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 .
Please note: The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action.
Interview Summary
Applicant’s summary of the Interview conducted on March 11, 2026, submitted on 3/24/2026, is acknowledged.
Claim Status
In the most recent amendment to the claims, submitted on 1/23/2026, claim 79 has been cancelled and claim 84 is new.
Claims 1-2, 7, 12-14, 21, 25-27, 30, 39, 43, 46-49, 57-58, and 84 are pending and being examined on the merits.
Drawings
Applicant’s submission of replacement drawings that do not contain color on 1/23/2026 is acknowledged. The drawings submitted on 1/23/2026 are accepted.
Claim Objections
The objection to claim 49 is withdrawn in light of Applicant’s amendment to the claim.
Claim Interpretation
Claim 1 is directed to a first encoding probe that comprises “an interrogatory region for interrogating a region of interest”. The specification states that the interrogatory region can be within one or both arms of the padlock probe (paragraph [0235]). Therefore, for purposes of examination, the interrogatory region can be a sequence that is complementary to, and hybridizable, to the target nucleic acid.
Claim 7 is directed to the method of claim 1 “wherein the first encoding probe comprises one or more ribonucleotides and/or the one or more second encoding probes comprise one or more ribonucleotides”. Given the use of the word “comprising” in the claim, the broadest reasonable interpretation of this limitation is that the probes contain 1 or more ribonucleotides and can additionally contain deoxyribonucleotides.
Claim 27 is directed to the method of claim 1 “wherein the one or more second encoding probes and/or the first encoding probe collectively comprise one or more hybridization barcode sequences that correspond to the target nucleic acid”. The specification describes the hybridization barcode region in paragraph [0233] as follows: “The barcode region may comprise barcode sequences, e.g., one or more hybridization barcode sequences (e.g., for detection by FISH, such as single molecule FISH (smFISH)), that can be detected in sequential hybridization cycles to decode the target nucleic acid or sequence thereof”. For purposes of examination, “hybridization barcode sequence” is interpreted to mean a barcode is a sequence that can serve as identification of a particular target nucleic acid via hybridization of barcode specific probes. Additionally, according to the specification, the hybridization barcode sequence is a type of barcode sequence and can be the same sequence as the “amplifiable barcode sequence” (cited in claim 30; see paragraph [0233] of the instant specification).
Claim Rejections - 35 USC § 103
Withdrawn:
The rejection of claims 1-2, 7, 12, 14, 21, 25-26, 27, 30, 39, 43, 46-49, 57, and 79 under 35 U.S.C. 103 as being unpatentable over Nilsson (Nilsson et al., WO 2019/068880 A1) in view of Glezer (Glezer et al., US 2022/0042083 A1; EFD of 8/6/2020) and Pinard (Pinard et al., US 2021/0403992 A1; EFD 6/29/2020) is withdrawn in light of Applicant’s amendments to the claims.
The rejection of claim 13 under 35 U.S.C. 103 as being unpatentable over Nilsson (Nilsson et al., WO 2019/068880 A1) in view of Glezer (Glezer et al., US 2022/0042083 A1; EFD of 8/6/2020) and Pinard (Pinard et al., US 2021/0403992 A1; EFD 6/29/2020) as applied to claims 1-2, 7, 12, 14, 21, 25-26, 27, 30, 39, 43, 46-49, 57, and 79 above, and further in view of Zhang (Zhang et al., Chem. Soc. Rev., 2020) is withdrawn in light of Applicant’s amendments to the claims.
The rejection of claim 58 under 35 U.S.C. 103 as being unpatentable over Nilsson (Nilsson et al., WO 2019/068880 A1) in view of Glezer (Glezer et al., US 2022/0042083 A1; EFD of 8/6/2020) and Pinard (Pinard et al., US 2021/0403992 A1; EFD 6/29/2020) as applied to claims 1-2, 7, 12, 14, 21, 25-26, 27, 30, 39, 43, 46-49, 57, and 79 above, and further in view of Wu (Wu et al., Communications Biology, 2018; cited on IDS of 9/19/2023) is withdrawn in light of Applicant’s amendments to the claims.
New (Necessitated by Amendments):
Claims 1-2, 7, 12, 14, 21, 25-27, 30, 39, 43, 46-49, 57, and 84 are rejected under 35 U.S.C. 103 as being unpatentable over Nilsson (Nilsson et al., WO 2019/068880 A1) in view of Glezer (Glezer et al., US 2022/0042083 A1; EFD of 8/6/2020) and Pinard (Pinard et al., US 2021/0403992 A1; EFD 6/29/2020).
