DETAILED ACTION
The present Office Action is responsive to the Amendment received on September 10, 2025.
Preliminary Remark
Claims 13 and 15 are canceled.
Claims 21 and 22 are new.
Information Disclosure Statement
The IDS received on September 10, 2025 is proper and is being considered by the Examiner. The IDS was received with the fee under 37 CFR 1.17(p).
Specification
The objection made to the specification for reasons set discussed in the Office Action mailed on June 11, 2025 is withdrawn in view of the Amendment received on September 10, 2025.
Claim Objections
The objection made to claim 2 for the reasons discussed in the Office Action mailed on June 11, 2025 is withdrawn in view of the Amendment received on September 10, 2025.
Claim Rejections - 35 USC § 112
The rejection of claims 7 and 13 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to particularly point out and distinctly claim the subject matter, made in the Office Action mailed on June 11, 2025 is withdrawn in view of the Amendment received on September 10, 2025.
Rejection – New Grounds, Necessitated by Amendment
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 21 and 22 are rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 21 is indefinite because the claim recites a Markush group, but the members of the Markush are not definite. Specifically, the claim recites that the members of the Markush includes “other means of localized/clonal amplification or wherein the amplification process is based on the use of exonuclease, endonuclease, transposase and CRISPR/Cas-based enzymatic activity prior to amplification”
The usage of “or” when reciting the members of a Markush claim is indefinite because the set of members must be a specific set, utilizing the conjunction, “and”. Additionally, the breadth covered by the member, “other means of localized amplification” entirely too broad to the extent unclear what such means can be compatible with the claimed method. As well, the phrase, “amplification process is based on … prior to amplification”. The claim is defining a Markush group that amplifies the target biomolecules. Therefore, any members reciting steps that occur “prior to amplification” cannot be listed as a member of this Markush group.
Claim 22 recites the phrase, “method can simultaneously detect” does not actively limit the parent claim and if it does, unclear just exactly how the claim further limits the parent claim.
Claim Rejections - 35 USC § 102
The rejection of claim 13 under 35 U.S.C. 102(a)(1) as being anticipated by Wang et al. (Bioconjugate Chem., 2007, vol. 18, pages 297-301), made in the Office Action mailed on June 11, 2025 is withdrawn in view of the Amendment received on September 10, 2025, canceling the rejected claim.
Rejection - Maintained
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
The rejection of claims 1-10, 13, 14, 16, 19, and 20 under 35 U.S.C. 102(a)(1) as being anticipated by Wang et al. (Bioconjugate Chem., 2007, vol. 18, pages 297-301), made in the Office Action mailed on June 11, 2025 is maintained for the reasons of record.
Applicants’ arguments presented in the Amendment received on September 10, 2025 have been carefully considered but they have not been found persuasive for the reasons discussed in the, “Response to Arguments” section.
The Rejection:
With regard to claim 1, Wang et al. teach a method for multiplexed detection of a plurality of target biomolecules using optical encoding, wherein each target biomolecule has at least one detection target, comprising the steps of:
providing a plurality of nanoparticle types comprising a plurality of nanoparticles (“[w]e have developed triple-dye-doped FRET silica nanoparticles (NP) … to serve as promising substrates for multiplexed bacteria monitoring”, page 297, 1st column), each nanoparticle having a coating that provides binding affinity of the nanoparticle to a type-specific detection target (see Figure 1, strategies A, B, and C, “[c]arbodiimide chemistry was used to cross-link free carboxylic acid groups on NPs with amine-containing antibodies … Streptavidin conjugation to NPs used the above procedure except that the antibody was changed to streptavidin … biotin-labeled antibody weas added … Amine-modified NPs were reacted with the activated ester of biotin … Streptavidin and NP-biotin were then mixed”, page 298, 1st column), and wherein each nanoparticle comprises a plurality of fluorophores that generates a signal which is unique for each nanoparticle type (“the TMR-APTS conjugate or the three tandem dye-APTS conjugates (FAM-APTS, R6G-APTS, ROX-APTS) mixed at desired ratios (1:0.5:1, 0.5:1:4, 0.5:0.5:3)”, page 298, 1st column; “worked described here assesses the use of FRET NPs for multiplexed bacterial detection … results show that FRET NPs can be used as highly sensitive and selective platforms for sensing multiple microorganisms simultaneously”, page 297, 2nd column);
providing a sample comprising a plurality of target biomolecules (“[a]ntibody-labeled NPs and bacteria were mixed at an experimentally optimized ratio”, page 298, 2nd column);
contacting the sample with a plurality of nanoparticle types, thereby allowing the nanoparticles to bind with the detection targets of the target biomolecules (see above); and
optically decoding the fluorophore signals emitted by the nanoparticle of the nanoparticle types bound to the detection target of the target biomolecules by measuring the wavelength and intensity of the emitted signals thereby detecting the presence and identity of the target biomolecules (“[s]imultaneous multiple bacterial monitoring was achieved using color-encoded FRET NPs … In the FRET NPs, FAM was used as a common donor for R6G and ROX, while R6G acted as both an acceptor for FAM and donor for ROX. By exciting at the maximum excitation wavelength for FAM (488 nm), efficient energy transfer between the three dyes in the same NP occurred, and NPs exhibited three emission peaks corresponding to the three encapsulated dyes”, page 299, 2nd column; “we have been the first to apply multicolor FRET NPs for multiplexed monitoring of bacteria species … fast and accurate with high sensitivity”, page 300, 1st column).
