control of material stiffness and the attachment of cell adhesive ligands” (Abstract).
Regarding claims 17 and 18, Norris teaches a polyacrylamide hydrogel formed from polymerizing acrylamide and a photocleavable cross-linker wherein the photocleavable cross-linker is ortho-nitrobenzyl (o-NB) bis-acrylate (“photo-degradable polyacrylamide hydrogel networks are produced by copolymerizing acrylamide and a photocleavable ortho-nitrobenzyl (o-NB) bis-acrylate cross-linker” Abstract).
In view of Norris’s teachings that polyacrylamide hydrogels are advantageous in that they allow for the attachment of cell adhesive ligands, it would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to combine the teachings of De Rutte with the teachings of Norris to obtain the claimed invention. Because De Rutte’s method may utilize a blocking particle wherein “the blocking particle is a polymer hydrogel” and “the blocking particle is coated with one or more affinity agents” (De Rutte [0007]), one of ordinary skill in the art would be motivated to combine the method with Norris’s teaching that polyacrylamide hydrogels can have cell adhesive ligands, a type of affinity agent, attached to the hydrogel. One of ordinary skill in the art would have a reasonable expectation of success because the hydrogel of De Rutte and hydrogel of Norris are both used for attachment of affinity groups and would thus be interchangeable. Therefore, the invention as a whole of claims 15 and 17-18 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 16 is rejected under 35 U.S.C. 103 as being unpatentable over De Rutte et al. (US 2025/0264477, effectively filed date 14 April 2021), herein De Rutte, as applied to claims 1, 4-8, 10, 12, and 14-15 above, in view of Wu et al. (cited in IDS)(US Patent 11,180,750, issued 23 November 2021), herein Wu.
Regarding claim 16, De Rutte teaches the method of claim 1, as described in the 35 U.S.C. 102 rejection above. However, De Rutte does not teach hydrogel beads comprising polyacrylamide that is cross-linked with Bis-acrylylcystamine. This deficiency is made up for in the teachings of Wu.
Regarding claim 16, Wu teaches a nucleic acid reaction method using a hydrogel bead (“the hydrogel beads provide a microenvironment for controlled reactions of long DNA fragments” col 8 lines 55-56) that comprises polyacrylamide that is cross-linked with Bis-acrylylcystamine (“one embodiment includes a bead including a hydrogel polymer” col 4 lines 61-62, “examples of hydrogel polymers, which may include one or more crosslinkers, include but are not limited to […] acrylamide, N,N’-bis(acrolyoyl)cystamine, […] or combinations thereof” col 5 lines 13-36, “reversible crosslinker may be N,N’-bis(acryloyl)cystamine, a reversible crosslinker for polyacrylamide gels” col 6 lines 26-27).
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 hydrogel beads taught by Wu for the hydrogel beads in the method of De Rutte (see 35 U.S.C. 102 rejection of claim 1). One of ordinary skill in the art would be able to substitute the hydrogel beads of Wu for the hydrogel beads in the method of De Rutte and would expect that the results of the substitution would be predictable because both hydrogel beads are compatible with nucleic acid reactions. Therefore, the invention as a whole of claim 16 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
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.
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/JEFFREY BELLAH/Examiner, Art Unit 1683
/ANNE M. GUSSOW/Supervisory Patent Examiner, Art Unit 1683