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.
Claim Status
Claims 1-11 and 18-26 are pending.
Claims 18-26 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 11/5/2025.
Claims 1-11 are being examined on the merits.
Information Disclosure Statement
The listing of references in the specification is not a proper information disclosure statement (see paragraphs [0055, 0079, and 0084] for examples). 37 CFR 1.98(b) requires a list of all patents, publications, or other information submitted for consideration by the Office, and MPEP § 609.04(a) states, "the list may not be incorporated into the specification but must be submitted in a separate paper." Therefore, unless the references have been cited by the examiner on form PTO-892, they have not been considered. No IDS was filed with the reply of 3/26/2026, and therefore any references in the specification not present in the previous IDSs remain unconsidered unless cited by the examiner.
Specification
Applicant’s amendments to the specification, submitted on 3/23/2026, to properly note trade names and marks used in commerce are acknowledged.
Claim Objections
The objections to claims 4 and 6 are withdrawn in light of Applicant’s amendments to the claims.
Claim Rejections - 35 USC § 112b - Indefiniteness
The rejection of claims 1, 3-6, and 8-11 under 35 U.S.C. 112(b) are withdrawn in light of Applicant’s amendments to the claims and provided clarifications regarding “a first assay” result in claim 1.
The rejection of claims 2 and 6 (below) 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 are maintained.
Claim 2 recites the limitation “introducing the extracted recombinant nucleic acids into a well and/or microchannel of a microfluidic device”. It is unclear how a feature of a microfluidic device could be both a well AND a microchannel. Clarification is required.
Claim 7 recites the limitation “introducing the extracted proteins of the particles into a well and/or microchannel of a microfluidic device”. It is unclear how a feature of a microfluidic device could be both a well AND a microchannel. Clarification is required.
Response to Remarks
Applicant has responded to the 112b rejections above with clarifying remarks (pg 11 of Remarks of 3/23/2026). “Applicant submits that the extracted recombinant nucleic acids [or extracted recombinant proteins] may be introduced into either or both a well and a microchannel of a microfluidic device”. However, this does not clarify the issue. It is still unclear how a feature can be BOTH a well AND a microchannel. If it’s a case wherein the extracted recombinant molecules are flown through a well and then a microchannel, it is additionally unclear as to which feature contains the detectable label.
Maintained Claim Rejections - 35 USC § 103
Claims 1-3, 5, and 8-10 are rejected under 35 U.S.C. 103 as being unpatentable over Wu (Wu et al., US 2023/0126263 A1; EFD of 9/3/2021) in view of Hutanu (Hutanu et al., Electrophoresis 2021) and Park (Park et al., WO 2005/068993 A1; cited on IDS of 10/3/2023).
Regarding claim 1: Wu teaches a method of characterizing fluid samples containing particles to determine a ratio of full particles to empty particles in the fluid sample (Abstract). Wu teaches applying a sample containing particles to a capillary containing a separation matrix, labeling the target component (such as nucleic acid) with a selective binding agent (which can be added before or after separation (paragraph [0056]), detecting a signal and determining the quantity/characteristic of the nucleic acid based on the migration through the separation matrix (paragraphs [0005-0006, 0025]). Wu teaches that the nucleic acid can be recombinant nucleic acid within a viral particle such as adeno-associated virus (paragraphs [0022 and 0023]). Wu teaches comparing the data obtained from a test sample of particles to data obtained from a sample with known proportions of full/empty particles (Abstract). This allows for the establishment of a relationship between nucleic acid content and proportions of particles in a “full” state which allows for prediction of proportions of particles in full, partial, and empty states in particles of an unknown state (Abstract, paragraph [0019]). Comparison of the test sample to a reference standard representative of full particles allows for determining of the ratio of full particles to partially full particles to produce a first assay result and characterize the population of particles in the fluid sample. Wu teaches that multiple aliquots can be obtained from the same sample to run samples through two different analyses, one targeting nucleic acid and the other protein (paragraph [0054]).
Wu does not teach extracting the recombinant nucleic acids from the first aliquot of the sample or flowing the sample through a polymeric separation medium within the microchannel to separate the labeled nucleic acids by size and characterizing the nucleic acids by the time it takes to flow through the polymeric separation channel. However, extraction of recombinant nucleic acids from viral particles to characterize the nucleic acid sizes by time it takes to travel through a polymeric separation medium is known in the art, as taught by Hutanu.