With regard to claim 2, each nanoparticle type is optically encoded by comparing controlled ratios of the plurality of fluorophores, thereby controlling the emission wavelength and intensity from the nanoparticle type, or altering the properties of the fluorophore affecting its emission intensity (“by varying the ratios of three fluorescence energy transfer tandem dyes coencapsulated into the silica NPs, excitation with a single wavelength leads to emission of numerous different colors”, page 297, 1st column).
With regard to claim 3, the binding affinity of the coating of the nanoparticle type is provided by a detection probe X (the antibody) attached via a linker to the nanoparticle, wherein the detection probe X is chosen from nucleic acid molecule, an antigen, or an antibody (see above for linkage description).
With regard to claim 4, the coating of the nanoparticle comprises functional group Y (biotin) via a linker (PEG) to the nanoparticle, wherein the group Y is negatively charged (biotin is negatively charged with -COO- at reaction pH).
With regard to claim 5, the linker (PEG) comprises an anchor group that tether the coating to the nanoparticle (“Amine-modified NPs were reacted with the activated ester of biotin (NHS-PEG500-biotin) in the phosphate buffer … to couple biotin molecules to the NPs”, page 298, 1st column).
With regard to claim 6, more than one linkers are provided on each NPs to immobilized more than one probe per nanoparticle (see above).
With regard to claim 7, the plurality of fluorophores are organic fluorophores (FAM, ROX, and R6G).
With regard to claims 8-10 and 19, the nanoparticles are silica particles (“dye-doped NPS, each encapsulating ~10,000 dye molecules in a ~60 nm silica sphere”, page 297, 1st column).
With regard to claims 13 and 20, the detection is via optical decoding (see above and “[o]ptical and fluorescence images were obtained…”, page 297, 2nd column).
With regard to claim 14, the method employes three different NPs labeled with probes (see above).
With regard to claim 16, the linker is a spacer group (PEG).
Therefore, the invention as claimed is deemed anticipated by Wang et al.
Response to Arguments:
Applicants traverse the rejection.
Applicants contend that Wang et al. fail to teach measuring both the wavelength and intensity of the signals emitted by the nanoparticles (page 12, bottom paragraph, Response), and that visually analyzing color emitted by the nanoparticles is not equivalent to “optically decoding the fluorophore signals emitted by the nanoparticle of the nanoparticle types bound to the detection target of the target biomolecules” as presently claimed (page 12, bottom paragraph, Response).
Applicants further state that claim 1 has also been amended to recite that the method further involve the step of, “optically decoding the fluorophore signals emitted by the nanoparticle of the nanoparticle types bound to the detection target of the target biomolecules by measuring the wavelength and intensity of the emitted by signals to ratiometrically identify the optical code incorporated in the nanoparticle types”, and that this optical encoding is uniquely separable when decoded due to the unique combination of wavelength and intensity of the emitted signals being analyzed (page 13, Response).
These arguments have been carefully considered but they have not been found persuasive for the following reasons.
As to Applicants’ contention that Wang fail to teach measuring both the wavelength and intensity of the signals emitted by the nanoparticles, wherein visually analyzing color emitted by the nanoparticle is “not equivalent” to an “optical decoding”, the Office respectfully disagrees.
