.DETAILED ACTION
The Applicant’s response, received 24 November 2025 has been fully considered. The following rejections and/or objections are either reiterated or newly applied. They constitute the complete set presently being applied to the instant application.
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 Claims
Claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 are pending.
Claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 are rejected.
Priority
There are no domestic or foreign applications for which benefit of priority is claimed.
The effective filing date of the claimed invention is 21 February 2024.
Claim Rejections - 35 USC § 112
The amendment and arguments/remarks received 24 November 2025 have been fully considered, however after further consideration, the rejection raised under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, in the Office action mailed 03 June 2025 is maintained in view of the amendment and arguments/remarks, as noted below.
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 2, 16, 18, and 19 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. ii
Claim 2 is indefinite for reciting the limitation “wherein generating the count of the number of target molecules of the sample distributed across the set of partitions is performed within 5 seconds” because it is not clear as to whether the “performed within 5 seconds” limitation means that the light sheet scanning of partitions within the container is performed within 5 seconds, or alternatively if it is the quantification and/or enumeration of the fluorescent signals generated by the light-sheet scan that is performed with 5 seconds. This limitation is interpreted to mean quantification and/or enumeration of the fluorescent signals generated by the light-sheet scan.
Claim 16 is indefinite for reciting the limitation “wherein generating the count of the number of target molecules of the sample distributed across the set of partitions comprises omitting processing of a subset of sheets” because the claim from which claim 16 depends (claim 1) only recites “scanning a sheet of partitions.”
Claim 18 is indefinite for reciting the limitation “wherein generating the count of the number of target molecules of the sample distributed across the set of partitions comprises generating the count in less than 90 seconds” because it is not clear as to whether the “generating the count in less than 90 seconds” limitation means that the light sheet scanning of partitions within the container is performed in less than 90 seconds, or alternatively if it is the quantification and/or enumeration of the fluorescent signals generated by the light-sheet scan that is performed in less than 90 seconds. This limitation is interpreted to mean quantification and/or enumeration of the fluorescent signals generated by the light-sheet scan.
Claim 19 is indefinite for reciting the limitation “enumerating positive partitions of the set of partitions upon scanning a set of sheets of partitions spanning less than 25% of an internal volume of the container using light sheet scanning, and enumerating positive partitions identified from the set of sheets” because it is not clear as to how the first enumerating step differs from the second enumerating step.
Response to Arguments
The Applicant’s arguments/remarks received 24 November 2025 have been fully considered, but are not persuasive.
The Applicant states on page 6 (para. 3) of the Remarks that a skilled artisan in reading claim 2 would understand that the 5 seconds refers to “generating the count of the number of target molecules of the sample distributed across the set of partitions,” of claim 1 and accordingly would consider claim 2 definite.
This argument is not persuasive, because claim 1 also recites the wherein clause “wherein generating the count comprises scanning a sheet of partitions within the container using light sheet scanning,” and therefore claim 2 is indefinite because it is not clear as to whether the further limitation reciting “performed within 5 seconds” also comprises the step of performing the light sheet scanning of the sheet of partitions. For example, if ‘generating’ involves a step assessing the various criteria it is unclear if this is accomplished within 5 seconds, or if only once the data is obtained the assessment of the data requires 5 seconds.
More clearly setting forth what is required of generating and what is performed in 5 seconds would address the basis of the rejection of limitation for ‘the count’ accomplished.
The Applicant states on page 6 (para. 4) of the Remarks that a skilled artisan would understand that claim 16 involves “omitting processing of a subset of sheets positioned away from the center of the container along the lateral axis,” and that claim 16 is definite.
This argument is not persuasive, because claim 1 only recites a single sheet, i.e., “scanning a sheet of partitions,” and therefore claim 16 is indefinite because it is not clear as to how there can be a further limiting step of “omitting processing of a subset of sheets” (emphasis added). It is unclear how a subset is generated if there is only one.
The Applicant states on page 6 (para. 5) of the Remarks that the Office alleges that claim 18 is indefinite for similar reasons as claim 1, and that as described above, a skilled artisan, in reading claim 18, would consider it definite.
This argument is not persuasive, because first, claim 1 is not rejected under 35 U.S.C. 112(b) in the Office action mailed 03 June 2025. Second, this argument is not persuasive for the same reason provided above in the foregoing response to argument regarding claim 2.
