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 .
Status of the Application
Claims 1-17 and 44 are pending and under examination
Claims 18-19, 23, and 27 are withdrawn.
Applicant’s election without traverse of Group I, which includes Claims 1-17 and 44 in the reply filed on 05/11/2026 is acknowledged.
Claim(s) 18-19, 23, and 27 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected aptamer detection probe set, there being no allowable generic or linking claim. Election was made in the reply filed on 05/11/2026.
Claim Rejections - 35 USC § 112(b)
The following is a quotation of 35 U.S.C. 112(b):
(b) CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim(s) 13 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 13 recites “extending the ligated first oligonucleotide and second oligonucleotide and the hybridized third oligonucleotide to generate first and second complementary strands comprising 5′ and 3′ adapters for sequencing.” The phrase does not identify which oligonucleotide(s) function as primers or templates. It is also unclear how the first and second complementary strands are generated.
For purposes of examination only, and to facilitate a complete analysis of the claim, the Examiner interprets claim 13 as requiring third oligonucleotide to act as primer and extended using ligated first oligonucleotide and second oligonucleotide as template. This interpretation is adopted solely for examination and does not resolve the lack of clarity in the claim language.
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.
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.
Terbrueggen et al., Zhang et al., and Gold et al.
Claim(s) 1-9 and 16-17 is/are rejected under 35 U.S.C. 103 as being unpatentable over Terbrueggen et al. (US9745616B2, filed May 17th 2012) in view of Zhang et al. (US20190185933A1, EFD: Dec 20th 2018) and Gold et al. (PLoS One. 2010;5(12):e15004. (2010)).
Regarding claim 1, Terbrueggen discloses a method of nucleic acid detection, comprising: contacting the nucleic acids with a mixture of first probes, wherein a first complementary region of each first probe of the mixture hybridizes to a first region of the individual nucleic acid and first probes in the mixture are coupled to an affinity tag; contacting the individual nucleic acid with a second probe to hybridize a second complementary region of the second probe to a second region of the individual nucleic acid and wherein the second probe comprises a nonhybridizing region extending from the complementary region, the nonhybridizing region comprising an identification sequence uniquely identifying for the individual nucleic acid, wherein the first complementary region and the second complementary region uniquely hybridize to the individual nucleic acid; capturing a first probe of the mixture via binding of the affinity tag to an affinity tag binder to capture the individual nucleic acid and the second probe hybridized to the second region of the individual nucleic acid, wherein the first probe is in the subset coupled to the affinity tag; and detecting the identification sequence of the captured second probe. (e.g. method for direct detection of nucleic acid target using a target specific probe pair. The probe pair consists of a Capture Probe which carries 3′ biotin capture handle and a pair of Chemical Ligation Dependent Probe Amplification (CLPA) probes, which can be chemically ligated into 1 probe [ Scheme 1 : Chemical ligation reaction, columns 43 and 44] and carries 5’ unique identifier barcode [Fig 10 shown below]. Both probes hybridize to the same target nucleic acid. The biotin label at the 3′ end of the capture probes is used to attach the complexes to streptavidin-coated immobile phase. The probes are amplified and sequenced [Abstract and Fig. 1 (shown below)].)
PNG
media_image1.png
990
787
media_image1.png
Greyscale
PNG
media_image2.png
1103
878
media_image2.png
Greyscale
Terbrueggen does not disclose contacting analytes of a sample with a plurality of aptamers under conditions that permit analyte-aptamer complexes to form, wherein different aptamers of the plurality of aptamers have specific affinity for respective different analytes of the analytes; and detecting the analytes by detecting aptamers of the analyte-aptamer complexes. And that only a subset of the first probes in the mixture are coupled to an affinity tag.
Zhang discloses a nucleic acid enrichment method employing positive-selection Probes and negative selection probe Sinks that are thermodynamically competitive. Zhang further teaches that Probes maybe either directly functionalized with a biotin or hybridized to a universal oligonucleotide functionalized with a biotin and collected using streptavidin-coated magnetic beads. In some embodiments, Sinks are not functionalized and compete with Probes for binding, thus controlling which target nucleic acids are collected [¶0006-0007, ¶0022 and Fig 1]. Zhang also teaches optimization of Probes/Sinks sequence and stoichiometry to improve enrichment [¶0091].
Gold discloses contacting a sample with a mixture of different aptamers selected to bind respective protein analytes, thus forming aptamer-protein complexes, and subsequently detecting the proteins by detecting the corresponding aptamers through hybridization to sequence specific complementary DNA probes. Gold teaches the detected aptamer population reflects the quantity of respective proteins in the sample [abstract and Fig 7 A-H].