The act of decoding results in the identification of which targets are bound by which probes comprised by dye-doped. And as shown by Wang et al., nanoparticles which comprises a specific target binding probe are doped with a combination of fluorophores, wherein the varying ratio in the combination of said fluorophores result in different exhibited color (or wavelengths):
“we have developed triple-dye-doped NPs [nanoparticles], each encapsulating ~10 000 dye molecules … by varying the ratio of the three fluorescent energy transfer tandem dyes coencapsulated into the silica NPs, excitation with a single wavelength leads to emission of numerous different colors, thereby permitting simultaneous and sensitive detection of multiple targets” (page 297, 1st column)
“Simultaneous multiple bacteria monitoring was achieved using color-encoded FRET NPs. To prepare the FRET NPs, the three amine-reactive energy-transfer tandem dyes (FAM-SE, R6G-SE, ROX-SE) were first covalently linked to the silane coupling agent APTS. The three APTS-dye conjugates were then mixed at desired ratios (1:0.5:1, 0.5:1:4, 0.5:0.5:3) … By exciting at the maximum excitation wavelength for FAM (488 nm), efficient energy transfer between the three dyes in the same NP occurred, and NPs exhibited three emission peaks corresponding to the three encapsulated dyes, while the combinatorial color varies due to the different dye doping ratio.” (page 299, 2nd column)
Therefore, contrary to Applicants’ characterization of Wang’s teachings, the artisans teach signals that result from emission wavelengths from the ratios of the dyes present in the nanoparticles, wherein the intensity of the signal is based on the varying amounts of the dyes co-encapsulated in the nanoparticle.
Therefore, the resulting signals captured by the camera is based on the measurement of the wavelength and intensity of the signals produced from the NPs, and the color/signal observed is based on the different ratio of fluorescent dyes that are encapsulated in the NPs.
And because the varying ratios of dyes that are doped into the NPs, the resulting optical encoding is uniquely separable when decoded due to the unique combination of wavelength and intensity of the emitted signals being analyzed (i.e., observed by different colors, see Figure 4, also “resultant bacteria-NP complexes shows that three bacteria species were specifically covered with their corresponding antibody-labeled NPs and exhibit the coding colors of the attached NPs”, page 300, 1st column).
If Applicants are contending that the actual determination involves the spectra and intensity of the each of the dyes in the encapsulated in nanoparticle must be determined when decoding, then this should be explicitly recited. Generically reciting that the optical decoding is ratiometrically made does not clearly distinguish a decoding via color that is produced from a varying “ratio” of dyes encompassed by the nanoparticles of Wang et la.
Therefore, Applicants’ arguments are not found persuasive and the invention as claimed is anticipated by Wang et al.
Claim Rejections - 35 USC § 103
Maintained & New, Necessitated by Amendment
The following is a quotation of 35 U.S.C. 103 which forms the basis for all obviousness rejections set forth in this Office action:
A patent for a claimed invention may not be obtained, notwithstanding that the claimed invention is not identically disclosed as set forth in section 102, if the differences between the claimed invention and the prior art are such that the claimed invention as a whole would have been obvious before the effective filing date of the claimed invention to a person having ordinary skill in the art to which the claimed invention pertains. Patentability shall not be negated by the manner in which the invention was made.
The rejection of claims 11 and 12 under 35 U.S.C. 103 as being unpatentable over Wang et al. (Bioconjugate Chem., 2007, vol. 18, pages 297-301) in view of Nallur et al. (Nucleic Acids Research, 2001, vol. 29, no. 23, e118, pages 1-9), made in the Office Action mailed on June 11, 2025 is maintained for the reasons of record.
In addition, claims 21 and 22 are rejected herein as being necessitated by Amendment (its addition).
Applicants do not present any new arguments for the instant rejection but rely solely on their arguments presented for the anticipation rejection over Wang et al. which have been fully responded to above.
Therefore, the rejection is maintained for the reasons already of record.
The Rejection:
The teachings of Wang et al. have already been discussed above.
As previously discussed, Wang et al. already teach that a combination of fluorescent dyes embedded in a nanoparticle can be utilized to generate unique emission fluorescence for multiple analyte detection.
Wang et al. do not employ their method for the detection of target nucleic acid, and therefore, do not perform an RCA for detecting the RCA products, utilizing their nanoparticle labeled probes (claims 11 and 12).