Regarding claim 1 and 84: Nilsson teaches a method of analyzing a biological sample comprising contacting the biological sample with a plurality of encoding probes comprising a first encoding probe that is capable of hybridizing to a first target sequence in a target nucleic acid in the biological sample (claim 1a; Abstract). Nilsson teaches that the first encoding probe is circularizable and comprises an interrogatory region for interrogating a region of interest in the first target sequence (claim 1a; pg 6, ln 11-29, pg 7, ln 19-25, pg 47 ln 21-24, and Figure 1A). Nilsson teaches circularizing the first encoding probe to generate a circularized first encoding probe (claim 1b; pg 6, ln 11-29). Nilsson teaches generating a rolling circle amplification (RCA) product of the circularized first encoding probe (claim 1e; pg 7, ln 4-5) and detecting a signal associated with the RCA product (claim 1f; pg 7, ln 6-7).
Nilsson does not teach that there are one or more second encoding probes contacted with the biological sample, which are circular or circularizable (claim 1a). However, use of multiple circular or circularizable probes for targeting a nucleic acid molecule in a biological sample is known in the art, as taught by Glezer.
Glezer teaches a method of detecting target nucleic acids such as RNA in a cell in situ using barcoded oligonucleotide padlock probes (paragraph [0006]). Glezer teaches that multiple probes can be designed to target multiple regions on the RNA of interest (paragraph [0192]). This reads on one or more second encoding probes that are capable of hybridizing to a second target sequence. These one or more second encoding probes are also circularizable padlock probes similar to the “first” encoding probe (Figure 4-6).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Nilsson to include one or more second encoding probes capable of hybridizing to the same target nucleic acid, as taught by Glezer. One would be motivated to include one or more secondary encoding probes given the teaching by Glezer that this can “enhance the signal, and/or to provide a level of redundancy of targeting” (paragraph [0192]). One would have a reasonable expectation of success given that Glezer teaches targeting RNA molecules in situ, hybridizing multiple padlock probes to the same nucleic acid target, and performing RCA to detect said target in a manner similar to the methodology employed by Nilsson.
Nilsson in view of Glezer do not teach contacting the biological sample with one or more primary detectable probes that hybridize to the first encoding probe and/or second encoding probes and detecting a signal associated with the one or more primary detectable probes. However, detection of an encoding probe prior to amplification via RCA is known in the art, as taught by Pinard.
Pinard teaches using padlock probes in combination with fluorescent in situ hybridization to detect RNAs in biological samples (Abstract). Pinard teaches hybridizing padlock probes to a specific target and then using FISH probes to identify the padlocks that are bound to mRNA (paragraph 0032]). These FISH probes, applied prior to amplification via RCA, read on the one or more primary detectable probes that hybridize to the first encoding probe. Pinard teaches identifying/detecting the fluorescent signal in order to determine the barcode in the padlock probe (paragraph [0032]).
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Nilsson in view of Glezer to detect a signal associated with one or more primary detectable probes that are hybridized to the first encoding probe and second encoding probes. One would be motivated to do so given the teaching by Pinard that this would enable “selective RCA” in which only specific padlock probes would be amplified using the barcode as a priming site (paragraphs [0032] and [0055]). This enables control and which targets are amplified/detected in the reaction (paragraph [0034]). One would have a reasonable expectation of success given that Pinard demonstrates usage of padlock probes, hybridization of primary detection probes, and then RCA on said padlock probes for detection of nucleic acids of interest.
Regarding claim 2: Nilsson teaches using the first target sequence as a template to ligate and circularize the first encoding probe after hybridization of the interrogatory region to the region of interest (pg 10, ln 31-35).
Regarding claim 7: Nilsson teaches that the first encoding probe comprises one or more ribonucleotides (pg 1, ln 6 and pg 4, ln 5-6).
Regarding claim 12: Nilsson teaches that the target nucleic acid comprises RNA (pg 5, ln 24-26).
Regarding claim 14: Nilsson teaches that the hybridization region in the first encoding probe is split and contains a 5’ hybridization region and a 3’ hybridization region (pg 11, ln 1-5 and Fig 1A).
Regarding claim 21: Glezer teaches that each second encoding probe comprises a second hybridization region that is complementary to the second target sequence (paragraph [0192] and Fig 4-6).
Regarding claims 25 and 26: Glezer teaches that the padlock probes, a first and any one or more secondary, each independently comprise one or more barcode regions (paragraph [0192] and Fig 6), and that these barcodes can be different depending on the region of the nucleic acid molecule targeted (paragraph [0192]).