The Applicant states on page 6 (para. 6) of the Remarks that a skilled artisan in reading claim 19 would understand that “enumerating positive partitions of the set of partitions,” and “enumerating positive partition identified from the set of sheets,” are different, and accordingly, that claim 19 is definite.
This argument is not persuasive, because both the first and second enumerating steps recite “enumerating positive partitions,” and therefore it is not clear as to how the enumerating steps are different, and therefore the claim is indefinite.
Claim Rejections - 35 USC § 101
The amendment and arguments/remarks received 24 November 2025 have been fully considered, however after further consideration, the rejection raised under 35 U.S.C. 101 in the Office action mailed 03 June 2025 is maintained in view of the amendment and arguments/remarks, as noted below.
35 U.S.C. 101 reads as follows:
Whoever invents or discovers any new and useful process, machine, manufacture, or composition of matter, or any new and useful improvement thereof, may obtain a patent therefor, subject to the conditions and requirements of this title.
Claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 are rejected under 35 U.S.C. 101 because the claimed invention is directed to an abstract idea without significantly more. The claims recite both: (a) mathematical concepts, (e.g., mathematical relationships, formulas or equations, mathematical calculations); and (b) mental processes, i.e., concepts performed in the human mind, (e.g., observation, evaluation, judgement, opinion).
Subject matter eligibility evaluation in accordance with MPEP 2106.
Eligibility Step 1: Step 1 of the eligibility analysis asks: Is the claim to a process, machine, manufacture or composition of matter?
Claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 are directed to a method (i.e., a process) for generating a count.
Therefore, these claims are encompassed by the categories of statutory subject matter, and thus, satisfy the subject matter eligibility requirements under step 1.
[Step 1: YES]
Eligibility Step 2A: First it is determined in Prong One whether a claim recites a judicial exception, and if so, then it is determined in Prong Two whether the recited judicial exception is integrated into a practical application of that exception.
Eligibility Step 2A Prong One: In determining whether a claim is directed to a judicial exception, examination is performed that analyzes whether the claim recites a judicial exception, i.e., whether a law of nature, natural phenomenon, or abstract idea is set forth or described in the claim.
Independent claim 1 recites the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas:
generating a count of a number of target molecules of a sample distributed across a set of partitions by identifying partitions of the set of partitions comprising a target molecule based on satisfaction of a signal criterion, an optical property criterion, a shape criterion, and a morphology criterion, wherein each partition of the set of partitions contains less than two target molecules (i.e., mental processes and mathematical concepts);
wherein the signal intensity criterion is evaluated for a set of pixels depicting a partition of the set of partitions, wherein the signal intensity criterion is based upon a maximum signal intensity of the set of pixels (i.e., mental processes); and
wherein the count has: a precision characterized by a coefficient of variation less than 7%, and an accuracy greater than 95% (i.e., mental processes and mathematical concepts).
Dependent claims 6-9, 12-14, 16, 19, 20, and 22-24 further recite the following steps which fall within the mental processes and/or mathematical concepts groupings of abstract ideas, as noted below.
