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 .
Priority
Applicant’s claim for the benefit of a prior-filed application under 35 U.S.C. 119(e) or
under 35 U.S.C. 120, 121, 365(c), or 386(c) is acknowledged. This application is a 371 of
PCT/CL2021/050039 filed 05/14/2021 which claims benefit of Application No. 63/025,534 filed
05/15/2020. Based on the filing receipt, the effective filing date of this application is May 15,
2020 which is the filing date of Application No. 63/025,534 from which the benefit of priority is
claimed.
Withdrawn Objections/Rejections
The objection to the misnumbered claims 3-10 has been withdrawn, due to the cancellation of the claims in the amended claims filed 02/17/2026.
The objection to claims 1-11 for unconventional claim language has been withdrawn, necessitated by amendments filed 02/17/2026. In the amended claims, the unconventional claim language has been removed.
The objection to claim 1 for a clerical error has been withdrawn, necessitated by the claim amendments filed 02/17/2026 which removed the error.
The objection to claim 4 for a clerical error has been withdrawn, necessitated by the claim amendments filed 02/17/2026 which removed the error.
The rejection of claims 1-11 under 35 U.S.C. 112(b) for insufficient antecedent basis regarding “the microorganisms” in step b of claim 1 has been withdrawn, necessitated by claim amendments filed 02/17/2026.
The rejections of claims 4 and 6 on the grounds of 35 U.S.C. 112(b) have been withdrawn, due to the cancellation of the claims in amendments filed 02/17/2026.
The rejections of claims 1-3 on the grounds of 35 U.S.C. 103 have been withdrawn, necessitated by claim amendments filed 02/17/2026. Specifically, the addition of the limitation “wherein the inert media is a medium capable of forming a density gradient after centrifugation” in independent claim 1 was not taught or suggested in the said rejection. New rejections to replace the withdrawn rejection are set forth below.
The rejections of claims 4-11 on the grounds of 35 U.S.C. 103 have been withdrawn, due to the cancellation of the claims in claim amendments filed 02/17/2026.
New Rejections and Objections
Claim Objections
Claim 1 is objected to because of the following informalities:
Claim 1 recites, “centrifuging for from 45 seconds to 2 minutes”. The claim should recite, “centrifuging for between 45 seconds to 2 minutes”.
Claim 1 recites, “at a rate at or between 150 and 250 g”. The claim should recite “at a rate of between 150 and 250 g”.
Appropriate correction is required.
Claim Rejections - 35 USC § 103
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, 12, and 18-19 are rejected under 35 U.S.C. 103 as being unpatentable over Salema, et al. (“Escherichia coli surface display for the selection of nanobodies”, published 2017, cited in IDS filed 11/14/2022) in view of Jung, et al. (“Binding and Enrichment of Escherichia coli Spheroplasts Expressing Inner Membrane Tethered scFv Antibodies on Surface Immobilized Antigens”, published 2007-03-09, cited in PTO-892 dated 08/14/2025), a ResearchGate comment from S Ashraf Imam (https://www.researchgate.net/post/what_is_the_centrifugationspeed_used_to_pelletize_living_cells#:~:text=it%20is%20pertinent%20to%20use,with%20constant%20acceleration%20and%20deceleration, published 2018, cited in PTO-892 dated 08/14/2025), Zhao (CN 110964101 A, published 2020-04-07, cited in PTO-892 dated 08/14/2025), Yan (US 20200150114 A1, published 2020-05-14, cited in PTO-892 dated 08/14/2025), and Richman, et al. (“Development of a novel strategy for engineering high-affinity proteins by yeast display”, published 2006-06, cited in PTO-892 dated 08/14/2025) as evidenced by the material safety data sheet of Agarose beads from Gold Biotechnology (https://goldbio.com/uploads/documents/1cffa7e57257420a72f637727ef2644a.pdf, cited in PTO-892 dated 08/14/2025) and the product data sheet of PBS buffer from Biosolve Chemical (https://shop.biosolve-chemicals.eu/detail.php?id=2093, cited in PTO-892 dated 08/14/2025).