As of the application’ s effective filing date, it would have been prima facie obvious to a person of ordinary skill in the art to modify Terbrueggen’s method by including a non-affinity tagged competing fraction of capture probes because Zhang teaches that functionalized and non-functionalized competitive probes maybe used together with optimized relative composition enable rare analyte recovery [abstract]. Such modification would predictably reduce capture and detection of selected highly abundant analytes while retaining Terbrueggen’s separate barcode reporter probes. It further would have been obvious to modify Terbrueggen’s method to aptamer of Gold because Gold teaches aptamers are nucleic acids that bind to specific analytes and are detected by sequence specific hybridization. The combination would therefore provide a predictable method for detecting analytes by affinity capturing and barcode detecting their corresponding aptamers while attenuating recovery of aptamers associated with highly abundant analytes. See KSR International Co. v. Teleflex Inc., 550 U.S. 398, 415-421, USPQ2d 1385, 1395 — 97 (2007) (MPEP § 2143).
Regarding claim 2, Terbrueggen does not specify the distance between capture probe and CLPA probe. However, Terbrueggen teaches separately select target specific probe binding regions [Fig 10]. One of ordinary skill in the art would have recognized that such region could be selected as either continuous or noncontinuous depending on sequence accessibility and probe design constraints. Selecting noncontinuous regions separated by at least one nucleotide would have been a routine and predictable probe placement that maintain independent hybridization function of each probe.
Regarding claim 3, Terbrueggen discloses affinity tag is biotin and the affinity tag capture molecule is streptavidin. [Fig 10]
Regarding claim 4, Terbrueggen discloses second probe comprises priming site for amplification.[Fig 10]
Regarding claim 5, Terbrueggen discloses the primers comprise a first primer that binds to a first primer binding region of the nonhybridizing region and a second primer that binds to a second primer binding region of the nonhybridizing region, wherein the first primer binding region and the second primer binding region flank the identification sequence. (e.g. upon chemical ligation of CLPA probe set Fig 10 and Fig 1, the probe comprises primer binding sites at both 3’ and 5’ ends of the CLPA probe (e.g. X and Y) and the unique identifier barcode/spacer locates between the primer binding sites).
Regarding claim 6, Terbrueggen discloses the first primer comprises a first sequencing primer and the second primer comprises a second sequencing primer such that the amplification product comprises the first sequencing primer and the second sequencing primer. (e.g. Fig 1 above shows both X and Y primer binding sites at both ends of amplification product)
Regarding 7, Terbrueggen discloses detecting the identification sequence of the captured second probe comprises sequencing the amplification product. [column 1, line 67]
Regarding 8, Terbrueggen discloses generating a detectable assay output corresponding to the identification sequence of the detected target nucleic acid, such as an electrophoretic peak [Fig 1], mass spectrometry, or microarray signal associate with target specific ligation product [column 3, lines 57-64].
Gold teaches detected aptamer sequence corresponds to identify the respective analyte bounded by aptamer [abstract and Fig 7 A-H].
As of the application’ s effective filing date, it would have been prima facie obvious to a person of ordinary skill in the art to generate or display an indication identifying the aptamer correspond to the detected barcode/identifier sequence because reporting the sequence and quantity of a detected target is predictable use of the assay and would permit user to interpret the detection data. The generated indication constitutes a notification related to the individual aptamer base on detection the identification sequence. See KSR International Co. v. Teleflex Inc., 550 U.S. 398, 415-421, USPQ2d 1385, 1395 — 97 (2007) (MPEP § 2143).
Regarding claim 9, Terbrueggen discloses separating the captured second probe from the captured first probe and the captured individual aptamer before detecting the identification sequence. (e.g. Amplification of the target may also include turnover of the ligation product, in which the ligation product has a lower or comparable affinity for the template or target nucleic acid than do the separate ligation probes. Thus, upon ligation of the hybridized probes, the ligation product is released from the target, freeing the target to serve as a template for a new ligation reaction [column 9, lines 45-53].)
Regarding claim 16, Terbrueggen discloses removing uncaptured probes before detecting the identification sequence. [Fig 1]
Regarding claim 17, Zhang teaches providing a mixture containing functionalized positive selection probe and nonfunctionalized competitive sinks, wherein the nonfunctionalized sinks lack the affinity tag and compete with the functionalized probes for binding to the target nucleic acid. As discussed in claim 1, it would have been obvious to modify Terbrueggen’s affinity tag capture probe to include nonaffinity competitive sinks. Accordingly, complexes with affinity tag are retained by the affinity tag binder and complexes without affinity tags are washed away/removed. Hence, the uncaptured probes include probes of the mixture that are outside the affinity tag subset and are not coupled with affinity tag.