Dependent claim 6 further recites:
if the number of target molecules within the set of partitions is within a first range, generating the count comprises performing a first set of operations, if the number of target molecules within the set of partitions is within a second range, generating the count comprises performing a second set of operations different than the first set of steps, and if the number of target molecules within the set of partitions is within a third range, generating the count comprises generating a third set of operations different than the first set of operations and the second set of operations (i.e., mental processes and mathematical concepts);
Dependent claim 7 further recites:
wherein the first range comprises a range from 1 target molecule to 100 target molecules (i.e., mental processes and mathematical concepts);
Dependent claim 8 further recites:
wherein the third range comprises a range from 100,000 target molecules to 1,000,000 target molecules (i.e., mental processes and mathematical concepts);
Dependent claim 9 further recites:
wherein the second range comprises 100 target molecules to 100,000 target molecules (i.e., mental processes and mathematical concepts);
Dependent claim 12 further recites:
wherein the optical property criterion is evaluated for a set of pixels depicting a droplet of the set of partitions, and wherein the optical property criterion is based upon a lateral intensity gradient across the set of pixels (i.e., mental processes);
Dependent claim 13 further recites:
wherein the shape criterion is evaluated for a set of pixels depicting a partition of the set of partitions, and wherein the shape criterion is based upon a droplet radius determined from the set of pixels and a partition eccentricity determined from the set of pixels (i.e., mental processes and mathematical concepts);
Dependent claim 14 further recites:
wherein the morphology criterion is evaluated for a set of pixels depicting a partition of the set of partitions, and wherein the morphology criterion is based upon determination of a correlation with a three-dimensional profile of a representative positive partition determined from a set of control samples (i.e., mental processes);
Dependent claim 16 further recites:
wherein the container comprises a width and a lateral axis referenced to a center of the container, and wherein generating the count of the number of target molecules of the sample distributed across the set of partitions comprises omitting processing of a subset of sheets positioned away from the center of the container along the lateral axis (i.e., mental processes and mathematical concepts);
Dependent claim 19 further recites:
enumerating positive partitions identified from the set of sheets (i.e., mental processes and mathematical concepts);
Dependent claim 20 further recites:
projecting candidate positive partitions onto a two-dimensional mapping (UMAP) of droplets, and counting only partitions of a cluster of positive partitions from the two-dimensional mapping of partitions (i.e., mental processes and mathematical concepts);
Dependent claim 22 further recites:
wherein the signal intensity criterion is further based upon a maximum convoluted intensity of the set of pixels, a difference between the maximum intensity and the minimum intensity of the set of pixels, and a total intensity of the set of pixels (i.e., mental processes and mathematical concepts);
Dependent claim 23 further recites:
wherein the optical property criterion is further based upon second order intensity gradients across the set of pixels along a set of directions, and intensity uniformity across the set of pixels (i.e., mental processes and mathematical concepts); and
Dependent claim 24 further recites:
wherein the count of the number of target molecules of the sample distributed across the set of partitions is between 1 target molecule and 1,000,000 target molecules (i.e., mental processes and mathematical concepts).
The abstract ideas recited in the claims are evaluated under the broadest reasonable interpretation (BRI) of the claim limitations when read in light of and consistent with the specification. As noted in the foregoing section, the claims are determined to contain limitations that can practically be performed in the human mind with the aid of a pen and paper (e.g., generating a count of a set of targets in a sample (e.g., paras. [00115] & [00146] in the specification; FIG. 1A) upon conversion to data (i.e., information) can be practically performed in the human mind with or without a physical aid), and therefore recite judicial exceptions from the mental process grouping of abstract ideas. Additionally, the recited limitations that are identified as judicial exceptions from the mathematical concepts grouping of abstract ideas (e.g., generating a count) are abstract ideas irrespective of whether or not the limitations are practical to perform in the human mind.
Therefore, claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 recite an abstract idea.
[Step 2A Prong One: YES]
Eligibility Step 2A Prong Two: In determining whether a claim is directed to a judicial exception, further examination is performed that analyzes if the claim recites additional elements that when examined as a whole integrates the judicial exception(s) into a practical application (MPEP 2106.04(d)). A claim that integrates a judicial exception into a practical application will apply, rely on, or use the judicial exception in a manner that imposes a meaningful limit on the judicial exception. The claimed additional elements are analyzed to determine if the abstract idea is integrated into a practical application (MPEP 2106.04(d)(I); MPEP 2106.05(a-h)). If the claim contains no additional elements beyond the abstract idea, the claim fails to integrate the abstract idea into a practical application (MPEP 2106.04(d)(III)).
The judicial exceptions identified in Eligibility Step 2A Prong One are not integrated into a practical application because of the reasons noted below.
Dependent claims 6-9, 16, 20, and 24 do not recite any elements in addition to the judicial exception, and thus are part of the judicial exception.
The additional elements in independent claim 1 include:
target molecules of a sample distributed across a set of partitions;
wherein each partition of the set of partitions contains less than two target molecules;
wherein the set of partitions are stabilized in position within a container;
scanning a sheet of partitions within the container using light sheet scanning; and
a set of pixels (i.e., digital image).
The additional elements in dependent claims 2, 3, 5, 12, 13, 14, 18, 19, 22, and 23 include:
wherein generating the count of the number of target molecules of the sample distributed across the set of partitions is performed within 5 seconds (claim 2);
wherein the set of partitions comprises at least 20 million partitions (claim 3);
wherein the container comprises a volumetric capacity of 50 microliters, and wherein the set of partitions comprises at least 30 million partitions (claim 5);
wherein generating the count of the number of target molecules of the sample distributed across the set of partitions comprises generating the count in less than 90 seconds (claim 18);
scanning a set of sheets of partitions spanning less than 25% of an internal volume of the container using light sheet scanning (claim 19); and
a set of pixels (i.e., a digital image) (claims 12, 13, 14, 22, and 23).