With respect to claim 1, Salema teaches a method of separation of nanoantibodies (HHV) against a specific antigen from a nanoantibody expression library wherein the method comprises the following steps: (a) binding the antigen of interest to beads (b) incubating the microorganisms of the expression library with the antigen of interest, where the expression library corresponds to microorganisms transformed with the cDNA of fragments corresponding to the HHV domains of an HHV-producing animal previously immunized with that antigen of interest; (c) obtaining the beads with the linked microorganisms, which correspond to those that express HHV that recognizes the antigen (see, e.g., p. 1474-1475, under “Selection of Nbs by E. coli display using magnetic cell sorting (MACS)”, para. 1-2; and p. 1475, under “Fig. 4.”, panels “(A)” and “(B)”).
Selema fails to teach placing protein-binding polymer beads with the linked organisms in a tube with an inert medium with a density greater than or equal to 1 g/mL and centrifuging for 45 s to 2 minutes at a rate between 150 and 250 g, and discarding the upper fraction and supernatant with the free microorganisms, and obtaining the beads with the linked microorganisms, which correspond to those that express HHV that recognizes the antigen.
However, Jung, Zhao, Imam, Yan¸ and Richman rectify these deficiencies. Jung teaches placing antigen-binding polymer beads with the microorganisms expressing antibodies in a tube with an inert medium with a density greater than or equal to 1 g/mL and centrifuging for 3 minutes at a rate of 3,000 rpm, and discarding the upper fraction and supernatant with the free microorganisms, and obtaining the beads with the bound microorganisms, which correspond to those that express antibodies that recognizes the antigen (see, e.g., antigen-binding polymer beads – p. 42, col. 1, under “Digoxigenin Immobilization Onto Beads”; placing antigen-binding polymer beads with microorganisms expressing antibodies in a tube with an inert medium with a density greater than or equal to 1 g/mL and centrifuging for 3 minutes at a rate of 3,000 rpm discarding the upper fraction and supernatant with the free microorganisms, and obtaining the beads with the linked microorganisms, which correspond to those that express antibodies that recognizes the antigen – p. 42, col. 1-2, under “Preparation of Spheroplasts and FC Analysis”: “Three hundred microliters of the spheroplasts and 50 µL of the 1% (wt/vol) digoxigenin immobilized beads were mixed in a glass test tube and incubated at room temperature for 2 h with shaking and washed in 1 mL of PBS to remove unbound spheroplasts by centrifugation at 3,000 rpm for 3 min”). PBS has density greater than or equal to 1 g/mL as evidenced by the product sheet for PBS Buffer from Biosolve Chemical (see, e.g., under “Product Specification”, under “Density of 1X conc (25°C)”). It is understood that the spheroplasts bound to the beads will be denser than the supernatant and unbound spheroplasts, therefore, to remove the unbound spheroplasts by centrifugation, the upper fraction and supernatant will be discarded.
Zhao discloses the use of antigen-beads composed of Human serum albumin cross-linked agarose gel beads for separating nanobodies bound to antigen-beads from unbound nanoantibodies by centrifugation in PBS at 100 g for 2 min (see, e.g., p. 6, under “Example 4”, under “Nanobody biological characterization includes the following steps:”, under “(1)Species specific detection of anti-human serum albumin nanobody:”).
In a ResearchGate comment discussing centrifuging cells, Imam recommends “using 200 g centrifugation […] most cells do not require more than 5 minutes of centrifugation”.
In a patent application directed to a method for capturing target cells or molecules in solution, Yan discloses that density gradient centrifugation and magnetic activated cell sorting (MACS) are functional equivalents for sorting cells (see, e.g., para. [0003]). Yan teaches density gradient centrifugation technologies are simple to operate and enable the capture of cells (see, e.g., para. [0003]).
Selema, Jung, Imam, Zhao and Yan are analogous to the field of the claimed invention because they are all in the field of microbiology. One of ordinary skill in the art before the effective filing date of the application would have found it obvious to simply substitute the separation method of Salema with the separation method of Jung and Zhao and use the centrifugation rate recommended by Imam. An artisan would have been motivated to do so because Yan discloses that both separation methods are functional equivalents for sorting cells and density gradient centrifugation technologies have the advantage of being simple to operate and enable the capture of cells.