Nallur et al. teach a method of detecting a target nucleic acid via use of RCA, wherein the artisans generate an RCA product, which is detected with a labeled probe (see Figure 1, also below):
“Microarrays containing immobilized P1 ligation oligonucleotides were incubated with 8 ml of a solution containing 200 nM P2 (one each for the wild-type and mutant alleles) …” (page 3, 1st column)
“an immobilized oligonucleotide probe (P1), complementary to the target sequence and containing a phosphate group at the 5’ terminus, underwent allele-specific ligation to one of a pair of oligonucleotide probes (P2mu and P2wt) in the presence of oligonucleotide target. The P2 probes posses two 3’ ends, by virtue of reversed back-bone synthesis; one end that is complementary to a 20 base sequence of the target contiguous with P1 and the other end having a sequence that does not hybridize with the target, but hybridizes to an RCA circle. At the gene-specific end, the two P2 probes differ in sequence by a single 3’-terminal nucleotide. After hybridization of the target to the immobilized P1 probes, a thermostable DNA ligase joins the cognate P2 to P1 probe thereby immobilizing P2 on the microarray … two circles containing sequences complementary to P2wt or P2mut, respectively are hybridized with the array. RCA is performed and the amplified product from the wild-type primer and circle is decorated by hybridization with an oligonucleotide labeled with Cy5, while the RCA product amplified from the mutant primer and circle is decorated by hybridization with an oligonucleotide labeled with Cy3” (page 6, 2nd column, bottom paragraph to page 7, 1st column, 1st paragraph)
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 combine the teachings of Wang et al. and Nallur et al., thereby arriving at the invention as claimed for the following reasons.
The motivation to do so is provided by the rationale of obviousness discussed by the Supreme Court in KSR, wherein the Supreme Court particularly emphasized “the need for caution in granting a patent based on the combination of elements found in the prior art,” Id. at 415, 82 USPQ2d at 1395, and discussed circumstances in which a patent might be determined to be obvious. Importantly, the Supreme Court reaffirmed principles based on its precedent that “[t]he combination of familiar elements according to known methods is likely to be obvious when it does no more than yield predictable results.” Id. at 415-16, 82 USPQ2d at 1395. The Supreme Court stated that there are “[t]hree cases decided after Graham [that] illustrate this doctrine.” Id. at 416, 82 USPQ2d at 1395. (1) “In United States v. Adams, . . . [t]he Court recognized that when a patent claims a structure already known in the prior art that is altered by the mere substitution of one element for another known in the field, the combination must do more than yield a predictable result.”
While Wang et al. did not explicitly teach that their labels can be used for detection of other target analytes, one of ordinary skill in the art would have recognized that the nanoparticles of Wang et al. which generate different emission spectra would have also yielded the predictable outcome of providing distinguishing detection signals when used to as a label in other types of detection assays. Given that Nallur et al. teach a method of amplifying a target nucleic acid via RCA, wherein multiple RCA products are detected with two different target specific probes labeled with different fluorescent moieties, one of ordinary skill in the art would have been motivated to take the teachings of Wang et al. to detect multiple target nucleic acid from RCA reaction products with using the uniquely labeled nanoparticles.
Therefore, the invention as claimed is deemed prima facie obvious over the cited references.
Response to Arguments:
With regard to claim 21, the rationale to amplify the target molecules before the detection means of Wang et al. is based on the conventionally adopted practice for amplifying starting target molecules via thermocycling or isothermal means (such as PCR, or RCA), wherein the motivation for doing so is for the advantage of increasing the sensitivity of the assay.
With regard to claim 22, it is asserted that the method of Wang et al. can detect as many as at least 50 target biomolecules as the different dye ratio and variety can be employed to detect at least 50 same target biomolecules or 50 different target biomolecules. One of ordinary skill in the art would have had a reasonable expectation of success at doing so as Wang et al. already provided a framework from which different optical signals can be generated (by doping varying ratios of different combinations of dyes onto nanoparticles) and experimenting varying ratios and combinations of different fluorescent dyes which are distinguishable would have involved routine.
The rejection of claims 17 and 18 under 35 U.S.C. 103 as being unpatentable over Wang et al. (Bioconjugate Chem., 2007, vol. 18, pages 297-301) in view of Xu et al. (Nucleic Acids Research, 2003, vol. 31, no. 8, e43, pages 1-10), made in the Office Action mailed on June 11, 2025 is maintained for the reasons of record.
Applicants do not present any new arguments for the instant rejection but rely solely on their arguments presented for the anticipation rejection over Wang et al. which have been fully responded to above.
Therefore, the rejection is maintained for the reasons already of record.
The Rejection:
The teachings of Wang et al. have already been discussed above.