Hutanu teaches using capillary gel electrophoresis to characterize nucleic acids from viral particle samples (3.3 Capillary gel electrophoresis). Hutanu teaches application of nucleic acids from viral particles to a separation matrix comprised of 1% PVP (a polymeric separation medium) and SYBR-Green II (for labeling the nucleic acid; 3.3 Capillary gel electrophoresis). Hutanu teaches that for proper size distinction by gel electrophoresis, the nucleic acid needs to be released from the virus (3 Results and discussion) and could be achieved using the QIAquick PCR purification kit (2 Materials and methods – CGE). Hutanu teaches determining the size of the nucleic acids by the time it takes to travel through the separation medium and comparison to known size standards (Figure 3).
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 Wu with the size separation medium as taught by Hutanu. One would be motivated to characterize nucleic acids according to size given the assertion by Hutanu that with CGE it “is possible to separate all DNA size species, depending on their sequence length and conformation” and “quantify by signal intensity using an appropriate reference standard” (3.3 Capillary gel electrophoresis). Additionally, Hutanu demonstrates that the resolution that can be achieved with CGE allows for determination of improperly filled viral particles as well (fragments or over fill), which may not be possible with other methods (3.3 Capillary gel electrophoresis). One would have a reasonable expectation of success in using this methodology given that Wu teaches that their method can also be applied to information obtained from systems that employ capillaries with a separation matrix that separates nucleic acids by size based on voltage application (paragraph [0090]).
Wu in view of Hutanu does not teach flowing the labeled extracted recombinant nucleic acids from a polymeric separation medium in a microchannel into a detection region that is in fluid communication with said microchannel and which comprises a sensor capable of detecting a signal. However, microfluidic devices that allow for separation of nucleic acids through a separation medium in a microchannel that is in fluid communication with a detection region and a sensor is known in the art, as taught by Park.
Park teaches a method for spatially separating components in a sample using a microfluidic channel in a microfluidic device (Abstract). Park teaches applying analytes (such as nucleic acids or proteins) to a separation channel segment comprised of through separation media to affect migration of the analytes by size to resolve and identify/quantitate them (pg 3, ln 19; pg 4, ln 14-21). Separation channel segment reads on polymeric separation medium in a microchannel (pg 4, ln 14-21; pg 20, ln 19-21; pg 24, ln 15-25). Park teaches a detection region in fluid communication with the microchannel containing the polymeric separation medium (pg 37, ln 26-27) in which there is a sensor that can detect a signal from a nucleic acid or protein which has been labeled as they exit the polymeric separation medium (pg 4, ln 23-26; pg 8, ln 14-18; pg 22, ln 1-2; pg 37, ln 26-27). Park teaches determining an amount of time taken by the labelled nucleic acids to flow through the polymeric separation medium in the microchannel into the detection region, which is indicative of the size of the nucleic acids in the fluid sample (pg 24, ln 15-25 and pg 25, ln 18-25).
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 performed the method of Wu in view of Hutanu with the microfluidic device as taught by Park. One would be motivated to do so given the assertion by Park that their system provides “highly repeatable analytical results with high sensitivity, speed and resolution” (pg 3, ln 9-10 and pg 11, ln 25-28). One would have a reasonable expectation of success given that Park teaches application of nucleic acids to a microfluidic channel that contains a separation medium such as polyvinylpyrrolidone (PVP, as used in Hutanu; pg 24, ln 22).
Regarding claim 2: Hutanu teaches that the polymeric separation medium comprises SYBR Green II, which binds to the nucleic acids upon introduction to the capillary (or microchannel) and produced labeled extracted nucleic acids (3.3 Capillary gel electrophoresis).
Regarding claim 3 and 5: Wu teaches determining amount of particle protein in a sample taken from a second aliquot by assaying the particle protein (paragraph [0054]).
Regarding claim 8 and 9: Wu teaches that the nucleic acid can be recombinant nucleic acid within a viral particle such as adeno-associated virus (paragraphs [0022 and 0023]). Wu teaches that recombinant AAVs contain plasmids up to 5kb in length (reads on single -stranded DNA genome in the range of 500-7000 nucleotides in length; paragraph [0023]).
Regarding claim 10: Hutanu teaches that there is good correlation in capillary electrophoretic methods between the fluorescence signal and the virus concentration when particles are in the range of 10^11 to 10^13 GC/mL and demonstrated results of an electropherogram of AAV samples at 10^13 viral particles per mL (Figure 4 and 3.4 Capillary zone electrophoresis).