Regarding claim 27: As noted in the claim interpretation section above, “hybridization barcode sequence” is interpreted to mean a barcode is a sequence that can serve as identification of a particular target nucleic acid via hybridization of barcode specific probes. In which case, Pinard teaches that the encoding probes “collectively” comprise one or more hybridization barcode sequences that correspond to the target nucleic acid (paragraph [0032]).
Regarding claim 30: Nilsson teaches that the first encoding probe comprises a barcode sequence that is present in multiple copies within the RCA product (pg 31, ln 28-35 and pg 32 ln 1-6). As noted in the claim interpretation above, a barcode is also an “amplifiable” barcode once amplified via RCA.
Regarding claim 39: Nilsson teaches that padlock probe template-dependent ligation is sensitive enough to distinguish SNPs within an RNA template (pg 3, ln 4-6). Glezer teaches using multiple circularizable probes for a single target to provide redundancy in cases where the template has been mutated. These mutations would prevent circularization of secondary probes but would allow circularization of others (paragraph [0192]). Thus, Glezer teaches a scenario in which a first encoding probe is circularized and one or more second encoding probes are not circularized.
Regarding claim 43: Pinard teaching removing the one or more primary detectable probes without removing the first encoding probe from the target nucleic acid and before amplification (paragraph [0052]).
Regarding claim 46-48: Glezer teaches generating RCA products of multiple encoding probes on the same nucleic acid target template (paragraph [0192] and Fig 6). Glezer teaches detecting a signal associated with the RCA products (both second and first, Fig 5B) and that the detecting of each is performed sequentially (paragraph [0011] and Fig 5B).
Regarding claim 49: Nilsson teaches detecting the RCA product with detectable probes that bind to the RCA product (pg 44-45, ln 28-35 and ln 1-2, respectively)
Regarding claim 57: Glezer teaches that the probes hybridized to the same target nucleic acid can contain different primer binding sequences for RCA (Fig 6).
Claim 13 is rejected under 35 U.S.C. 103 as being unpatentable over Nilsson (Nilsson et al., WO 2019/068880 A1) in view of Glezer (Glezer et al., US 2022/0042083 A1; EFD of 8/6/2020) and Pinard (Pinard et al., US 2021/0403992 A1; EFD 6/29/2020) as applied to claims 1-2, 7, 12, 14, 21, 25-27, 30, 39, 43, 46-49, 57, and 84 above, and further in view of Zhang (Zhang et al., Chem. Soc. Rev., 2020).
The teachings of Nilsson in view of Glezer and Pinard are outlined above. Relevant to the instantly rejected claim, Nilsson in view of Glezer and Pinard teach use of a first encoding probe and one or more second encoding probes to detect target nucleic acid sequences of interest in a biological sample in situ. Nilsson in view of Glezer and Pinard teach that the first encoding probe comprises an interrogatory region that allows hybridization to the target nucleic acid and usage of said target nucleic acid as a template for ligation.
Nilsson in view of Glezer and Pinard do not teach that the biological sample comprises a counterpart target nucleic acid that does not allow hybridization to the interrogatory region in the first encoding probe or subsequent ligation of the first encoding probe using this counterpart target nucleic acid as a template. However, presence of counterpart target sequences that do not allow ligation of a target sequence probe is known in the art, as taught by Zhang.
Zhang teaches that a counterpart target nucleic acid may be present in a biological sample, such as the mutant or wild type allele of a specific target sequence (Fig 12G, in this particular example the first encoding probe is configured to hybridize and ligate on the mutant template while the “counterpart target nucleic acid” is a wild type allele; Fig 14B). Zhang teaches that the first encoding probe is unable to ligate the ends of the probe together on the counterpart target nucleic acid given that the region of interest (the SNP) is not complementary to the interrogatory region in the first encoding probe.
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Nilsson in view of Glezer and Pinard with that of Zhang. One would be motivated to employ this form of discriminatory detection given the assertion by Zhang that usage of padlock probes that discriminate between single nucleotide variants allows for efficient detection of these SNVs in situ (5. Bioanalytical and clinical applications). One would have a reasonable expectation of success given the teaching by Zhang that in situ SNV detection with the assistance of padlock probes is a commonly employed and successfully achieved assay.