A skilled artisan would have had a reasonable expectation of success in using an inert medium with a density greater than or equal to 1 g/mL and centrifuging for between 45 seconds to 2 minutes at a rate between 150-250 g in the method of Selema as modified by Jung and Zhao, Imam and Yan because Zhao teaches all of the conditions of the density gradient and further teaches centrifugation in PBS at 100g for 2 minutes and Imam makes clear that using about 200g of centrifugation, most cells do not require more than 5 minutes of centrifugation.
Salema, Jung, Imam, Zhao, and Yan teach as set forth above, but fail to teach the inert medium is a medium capable of forming a density gradient after centrifugation, such as a polysaccharide-based medium, as in claims 1 and 12. However, Richman teaches a selection strategy for engineering high-affinity proteins for a cell surface ligand by selecting with density centrifugation (see, e.g., p. 255, under abstract). Richman teaches the inert medium for centrifugation is Ficoll-Paque, as in claim 4 (see, e.g., p. 257, under “Density differential centrifugation”). It is understood that Ficoll-Paque is a polysaccharide-based medium because of the example on p. 12, para. 2 of the applicant’s specification.
Salema, Jung, Imam, Zhao, Yan, and Richman are analogous to the field of the claimed invention because they are all in the field of microbiology. One of ordinary skill in the art before the effective filing date of the application would have found it obvious to incorporate the inert medium, Ficoll-Paque, of Richman into the separation methods of Salema, Jung, Imam, Zhao, and Yan. An artisan would have been motivated to do so because Richman discloses that “the visible layer of cells above the Ficoll-Paque (the ‘interface’) was removed from each tube in a 1 mL volume” (see, p. 257, under “Density differential centrifugation”). An artisan would have recognized that the Ficoll-Paque creates a visible layer, rendering removal of the unbound cells facile. An artisan would have had a reasonable expectation of success based on the given disclosures.
With respect to claims 18, Salema teaches as set forth above and further teaches washing the microorganisms with lithium borate (LB) after separating microorganisms bound to antigen-beads from unbound microorganisms and plating the microorganisms bound to the antigen-beads on agar plates in culture medium to obtain isolated colonies, similar to claim 18 (see, e.g., p. 1476, under “Selection of Nbs by E. coli display using magnetic cell sorting (MACS)”, col. 1, para. 3). But, Selema fails to teach the beads are washed with PBS.
However, Zhao teaches washing the beads with PBS after separating nanoantibodies bound to antigen-beads from unbound nanoantibodies, as in claim 18 (see, e.g., p. 6, under “Example 4”, under “Nanobody biological characterization includes the following steps:”, under “(1)Species specific detection of anti-human serum albumin nanobody:”).
One of ordinary skill in the art before the effective filing date of the application would have found it obvious to simply substitute the wash buffer, LB, of Salema with the wash buffer, PBS, of Zhao because these wash buffers are functionally equivalent. A skilled artisan would have had a reasonable expectation of success in using PBS as a wash buffer in the modified method of Salema because both references teach PBS as a common and well-known buffers for use in assay. Salema discloses LB as an example buffer after separating microorganisms bound to antigen-beads from unbound microorganisms and further discloses “[a]ll steps were carried out at room temperature using mild buffers like PBS and LB, which were sufficient to wash out most non-specific binders and isolate antigen-specific clones” (see p. 1475, under “Selection of Nbs by E. coli display using magnetic cell sorting (MACS)”, col. 2, para. 1). The artisan would have a reasonable expectation of success based on the cumulative disclosures discussed above.
With respect to claim 19, Selema teaches as set forth above and further that the colonies correspond to microorganisms expressing HHVs capable of binding to the antigen of interest, as in claim 19 (see, e.g., p. 1476, under “Selection of Nbs by E. coli display using magnetic cell sorting (MACS)”, col. 1, para. 3).
Claims 2 and 3 are rejected under 35 U.S.C. 103 as being unpatentable over Salema, et al. (cited above) in view of Jung, et al. (cited above), a ResearchGate comment from S Ashraf Imam (cited above), Zhao (cited above), Yan (cited above), and Richman (cited above) as evidenced by the material safety data sheet of Agarose beads from Gold Biotechnology (cited above) and the product data sheet of PBS buffer from Biosolve Chemical (cited above), as applied to claims 1, 12, and 18-19 above, and further in view of Zucca, et al. (“Agarose and Its Derivatives as Supports for Enzyme Immobilization”, published 2016, cited in PTO-892 dated 08/14/2025).