The additional element of light sheet scanning (claims 1 and 19) is not an improvement to computer functionality itself, or an improvement to any other technology or technical field (see MPEP 2106.04(d)(1)), and therefore does not integrate the recited judicial exceptions into a practical application.
The additional element of a set of pixels (claims 1, 12, 13, 14, 22, and 23) is not an improvement to computer functionality itself, or an improvement to any other technology or technical field (see MPEP 2106.04(d)(1)), and therefore does not integrate the recited judicial exceptions into a practical application.
The additional elements of scanning a sheet of partitions within a container using light sheet scanning (claims 1 and 19); target molecules of a sample distributed across a set of partitions (claim 1); wherein each partition of the set of partitions contains less than two target molecules (claim 1); wherein the set of partitions are stabilized in position within a container (claim 1); wherein the set of partitions comprises at least 20 million partitions (claim 3); and wherein the container comprises a volumetric capacity of 50 microliters, and wherein the set of partitions comprises at least 30 million partitions (claim 5); are part of the data gathering process that generates the data used in the recited judicial exceptions, (MPEP 2106.05(g)), and also do not amount to more than generally linking the use of a judicial exception to a particular technological environment (MPEP 2106.05(h)), and therefore do not integrate the recited judicial exceptions into a practical application.
The additional elements of generating a count that is performed within 5 seconds (claim 2) and generating a count in less than 90 seconds (claim 18) do not amount to more than mere instructions to implement an abstract idea on a computer (MPEP 2106.05(f) (2) iii. (increased speed in the process comes solely from the capabilities of the general-purpose computer)).
Thus, the additionally recited elements merely invoke a computer as a tool, and/or amount to merely indicating a field of use or technological environment in which to apply a judicial exception, and/or amount to data gathering activity, and as such, when all limitations in claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 have been considered as a whole, the claims are deemed to not recite any additional elements that would integrate a judicial exception into a practical application, and therefore claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 are directed to an abstract idea (MPEP 2106.04(d)).
[Step 2A Prong Two: NO]
Eligibility Step 2B: Because the claims recite an abstract idea, and do not integrate that abstract idea into a practical application, the claims are probed for a specific inventive concept. The judicial exception alone cannot provide that inventive concept or practical application (MPEP 2106.05). Identifying whether the additional elements beyond the abstract idea amount to such an inventive concept requires considering the additional elements individually and in combination to determine if they amount to significantly more than the judicial exception (MPEP 2106.05A i-vi).
The claims do not include any additional elements that are sufficient to amount to significantly more than the judicial exception(s) because of the reasons noted below.
Dependent claims 6-9, 16, 20, and 24 do not recite any elements in addition to the judicial exception(s).
The additional elements recited in independent claim 1 and dependent claims 2, 3, 5, 12, 13, 14, 18, 19, 22, and 23 are identified above, and carried over from Step 2A: Prong Two along with their conclusions for analysis at Step 2B. Any additional element or combination of elements that was considered to be insignificant extra-solution activity at Step 2A: Prong Two was re-evaluated at Step 2B, because if such re-evaluation finds that the element is unconventional or otherwise more than what is well-understood, routine, conventional activity in the field, this finding may indicate that the additional element is no longer considered to be insignificant; and all additional elements and combination of elements were evaluated to determine whether any additional elements or combination of elements are other than what is well-understood, routine, conventional activity in the field, or simply append well-understood, routine, conventional activities previously known to the industry, specified at a high level of generality, to the judicial exception, per MPEP 2106.05(d).
The additional element of a set of pixels (claims 1, 12, 13, 14, 22, and 23) falls within the guidelines for conventional computer components and/or functions (see MPEP at 2106.05(b) and 2106.05(d)(II) regarding conventionality of computer components and computer processes).
The additional elements of generating a count that is performed within 5 seconds (claim 2) and generating a count in less than 90 seconds (claim 18) do not amount to more than mere instructions to implement an abstract idea on a computer (MPEP 2106.05(f)), and therefore do not amount to significantly more than the judicial exception.