Claims 4 and 11 are rejected under 35 U.S.C. 103 as being unpatentable over Wu (Wu et al., US 2023/0126263 A1; EFD of 9/3/2021) in view of Hutanu (Hutanu et al., Electrophoresis 2021) and Park (Park et al., WO 2005/068993 A1; cited on IDS of 10/3/2023) as applied to claims 1-3, 5, and 8-10 above, and further in view of Luo (Luo et al., LCGC Supplements, 2020).
The teachings of Wu in view of Hutanu and Park as they apply to claims 1 and 3, from which claims 4 and 11 depend, are detailed above. Relevant to the instantly rejected claims, Wu in view of Hutanu and Park teach a method of characterizing particles in a fluid sample through size separation polymeric medium within a microchannel of a microfluidic device. Wu in view of Hutanu and Park teach that the nucleic acid has to be extracted from the particles for accurate analysis in a polymeric separation medium.
Wu in view of Hutanu and Park do not teach determination of total number of particles in a fluid sample without assaying a particle protein in the fluid sample (claim 4) and they do not teach that the nucleic acid is extracted by contacting particles with a proteinase and a denaturing agent (claim 11). However, determination of a total number of particles in a fluid sample without assaying a particle protein in the fluid sample, and extraction of nucleic acid by proteinase and denaturing agents is known in the art, as taught by Luo.
Luo teaches determination of AAV viral integrity through analysis of nucleic acids within the AAV particles using capillary gel electrophoresis methods (Abstract). Luo teaches contacting the samples with proteinase K (a proteinase) and a denaturing agent (Abstract guanidine-hydrochloride; guanidinium chloride is identified as a denaturing agent in paragraph [0097] of the instant specification). Luo teaches using capillary gel electrophoresis to determine the amounts and purities of nucleic acids in viral samples (Abstract). Luo teaches that the measuring of peak area differences between known “full” and known “empty” capsid samples may offer a titer determination perspective (thus not requiring measurement of protein amounts in the sample to determine the total number of particles in the sample (Analysis of the Origin of Impurities, paragraph 2 and Conclusions).
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 Wu in view of Hutanu and Park with that of Luo. One would be motivated to perform proteinase and denaturing treatment on the particles to extract nucleic acids given the assertion by Luo that this method is recommended for “in-depth impurity analysis” (Conclusion). Additionally, one would be motivated to determine the total number of particles in the fluid sample without assaying particle protein through the measurement of signal intensities under peaks of nucleic acid sizes and comparing to known samples with known concentrations given that this would obviate the need for an extra step of protein measurement in the sample (Analysis of the Origin of Impurities). One would have a reasonable expectation of success given that Luo demonstrates the successful size separation of extracted nucleic acids from viral particles in a capillary gel electrophoresis method.
Claims 6 and 7 are rejected under 35 U.S.C. 103 as being unpatentable over Wu (Wu et al., US 2023/0126263 A1; EFD of 9/3/2021) in view of Hutanu (Hutanu et al., Electrophoresis 2021) and Park (Park et al., WO 2005/068993 A1; cited on IDS of 10/3/2023) as applied to claims 1-3, 5, and 8-10 above, and further in view of Zhang (Zhang et al., Human Gene Therapy 2021).
The teachings of Wu in view of Hutanu and Park as they apply to claims 1-3, 5, and 8-10 are detailed above. Relevant to the instantly rejected claims, Wu in view of Hutanu and Park teach a method of characterizing particles in a fluid sample through size separation polymeric medium within a microchannel of a microfluidic device. Wu in view of Hutanu and Park teach methodologies for size dependent separation of labeled proteins for characterization of protein compositions of samples (and specifically protein compositions of particle samples in Wu; Wu: paragraphs [0025, 0027, and 0054]; Park: pg 3, ln 19, pg 24, ln 15-25).
Regarding claim 6: Wu in view of Hutanu and Park teaches a second aliquot of fluid sample in which proteins of particles are analyzed by flowing through a polymeric separation medium in a microchannel into a detection region with a sensor capable of detecting a signal from labeled protein that are separated according to size (Wu: paragraphs [0006, 0025, 0027, and 0054] Park: pg 4, ln 14-21, pg 8, ln 21, pg 20, ln 19-21, pg 25, ln 18-25). Park teaches determining an amount of time taken by the labelled proteins to flow through the polymeric separation medium in the microchannel into the detection region, which is indicative of the size of the proteins in the fluid sample (pg 24, ln 15-25 and pg 25, ln 18-25). Wu teaches quantitating the amount of protein in the sample by analyzing the area under the curve associated with signal to quantitate amounts of protein in the sample (paragraph [0054]). Wu teaches comparing measurements to known references standards to determine the amount of protein in the sample to produce a second assay result (paragraph [0088]). Wu teaches comparing the first assay result (nucleic acid) to the second assay result (protein), to determine a ratio of full viral particles to empty viral particles in the fluid sample, thus characterizing the population of recombinant viral particles in the fluid sample (paragraph [0088]).