Claim 58 is rejected under 35 U.S.C. 103 as being unpatentable over Nilsson (Nilsson et al., WO 2019/068880 A1) in view of Glezer (Glezer et al., US 2022/0042083 A1; EFD of 8/6/2020) and Pinard (Pinard et al., US 2021/0403992 A1; EFD 6/29/2020) as applied to claims 1-2, 7, 12, 14, 21, 25-27, 30, 39, 43, 46-49, 57, and 84 above, and further in view of Wu (Wu et al., Communications Biology, 2018; cited on IDS of 9/19/2023).
The teachings of Nilsson in view of Glezer and Pinard are outlined above. Relevant to the instantly rejected claim, Nilsson in view of Glezer and Pinard teach use of a first encoding probe and one or more second encoding probes to detect target nucleic acid sequences of interest in a biological sample in situ. Nilsson in view of Glezer and Pinard teach detecting, with one or more primary detecting probes, the first encoding probe bound to the target nucleic acid.
Nilsson in view of Glezer and Pinard do not teach selecting a duration of RCA using the signals associated with the one or more primary detectable probes.
Wu teaches detecting a product RCA and using this readout of fluorescent measurement to determine the optimal duration of RCA (Results, paragraph 2). While Wu does not teach detecting the padlock probe pre-amplification to determine the optimal duration of RCA, Pinard teaches that hybridizing fluorescent probes to padlock probes allows for detection of abundance of an RNA transcript (paragraph [0010]). Claim 58 encompasses determining relative durations based on fluorescent read outs of a target (i.e., longer versus shorter durations based on the strength of signal, page 18 of instant specification). Where the prior art teaches a variety of RCA durations used for in situ transcript detection, a person of ordinary skill has good reason to pursue the known options within his or her technical grasp for optimization of relevant RCA durations. If this leads to the anticipated success, it is likely the product not of innovation but of ordinary skill and common sense to provide routine optimization.
It would have been prima facie obvious to one having ordinary skill in the art, before the effective filing date of the instant application, to have modified the method of Nilsson in view of Glezer and Pinard with that of Wu. One would be motivated to employ the optimization of RCA duration based on fluorescent signal as taught by Wu, given Wu’s assertion that this allows optimization of the signal-to-noise ratio and prevents rolonies from becoming so large that they “merge” with other rolonies from adjacent transcripts (Results, paragraph 2). One would have a reasonable expectation of success given that Pinard demonstrates that fluorescence and relative abundance of the padlock probes can be ascertained prior to amplification, and Wu teaches that measuring fluorescence of target transcripts allows for optimization of RCA duration.
Response to Remarks
Applicant's arguments filed 1/23/2026 have been fully considered but they are not persuasive for the following reasons.
Applicant argues on page 10 of Remarks that “Pinard also does not teach a method involving both detection of an encoding probe via primary probes at a location in a biological sample and detection of an RCP of an encoding probe at the location in the biological sample as required by claim 1”. In response to applicant's arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). Pinard is relied upon for teaching primary detectable probes that can be used to detected a barcode of a first encoding probe at a location in a biological sample, and the benefits of using said detectable probes prior to performing rolling circle amplification. Applicant argues that Pinard teaches performing sequencing of RCPs outside of the tissue sample and therefore the performance of RCA in situ and sequencing in situ changes the principle of operation of Pinard if it were to be combined with the teachings of Nilsson and Glezer (page 10 of Remarks). The examiner respectfully disagrees. Pinard teaches amplification of selected circularized encoding probes in situ (paragraph [0067]), and teaches that the sequencing of said RCPs can be performed in situ OR ex situ (paragraph [0079]). Therefore, the combination of the teachings of Nilsson in view of Glezer with Pinard would not fundamentally change the principle of operation of Pinard.
Applicant argues that “the cited references do not provide motivation to combine Nilsson in view of Glezer with Pinard in the manner suggested by the office”, specifically that “the cited references do not point to any benefit or additional information that would be gained by modifying Nilsson according to Pinard” (page 10 of Remarks). However, this benefit is explicitly stated in the Non-Final rejection above. While Nilsson teaches the benefits of detecting the RCP in situ, Pinard teaches that additional benefit of first identifying the specific circularized encoding probes before performing RCA, as this would enable “selective RCA” in which only specific padlock probes would be amplified using the barcode as a priming site (paragraphs [0032] and [0055]). This enables control and which targets are amplified/detected in the reaction (paragraph [0034]). While Nilssons teaches that their RCP detection method “allows a high degree of sequence specificity to be obtained”, one reading Pinard would recognize that one could also detect the encoding probes first to selectively decide which products to obtain this high degree of specificity from.