The additional element of light sheet scanning (claims 1 and 19) is conventional. Evidence is shown by Dell’Olio et al. (Laser Photonics, 2021, Vol. 15, pp. 1-29; as cited in the Office action mailed 03 June 2025). Dell’Olio et al. reviews photonic technologies for liquid biopsies (Title), and shows schematic descriptions of several advanced forms of high-resolution microscopy (Figure 2) including Bessel-beam microscopy wherein a highly focused, elongated excitation beam is swept across the sample, resulting in an extremely thin light sheet, and then the fluorescence is collected by a second microscopy objective lens held at 90 degrees with respect to the beam (Figure 2 E)); and further shows that one of the most used platforms for detecting point mutations in ctDNA, the ddPCR implemented in the commercial QX 200 system, utilizes fluorescence measurements to determine which droplets contain a nucleic acid target (page 21, column 1, para.1).
The additional elements of target molecules of a sample distributed across a set of partitions (claim 1); wherein each partition of the set of partitions contains less than two target molecules (claim 1); wherein the set of partitions are stabilized in position within a container (claim 1); wherein the set of partitions comprises at least 20 million partitions (claim 3); and wherein the container comprises a volumetric capacity of 50 microliters, and wherein the set of partitions comprises at least 30 million partitions (claim 5); are conventional. Evidence for the conventionality is shown by Xu et al. (Lab Chip, 2023, Vol. 23, pp. 1258-1278; as cited in the Office action mailed 03 June 2025). Xu et al. reviews advances in droplet digital polymerase chain reaction (Title) and discusses droplet generation methods and signal counting approaches, as well as applications in the fields of single-cell analysis, disease diagnosis, and pathogen detection (Abstract). Xu et al. further shows passive generation (Section 2.1) and active generation (Section 2.2) of droplets, and in particular shows an active generation process for centrifugal microdroplet generation wherein the emulsion is stably sitting at the bottom of a microcentrifuge tube after centrifugation (Fig. 3D (a)-(f)). Xu et al. further shows an example of a technology that involves partitioning a sample into millions of picoliter-sized droplets (Fig. 9), and that a droplet can theoretically contain at most one nucleic acid, although in practice it may contain more than one (Section 6: para. 1).
Therefore, when taken alone, all additional elements in claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 do not amount to significantly more than the above-identified judicial exception(s). Even when evaluated as a combination, the additional elements fail to transform the exception(s) into a patent-eligible application of that exception. Thus, claims 1-3, 5-9, 12-14, 16, 18-20, and 22-24 are deemed to not contribute an inventive concept, i.e., amount to significantly more than the judicial exception(s) (MPEP 2106.05(II)).
[Step 2B: NO]
Response to Arguments
The Applicant’s arguments/remarks received 24 November 2025 have been fully considered, but are not persuasive.
The Applicant states on page 7 of the Remarks that the instant claims are patent-eligible, for at least the reason that the instant claims are not directed to a judicial exception, and that furthermore, even if the claims were directed to a judicial exception, the claims recite additional elements that integrate the limitations into a practical application and contribute an inventive concept that amounts to significantly more than the judicial exception.
These arguments are not persuasive, because first, the foregoing arguments do not provide an explanation of which additional elements are purported to integrate the judicial exceptions into a practical application, and second, the foregoing arguments do not provide an explanation of which additional elements are purported to provide an inventive concept. Third, a claim is directed to a judicial exception when the claim is identified as reciting a judicial exception at Eligibility Step 2A Prong One, and when there are not any additional elements identified at Eligibility Step 2A Prong Two that are determined to integrate the judicial exception into a practical application, and when those additional elements are imported to Step 2B and determined to not provide an inventive concept. Thus, as noted and explained in the above rejection, the instant claims are directed to a judicial exception(s).
Claim Rejections - 35 USC § 103
The amendment and arguments/remarks received 24 November 2025 have been fully considered, however after further consideration, the rejections under 35 U.S.C. 103 in the Office action mailed 03 June 2025 are maintained in view of the amendment and arguments/remarks, as noted below.
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.
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.
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.