Terbrueggen et al., Zhang et al., Gold et al., and Larson et al.
Claim(s) 10 is/are rejected under 35 U.S.C. 103 as being unpatentable over Terbrueggen et al. (US9745616B2, filed May 17th 2012) in view of Zhang et al. (US20190185933A1, EFD: Dec 20th 2018), Gold et al. (PLoS One. 2010;5(12):e15004. (2010)), and Larson et al. (US10144962B2, EFD: June 24th 2017)
Regarding claim 10, Terbrueggen does not disclose ligating an oligonucleotide to an end of the captured second probe and extending the ligated oligonucleotide before detecting the identification sequence. However, it would have been prima facie obvious to a person of ordinary skill in the art to ligate an oligonucleotide adapter/universal sequences to an end of the captured second probe as needed for specific sequencing techniques because adapter ligation and template switch extension were well established techniques for converting captured nucleic acid molecules into amplifiable and sequenceable libraries (e.g. the techniques are disclosed by Larson et al. [column 6, lines 21-25 and column 2, lines 60-63]). Applying these known library preparation techniques to process Terbrueggen’s probes for downstream amplification and sequencing allow identification of sequence without changing the underlying aptamer detection chemistry.
Terbrueggen et al., Zhang et al., Gold et al., and Khodakov et al.
Claim(s) 11-13 is/are rejected under 35 U.S.C. 103 as being unpatentable over Terbrueggen et al. (US9745616B2, filed May 17th 2012) in view of Zhang et al. (US20190185933A1, EFD: Dec 20th 2018), Gold et al. (PLoS One. 2010;5(12):e15004. (2010)), and Khodakov et al. (Adv. Drug Deliv. Rev. 105 (2016): 3-19.).
Regarding claim 11, Terbrueggen does not disclose using the second probe as template for extension ligation of first oligonucleotide and second oligonucleotide.
Khodakov discloses Illumina Truseq ligation-assisted target enrichment method. Truseq involves hybridizing first and second oligonucleotides to regions flanking a target nucleic acid sequence, extending the first oligonucleotide by polymerase synthesis using target nucleic acid as template until the extended first oligonucleotide adjoins the second oligonucleotide, and subsequently ligating the two oligonucleotides. The ligated product then amplified [Fig 12a shown below].
PNG
media_image3.png
603
439
media_image3.png
Greyscale
As of the application’ s effective filing date, it would have been prima facie obvious to a person of ordinary skill in the art to apply Truseq extension ligation technique to Terbrueggen’s sequencing workflow of the second probe because extension ligation retains the identification sequence of the second probe. Applying Illumina Truseq would therefore predictably permit sequence amplification and detection of the identification sequence carried by second probe while utilizing well established hybridization, polymerase extension, and ligation techniques. See KSR International Co. v. Teleflex Inc., 550 U.S. 398, 415-421, USPQ2d 1385, 1395 — 97 (2007) (MPEP § 2143).
Regarding claim 12, Khodakov does not discloses that Illumina Truseq uses affinity tag for the oligonucleotide.
Terbrueggen discloses using affinity tag for enriching or manipulation of the ligated products.
As of the application’ s effective filing date, it would have been prima facie obvious to a person of ordinary skill in the art to couple an affinity tag to the second oligonucleotide in Illumina Truseq method for recovering/enriching the ligated products. Additionally, a skilled artisan would have known to select an orthogonal different affinity tag /binder pair so that the newly ligate product could be selectively recovered without recapturing the original template complex. See KSR International Co. v. Teleflex Inc., 550 U.S. 398, 415-421, USPQ2d 1385, 1395 — 97 (2007) (MPEP § 2143).
Regarding claim 13, Khodakov discloses hybridizing a third oligonucleotide to the ligated first oligonucleotide and second oligonucleotide and extending the ligated first oligonucleotide and second oligonucleotide and the hybridized third oligonucleotide to generate first and second complementary strands comprising 5′ and 3′ adapters for sequencing. (e.g. PCR primer correspond to the claimed third oligonucleotide and Khodakov discloses PCR amplification after extension ligation [fig 12a shown above]).
Terbrueggen et al., Zhang et al., Gold et al., and Beechem et al.
Claim(s) 14-15 is/are rejected under 35 U.S.C. 103 as being unpatentable over Terbrueggen et al. (US9745616B2, filed May 17th 2012) in view of Zhang et al. (US20190185933A1, EFD: Dec 20th 2018), Gold et al. (PLoS One. 2010;5(12):e15004. (2010)), and Beechem et al. (US20190249248A1, EFD: Feb 11th 2019).