Applicant argues on page 11 of Remarks that “the Examiner’s proposed modification to detect the padlocks of Nilsson via primary probe hybridization to identify the location of nucleic acids as in Pinard would fundamentally change the principle of operation of Nilsson”. The examiner respectfully disagrees. Using primary detectable probes, as taught by Pinard, would merely be an extra step before RCA as taught by Nilsson that would provide valuable information as to which barcodes were present at specific tissue locations and allow selective RCA of desirable products.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on nonstatutory double patenting provided the reference application or patent either is shown to be commonly owned with the examined application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. See MPEP § 717.02 for applications subject to examination under the first inventor to file provisions of the AIA as explained in MPEP § 2159. See MPEP § 2146 et seq. for applications not subject to examination under the first inventor to file provisions of the AIA . A terminal disclaimer must be signed in compliance with 37 CFR 1.321(b).
The filing of a terminal disclaimer by itself is not a complete reply to a nonstatutory double patenting (NSDP) rejection. A complete reply requires that the terminal disclaimer be accompanied by a reply requesting reconsideration of the prior Office action. Even where the NSDP rejection is provisional the reply must be complete. See MPEP § 804, subsection I.B.1. For a reply to a non-final Office action, see 37 CFR 1.111(a). For a reply to final Office action, see 37 CFR 1.113(c). A request for reconsideration while not provided for in 37 CFR 1.113(c) may be filed after final for consideration. See MPEP §§ 706.07(e) and 714.13.
The USPTO Internet website contains terminal disclaimer forms which may be used. Please visit www.uspto.gov/patent/patents-forms. The actual filing date of the application in which the form is filed determines what form (e.g., PTO/SB/25, PTO/SB/26, PTO/AIA /25, or PTO/AIA /26) should be used. A web-based eTerminal Disclaimer may be filled out completely online using web-screens. An eTerminal Disclaimer that meets all requirements is auto-processed and approved immediately upon submission. For more information about eTerminal Disclaimers, refer to www.uspto.gov/patents/apply/applying-online/eterminal-disclaimer.
Withdrawn:
The double patenting rejections over claims of U.S. Patent No. US 12,060,603 B2 and co-pending application 18/099,865 are withdrawn in light of Applicant’s filing of two approved Terminal Disclaimers.
The double patenting rejection over claims of US Patent No. US 12,400,733 B2 are withdrawn in light of Applicant’s amendments to the claims.
New (Necessitated by Amendments):
U.S. Patent No. US 12,400,733 B2
Although the claims at issue are not identical, they are not patentably distinct from each other because both sets of claims are drawn to the same limitations. Any additional limitations of the conflicting claims are encompassed by the open claim language “comprising” found in the instant claims.
Claims 1, 12, 25-27, 30, 43, and 46-48, are rejected on the ground of nonstatutory double patenting as being unpatentable over claims 16, 18, and 25 of U.S. Patent No. US 12,400,733 B2 in view of Pinard (Pinard et al., US 2021/0403992 A1; EFD 6/29/2020) according to citations and rationales provided above.
Claims 2, 7, 13-14, 21, 39, 49, 57-58, and 84 are rejected on the ground of nonstatutory double patenting as being unpatentable over claims 16, 18, and 25 of U.S. Patent No. US 12,400,733 B2 in view of Pinard (Pinard et al., US 2021/0403992 A1; EFD 6/29/2020) as applied to claims 1, 12, 25-27, 30, 43, and 46-48 above, and further in view of Nilsson (Nilsson et al., WO 2019/068880 A1), Glezer (Glezer et al., US 2022/0042083 A1; EFD of 8/6/2020), Zhang (Zhang et al., Chem. Soc. Rev., 2020), and Wu (Wu et al., Communications Biology, 2018; cited on IDS of 9/19/2023) according to citations and rationales provided above.
Response to Remarks
Applicant traverses the rejections on the ground of nonstatutory double patenting over claims 16, 18, and 25 of US Patent No. US 12,400,733 B2 in view of Pinard, Nilsson, Glezer, Zhang, and Wu (page 13 of Remarks) in view of the arguments and amendments provided in the Remarks of 1/23/2026. However, as noted in the 103 rejections above, Applicant’s arguments were not deemed persuasive, and as such the rejection on the grounds of nonstatutory double patenting are maintained, though amended as necessitated by the amendments to the claims.
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to KAILEY E CASH whose telephone number is (571)272-0971. The examiner can normally be reached Monday-Friday 8:30am-6pm ET.
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/KAILEY ELIZABETH CASH/Examiner, Art Unit 1683
/STEPHEN T KAPUSHOC/Primary Examiner, Art Unit 1683