As previously discussed, Wang et al. already teach that a combination of fluorescent dyes embedded in a nanoparticle can be utilized to generate unique emission fluorescence for multiple analyte detection:
While Wang et al. provide a combination of three different organic dyes in varying ratios to achieve this means, the artisans do not explicitly disclose that other fluorescent dye combinations can be utilized for the same purpose, such as those listed in claim 17, or inorganic fluorophores (claim 18).
Xu et al. teach a method of using quantum dot-encoded microsphere, wherein the artisans teach that a combination of different quantum dots can be encoded into microspheres to create a unique code for microsphere identification:
“we describe a new method for multiplexed SNP genotyping using the Qbead system … Qbead system uses combinations of different Qdot nanocrystals with various emission wavelengths and intensities to create unique codes for microsphere identification” (page 6, 1st column, bottom paragraph to 2nd column, 1st paragraph)
“The broad excitation spectra and narrow emission spectra of the system provides much more potential upside in terms of multiplexing since thousands more spectral codes are possible using nanocrystals instead of traditional dyes” (page 7, 2nd column)
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 combine the teachings of Wang et al. and Xu et al., thereby arriving at the invention as claimed for the following reasons.
The motivation to do so is provided by the rationale of obviousness discussed by the Supreme Court in KSR, wherein the Supreme Court particularly emphasized “the need for caution in granting a patent based on the combination of elements found in the prior art,” Id. at 415, 82 USPQ2d at 1395, and discussed circumstances in which a patent might be determined to be obvious. Importantly, the Supreme Court reaffirmed principles based on its precedent that “[t]he combination of familiar elements according to known methods is likely to be obvious when it does no more than yield predictable results.” Id. at 415-16, 82 USPQ2d at 1395. The Supreme Court stated that there are “[t]hree cases decided after Graham [that] illustrate this doctrine.” Id. at 416, 82 USPQ2d at 1395. (1) “In United States v. Adams, . . . [t]he Court recognized that when a patent claims a structure already known in the prior art that is altered by the mere substitution of one element for another known in the field, the combination must do more than yield a predictable result.”
Wang et al. already teach that a combination of optical labels such as organic fluorescent labels can be combined in different ratios in order to arrive at a unique emission spectrum, allowing for highly multiplexed detection capabilities.
Such a teaching would have provided the ordinarily skilled artisan to readily recognize that other types of dyes could have been utilized for the same purposes, with the predictable outcome, that is, expanding the multiplex capability in the detection method of Wang et al. with combination of other fluorescent labels commercially available in the art (i.e., the list of dyes in claim 17).
Similarly, based on the disclosure of Xu et al. who teach that quantum dots can also be combined into a bead in order to generate unique emission spectra, for the same purpose of expanding multiplex detection capabilities, one of ordinary skill in the art would have been motivated as well as would have had a reasonable expectation of success at utilizing a combination of inorganic labels, such as quantum dots, for the method of detection of Wang et al.
Therefore, the invention as claimed is deemed prima facie obvious over the cited references.
Conclusion
No claims are allowed.
The Office also notes that the decoding based on wavelength determination and their intensity is also taught by Ho, Winston (US 2002/0164271 A1, published November 7, 2002; IDS ref) and may be material to the invention should amendment make it necessary.
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.
Inquiries
Any inquiry concerning this communication or earlier communications from the Examiner should be directed to Young J. Kim whose telephone number is (571) 272-0785. The Examiner can best be reached from 7:30 a.m. to 4:00 p.m (M-F). The Examiner can also be reached via e-mail to Young.Kim@uspto.gov. However, the office cannot guarantee security through the e-mail system nor should official papers be transmitted through this route.
If attempts to reach the Examiner by telephone are unsuccessful, the Examiner's supervisor, Gary Benzion, can be reached at (571) 272-0782.
Papers related to this application may be submitted to Art Unit 1681 by facsimile transmission. The faxing of such papers must conform with the notice published in the Official Gazette, 1156 OG 61 (November 16, 1993) and 1157 OG 94 (December 28, 1993) (see 37 CFR 1.6(d)). NOTE: If applicant does submit a paper by FAX, the original copy should be retained by applicant or applicant’s representative. NO DUPLICATE COPIES SHOULD BE SUBMITTED, so as to avoid the processing of duplicate papers in the Office. All official documents must be sent to the Official Tech Center Fax number: (571) 273-8300. Any inquiry of a general nature or relating to the status of this application should be directed to the Group receptionist whose telephone number is (571) 272-1600.
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/YOUNG J KIM/Primary Examiner
Art Unit 1637 December 1, 2025
/YJK/