Claims 1, 3, 5-9, 12-14, 16, 19, and 24 are rejected under 35 U.S.C. 103 as being unpatentable over Shum et al. (Analytical Chemistry, Published 12 December 2022, Vol. 94, pp. 17868-17876, as cited in the Office action mailed 13 August 2024) in view of Liao et al. (PNAS, 2020, Vol. 117, No. 41, pp. 25628-25633, as cited in the Office action mailed 13 August 2024, with Supporting Information, pp. 1-37, as cited in the Office action mailed 03 June 2025) in view of Lai et al. (as cited in the Office action mailed 06 December 2024, and as cited in the Information Disclosure Statement (IDS) received 12 November 2024).
Regarding independent claim 1, Shum et al. shows a next-generation digital PCR (dPCR) system where DNA counting is performed in a single-molecule regimen through a 6-log dynamic range with each reaction divided into >30 million partitions (Abstract). Shum et al. further shows droplets with single-molecule occupancy (page 17868, col. 2, para. 3); partitions have no relative motion within the PCR tube and can be directly scanned inside the strip tubes after PCR via 3D light sheet microscopy (page 17871, col. 1, para. 1); coefficient of variation less than 7% (Table 1); and accuracy of the assay that is greater than 95% (Figure 6(E)).
Regarding dependent claim 3, Shum et al. further shows that every sample is partitioned into >30 million droplets (page 17868, column 2, para. 3).
Regarding dependent claim 5, Shum et al. further shows that every 50 μL sample is partitioned into >30 million droplets, enabling single-molecule occupancy, and that the system is extendable beyond 1 million DNA molecules (page 17868, column 2, para. 3).
Regarding dependent claims 6-9 and 19, the broadest reasonable interpretation of a method (or process) claim having contingent limitations requires only those steps that must be performed and does not include steps that are not required to be performed because the condition(s) precedent are not met (see MPEP 2111.04 II). Since there is no requirement that the number of target molecules is within any of the first, second, or third range, these limitations are not required (claims 6-9). Similarly, there is no requirement for enumerating positive partitions unless or until the limitation reciting “upon scanning a set of sheets of partitions spanning less than 25% of an internal volume of the container using light sheet scanning” is performed (claim 19). Therefore, claims 6-9 and 19 are rejected for the same reasons discussed above for claim 1.
Regarding dependent claim 24, Shum et al. further shows a serial dilution experiment targeting a 6-log dynamic range (i.e., 1 million) for single-molecule counting (page 17871, col. 1, para. 5).
Shum et al. does not show identifying partitions comprising a target molecule based on satisfaction of a signal criterion, an optical property criterion, a shape criterion, and a morphology criterion (claim 1); and wherein the signal intensity criterion is evaluated for a set of pixels depicting a partition of the set of partitions, wherein the signal intensity criterion is based upon a maximum signal intensity of the set of pixels (claim 1); or wherein the optical property criterion is evaluated for a set of pixels depicting a droplet of the set of partitions, and wherein the optical property criterion is based upon a lateral intensity gradient across the set of pixels (claim 12); or wherein the shape criterion is evaluated for a set of pixels depicting a partition of the set of partitions, and wherein the shape criterion is based upon a droplet radius determined from the set of pixels and a partition eccentricity determined from the set of pixels (claim 13); or wherein the morphology criterion is evaluated for a set of pixels depicting a partition of the set of partitions, and wherein the morphology criterion is based upon determination of a correlation with a three-dimensional profile of a representative positive partition determined from a set of control samples (claim 14); or wherein the container comprises a width and a lateral axis referenced to a center of the container, and wherein generating the count of the number of target molecules of the sample distributed across the set of partitions comprises omitting processing of a subset of sheets positioned away from the center of the container along the lateral axis (claim 16).
Regarding independent claim 1, Laio et al. shows three-dimensional digital PCR through light-sheet imaging of optically cleared emulsion (Title) and further shows the positive fluorescent droplets showed higher fluorescence intensity than either negative droplets or the backgrounds, and therefore searching local intensity maxima (voxels whose intensity is greater than those surrounding voxels) in the whole 3D space (Supplemental Information, page 5, #3); and further shows several digital image filters for identifying real positive droplets including an intensity filter (analogous to the instant claimed signal intensity criterion); a contrast filter (analogous to the instant claimed optical property criterion); a template matching filter (analogous to the instant claimed shape criterion); and a distance filter (analogous to the instant claimed morphology criterion); (Supplemental Information, pages 5-6).