Regarding claim 14, Terbrueggen does not disclose cleaving at least a portion of the nonhybridizing region comprising the identification sequence from the captured second probe before detecting the identification sequence.
Beechem discloses a probe comprises a target binding domain comprising a nucleic acid sequence that is complementary to a target nucleic acid. In the upper panel, the probe hybridizes to the target nucleic acid. In the lower panel, a UV photo-cleavable linker located between the target binding domain and the identifier oligonucleotide is cleaved, releasing the identifier oligonucleotide [Fig 11 shown below].
PNG
media_image4.png
1122
822
media_image4.png
Greyscale
As of the application’ s effective filing date, it would have been prima facie obvious to a person of ordinary skill in the art to incorporate Beechem’s cleavable linker into the second probe to enable the release of the identification sequence. Releasing the identification sequence would predictably remove the aptamer-protein binding complex from downstream processing, hence providing a smaller and simpler to handle nucleic acid templates for amplification and detection while retaining the information identifying the aptamer. See KSR International Co. v. Teleflex Inc., 550 U.S. 398, 415-421, USPQ2d 1385, 1395 — 97 (2007) (MPEP § 2143).
Regarding claim 15, Beechem discloses ligating a cleaved portion of the captured second probe to an end of an adapter after the cleaving. (e.g. (2) release the identifier oligonucleotide; (3) collecting the released identifier oligonucleotide; (4) ligating to the released identifier oligonucleotide at least one nucleic acid adapter [¶0005]).
Terbrueggen et al. and Gold et al.
Claim(s 44) is/are rejected under 35 U.S.C. 103 as being unpatentable over Terbrueggen et al. (US9745616B2, filed May 17th 2012) in view of Gold et al. (PLoS One. 2010;5(12):e15004. (2010)).
Regarding claim 44, Terbrueggen discloses a method of nucleic acid detection, comprising:
contacting an individual nucleic acid with a first reporter probe that hybridizes to a first region of the individual nucleic acid, wherein the first reporter probe comprises a first nonhybridizing region, and a second reporter probe that hybridize to a second region of the individual nucleic acid, wherein the second reporter probe comprises a second nonhybridizing region, the second nonhybridizing region comprising a identification sequence uniquely identifying for the individual aptamer; ligating ends of the first identification sequence and the second identification sequence to one another to generate ligated reporter probes; capturing ligated reporter probes using an affinity tag coupled to the first reporter probe or the second reporter probe; and detecting the first identification sequence and the second identification sequence via amplification of the captured ligated reporter probes to detect the individual aptamer.[Fig 1 below]
PNG
media_image2.png
1103
878
media_image2.png
Greyscale
Terbrueggen does not disclose nucleic acids are aptamers and that there is an additional identification sequence in the first probe.
Gold discloses contacting a sample with a mixture of different aptamers selected to bind respective protein analytes, thus forming aptamer-protein complexes, and subsequently detecting the proteins by detecting the corresponding aptamers through hybridization to sequence specific complementary DNA probes. Gold teaches the detected aptamer population reflects the quantity of respective proteins in the sample [abstract and Fig 7 A-H].
As of the application’ s effective filing date, it would have been prima facie obvious to a person of ordinary skill in the art to modify Terbrueggen’s method to aptamer of Gold because Gold teaches aptamers are nucleic acids that bind to specific analytes and are detected by sequence specific hybridization. The combination would therefore provide a predictable method for detecting analytes by affinity capturing and barcode detecting their corresponding aptamers while attenuating recovery of aptamers associated with highly abundant analytes. Additionally, it would have been obvious to provide each of Terbrueggen’s probe with a respective copy of target specific identifier sequence so that successful ligation products would contain two target identifier sequences. Because ligation only occurs when both probes hybridize to the same target, detecting both same identifiers would confirm that target specific hybridization event occurred, reducing false positive from nonspecific hybridization or unintended ligations. See KSR International Co. v. Teleflex Inc., 550 U.S. 398, 415-421, USPQ2d 1385, 1395 — 97 (2007) (MPEP § 2143).
Conclusion
No claims are allowed
Any inquiry concerning this communication or earlier communications from the examiner should be directed to Khai Quynh Tien Pham whose telephone number is (571)272-6998. The examiner can normally be reached M-T, 9-4 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, Heather Calamita can be reached at (571) 272-2876. 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.
/KHAI QUYNH TIEN PHAM/Examiner, Art Unit 1684
/JEREMY C FLINDERS/Primary Examiner, Art Unit 1684
Prosecution Insights