Wu in view of Hutanu and Park do not explicitly teach extracting the proteins from the population of particles. However, extracting proteins from a population of particles in order to perform size separation analysis is known in the art, as taught by Zhang.
Zhang teaches extracting proteins from AAV fluid samples for analysis via capillary gel electrophoresis in order to determine the size separation (composition of proteins in the viral particles) and the relative peak sizes for protein abundance (Abstract and Experimental - CE-SDS LIF sample preparation; Figure 1).
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 Wu in view of Hutanu and Park with that of Zhang. One would be motivated to do so given the assertion by Zhang that the practice of extracting the proteins from the sample is well-known and routinely practiced in the art (Introduction, paragraph 2). One would have a reasonable expectation of success given that Zhang successfully extracts proteins from fluid particle samples for separation via capillary gel electrophoresis.
Regarding claim 7: Wu teaches binding of detectable labels to proteins in the separation channel (paragraphs [0006, 0056]).
Response to Remarks
Applicant has traversed the rejections of claims 1-3, 5, and 8-10 over Wu in view of Hutanu and Park (pg 12-14 of Remarks of 3/23/2026). Applicant's arguments filed 3/23/2026 have been fully considered but they are not persuasive for the following reasons.
Applicant argues that the “entire basis” for Wu is the use of isoelectric focusing through a pH gradient for separation of particles. The examiner respectfully disagrees. The primary objective of Wu is to analyze “samples containing particles used for gene delivery to determine a quality of the sample and/or an indication that the gene delivery particles are in a full, partial, and/or empty state” and “determining a protein and/or NA content in samples with known proportions of gene delivery particles in a full, partial, and/or empty state and based on the determination, establish a relationship between NA content and proportions of gene delivery particles in a full state” (Abstract). Wu then provides a method for separating said particles to perform the analysis which does rely on separation of particles through a pH gradient using differences in isoelectric points. However, as taught by Hutanu, the same sort of separation of particles to determine differences between full, empty, and partial particles can be performed in another methodology, one which Hutanu teaches has specific benefits (as noted above). The use of a separate methodology to also enable analysis of particles in full, empty, or partial states, only separated by size rather than isoelectric point, does not fundamentally alter the aims of the analysis of Wu but merely provides a different methodology for achieving the separation of particles.
As noted in the previous Office Action of 12/5/2025 that “One would have a reasonable expectation of success in using this methodology given that Wu teaches that their method can also be applied to information obtained from systems that employ capillaries with a separation matrix that separates nucleic acids by size based on voltage application (paragraph [0090]).” Applicant disagrees with the Examiner’s interpretation of paragraph [0090] and states that “the Wu reference does not teach what the Examiner asserts”. Applicant argues that this paragraph “merely states a characteristic of ‘ProteinSimple’s® Simple Western® instrument’” (pg 14 of Remarks). The Examiner respectfully disagrees. Wu explicitly states that “the methods and processes described herein can be performed using existing systems similar to ProteinSimple’s® Simple Western® instrument” (emphasis added). Wu goes on to teach that such similar instruments may use different separation matrices to those taught by the instant application such as separation via size within a capillary, and that signals from particles run through said capillaries can be detected and analyzed to determine quantities of the components involved. Specifically, Wu indicates that “a sample containing a heterogeneous mixture of GD particles [viral vectors] can be loaded, and capillaries can be brought into contact with running buffer. Voltage can be applied to enable separation by molecular weight, isoelectric points, or other suitable characteristic”. The mention that viral vectors can be separated by weight to separate a heterogeneous mixture again points to the fact that Wu acknowledges that such matrices can be used for separation and classification of viral vectors/particles. Additionally, Wu teaches that GD particles (viral vectors) may be separated based on size to achieve the intended analytic output (paragraph [0060] and claims 29-32 in the embodiments listed in paragraph [0094]).
For these reasons, the rejections of claims 1-3, 5, and 8-10 over Wu in view of Hutanu and Park, the rejections of their dependent claims over Luo and Zhang under 35 USC 103 are maintained.
Conclusion
No claims are allowed.
THIS ACTION IS MADE FINAL. 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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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.
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/KAILEY ELIZABETH CASH/Examiner, Art Unit 1683
/ANNE M. GUSSOW/Supervisory Patent Examiner, Art Unit 1683
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