Regarding dependent claim 12, Laio et al. further shows that a true positive droplet should have a normal contrast against its adjacent voxels, and the contrast filters detect the valley between droplet signal and background noise (Supplemental Information, page 6, Contrast filter) (i.e., a lateral intensity gradient across a set of pixels).
Regarding dependent claim 14, Laio et al. further shows that to filter out artifact signals, a droplet template was generated based on manually selected voxels of several ground-truth droplets, and then correlated with each identified local maximum to evaluate the resemblance to a ground-truth spherical (3D) droplet (Supplemental Information, page 6, Template matching filter).
Shum et al. in view of Laio et al. does not show wherein the shape criterion is evaluated for a set of pixels depicting a partition of the set of partitions, and wherein the shape criterion is based upon a droplet radius determined from the set of pixels and a partition eccentricity determined from the set of pixels (claim 13); or wherein the container comprises a width and a lateral axis referenced to a center of the container, and wherein generating the count of the number of target molecules of the sample distributed across the set of partitions comprises omitting processing of a subset of sheets positioned away from the center of the container along the lateral axis (claim 16).
Regarding dependent claim 13, Lai et al. shows methods and systems for three-dimensional light sheet imaging (Title; and Abstract); and further shows the resulting cross-sectional plane images were combined to produce a three-dimensional image data set for each sample (para. [0101]); and a three-dimensional (3D) convolution was performed using a spherical template to identify substantially spherical fluorescent particles (corresponding to fluorescent lipid droplets) within an expected size range and eliminate fluorescent particles that do not conform to the expected size and shape parameters; and a 3D local maxima analysis was performed to identify potential fluorescent particle candidates having a positive fluorescent signal, and an additional 3D local maxima analysis was performed to filter out asymmetric particle candidates (e.g., non-spherical particle candidates) (para. [0102]).
Regarding dependent claim 16, Lai et al. further shows wherein the position of the tube cross-sectional plane image remains centered within the camera field of view during translation (para. [0099]); and further shows image processing methods to identify and quantify fluorescent particles within a three-dimensional image data set (para. [0055]); the resulting cross-sectional plane images may be combined to produce a three-dimensional image data set for each sample, and a region of interest within each cross-section may be selected outlining the confines of the sample (FIG. 5A) that may ensure that only the fluorescent particles inside the tube cross-section were considered (para. [0056]).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified Shum et al. by incorporating image processing methods to identify and quantify positive fluorescent droplets because Liao et al. shows a computational procedure for image-based droplet counting to process the raw light sheet fluorescent microscopy images (Methods, page 25632, col. 2, para. 5). One of ordinary skill in the art would have been motivated to modify Shum et al. with image-based droplet counting computational procedures, because Liao et al. shows that accurately identifying all of the positive droplets requires multiple filters to rule out those droplets that are not real positive droplets (Supplemental Information, page 5, para. 4). This modification would have had a reasonable expectation of success because both Shum et al. and Liao et al. utilize light-sheet fluorescence microscopy to acquire three-dimensional images of droplets to achieve digital counting of the positive droplets.
It would have been further obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified Shum et al. in view of Liao et al. by incorporating methods for automated detection and quantification of fluorescently labeled droplets imaged using light sheet imaging as shown by Lai et al., and discussed above. One of ordinary skill in the art would have been motivated to modify Shum et al. in view of Liao et al. with the automated detection and quantification methods shown by Lai et al. because Lai et al. shows that compared to one-dimensional and two-dimensional imaging techniques, three-dimensional imaging techniques have many advantages, such as higher throughput imaging methods and sequential imaging of multiple three-dimensional samples without user intervention, however compared to one-dimensional and two-dimensional imaging techniques, three-dimensional imaging techniques may have additional complications due to processing image data sets, such as uneven background intensities, uneven illumination intensity, or signal variability at different cross-sectional planes or positions within a cross-sectional plane. This modification would have had a reasonable expectation of success because both Shum et al. in view of Liao et al. and Lai et al. utilize light-sheet fluorescence microscopy to acquire three-dimensional images of droplets to achieve digital counting of the positive droplets, and Lai et al. further shows image processing methods for fluorescent particle quantification using a three-dimensional template to identify fluorescent particles substantially conforming to a desired shape and within an expected size range (para. [0057]).
Claims 2 and 18 are rejected under 35 U.S.C. 103 as being unpatentable over Shum et al. in view of Liao et al. in view of Lai et al. as applied to claims 1, 3, 5-9, 12-14, 16, 19, and 24 above, and further in view of Shum et al. (United States Patent Application Publication No.: US 2022/0389410, as cited in the Information Disclosure Statement (IDS) received 01 August 2024; previously cited).
Shum et al. in view of Liao et al. in view of Lai et al. as applied to claims 1, 3, 5-9, 12-14, 16, 19, and 24 above, do not show that generating the count is performed within 5 seconds (claim 2) or in less than 90 seconds (claim 18).
Regarding dependent claim 2, Shum et al. (US 2022/0389410) shows generating a plurality of droplets within a collecting container at an extremely high rate, e.g., of at least 1 million droplets per minute (Abstract); and that readout associated with digital analyses, e.g., counting, enumeration, etc., can be performed within a duration of 5 seconds or less (para. [0120]).
Regarding dependent claim 18, Shum et al. (US 2022/0389410) shows generating a plurality of droplets within a collecting container at an extremely high rate, e.g., of at least 1 million droplets per minute (Abstract); and that readout associated with digital analyses, e.g., counting, enumeration, etc., can be performed within a duration of 1 minute, 30 seconds, 20 seconds, 10 seconds, 5 seconds or less (para. [0120]).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to have modified Shum et al. in view of Liao et al. in view of Lai et al. as applied to claims 1, 3, 5-9, 12-14, 16, 19, and 24 above, by incorporating methods for digital analyses (e.g., counting, quantification, etc.) of readouts of fluorescent signals generated by a 3D scanning technique that can be performed within a duration of less than 90 seconds or within 5 seconds. One of ordinary skill in the art would have been motivated to modify Shum et al. in view of Liao et al. in view of Lai et al. as applied to claims 1, 3, 5-9, 12-14, 16, 19, and 24 above, because Shum et al. (US 2022/0389410) shows that readout associated with digital analyses, e.g., counting, enumeration, etc., can be performed within a duration of 5 seconds or less (para. [0120]). This modification would have had a reasonable expectation of success because both Shum et al. in view of Liao et al. in view of Lai et al. as applied to claims 1, 3, 5-9, 12-14, 16, 19, and 24 above, and Shum et al. (US 2022/0389410; at para. [0118]) show the utilization of light-sheet fluorescence microscopy to acquire three-dimensional images of droplets to achieve digital counting of the positive droplets.
Response to Arguments
The Applicant’s arguments/remarks received 24 November 2025 have been fully considered, but are not persuasive.
The Applicant states on page 7 (bottom) and page 8 (top) of the Remarks that claim 1 is not obvious over Shum because the relied upon portions of Shum do not recite every element of the claim, and neither the relied upon portions of Liao nor Lai cure this deficiency, and accordingly, claim 1 is not obvious over the combination of cited references. The Applicant further states (page 8, para. 2) that claims 3, 5-9, 12-14, 16, 19, and 24 are not obvious over the combination of cited references for at least the reason that these claims depend from and include all of the elements of claim 1 and recite additional elements of particular advantage and utility.
These arguments are not persuasive, because first, the primary reference of Shum is not relied upon to show each and every limitation of claim 1, as noted and explained in the above rejection. Second, the secondary references are relied upon to show the limitations identified in the rejection as not being shown by Shum, e.g., the secondary references of Laio and Lai are cited to show that the limitations of “identifying partitions of the set of partitions comprising a target molecule based on satisfaction of a signal criterion, an optical property criterion, a shape criterion, and a morphology criterion,” e.g., are obvious even though the secondary references describe the limitations with descriptive language that differs from the instant claim terms, as shown and explained in the rejection above.
That Applicant states on page 8 (para. 5) of the Remarks that claims 2 and 8 are not obvious over the relied upon portions of the cited references, for at least the reasons provided in the foregoing argument, and further because the cited reference Shum2 does not cure this deficiency.
These arguments are not persuasive, first because of the reasons given in the foregoing response to arguments, and second, because the further in view of reference (i.e., Shum2) shows the limitations of instant claims 2 and 8, as noted and as explained in the above rejection.
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.
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/S.W.B./Examiner, Art Unit 1687
/Joseph Woitach/Primary Examiner, Art Unit 1687