Selema, Jung, Imam, Zhao, Yan, and Richman teach as set forth above. Zhao teaches washing the beads with PBS after separating nanobodies bound to antigen-beads from unbound nanobodies (see, e.g., p. 6, under “Example 4”, under “Nanobody biological characterization includes the following steps:”, under “(1)Species specific detection of anti-human serum albumin nanobody:”). The material safety data sheet of Agarose beads from Gold Biotechnology discloses that agarose beads have a density of 1.05-1.03 g/mL, which is more than the stated 1.00-1.01 g/mL density of PBS cited above (see, e.g., col. 2, under “PHYSICAL AND CHEMICAL PROPERTIES”, under “Density”).
With respect to claims 2 and 3, Selema, Jung, Imam, Yan, and Richman fail to teach the use of the agarose beads of Zhao as discussed above.
In a journal article on the advantages of using agarose-based beads with bound-protein, Zucca gives motivation for substituting the beads of Selema with the agarose beads of Zhao. Zucca teaches “Agarose-based beads are highly porous, mechanically resistant, chemically and physically inert, and sharply hydrophilic” (see, e.g., p. 1, under “Abstract”).
Therefore, it would have been obvious to one of ordinary skill in the art before the effectively filing date of the instant application to use the agarose beads taught by Zhao in the modified method of Selema because Zucca teaches that agarose beads are chemically and physically inert and sharply hydrophilic, which are useful qualities for functionalizing proteins. The artisan would have a reasonable expectation of success based on the cumulative disclosures discussed above.
Claims 13 and 17 are rejected under 35 U.S.C. 103 as being unpatentable over Salema, et al. (cited above) in view of Jung, et al. (cited above), a ResearchGate comment from S Ashraf Imam (cited above), Zhao (cited above), Yan (cited above), and Richman (cited above) as evidenced by the material safety data sheet of Agarose beads from Gold Biotechnology (cited above) and the product data sheet of PBS buffer from Biosolve Chemical (cited above), as applied to claims 1, 12, and 18-19 above, and further in view of Fischinger, et al. (A high-throughput, bead-based, antigen-specific assay to assess the ability of antibodies to induce complement activation”, published 2019, cited in PTO-892 dated 08/14/2025).
Selema, Jung, Imam, Zhao, Yan, and Richman teach as set forth above. Jung teaches the beads stably bind to the antigen of interest in a time of between 2 to 12 hours (see, e.g., p. 42, under “Digoxigenin Immobilization Onto Beads”). It is understood that incubation overnight is between 2 to 12 hours.
With respect to claim 13, Selema, Jung, Imam, Zhao, Yan, and Richman fail to teach incubating the beads and the antigen of interest for a time between 2-12 hours.
In a journal article on a bead-based, antigen-specific assay that includes binding of antibodies to antigen-beads, Fischinger teaches the optimization of incubation time of the antigen with the beads (see, e.g., p. 6, under “Fig. 3.”, panel “D” and panel “E”).
With respect to claim 17, Selema, Jung, Imam, Zhao, Yan, and Richman fail to teach the antigen-beads are incubated with the microorganisms from the expression library for 20 to 60 minutes at room temperature.
However, Jung teaches the antigen-beads are incubated with the microorganisms from the expression library for 2 hours at room temperature, similar to claim 17 (see, e.g., p. 42, under “Preparation of Spheroplasts and FC Analysis”, col. 2).
Fischinger also teaches the optimization of incubation time of antibodies with the beads from 15 minutes to 2 hours (see, e.g., p. 6, under “Fig. 3.”, panel “J” and panel “K”).
Therefore, it would have been obvious to one of ordinary skill in the art before the effectively filing date of the instant application to modify the method of Selema as modified by Imam, Zhao, Yan, and Richman and to further incubate the beads with the antigen for a time between 2 and 12 hours as taught by Jung and incubating the microorganisms expressing the nanoantibody library for 20 to 60 minutes because Fischinger teaches that the incubation times for these steps can be optimized. Selecting the incubation steps of between 2 and 12 hours for the antigen with beads and 20 to 60 minutes for the antibodies with the beads is routine optimization for optimizing the signal to noise ratio, as taught by Fischinger (see, e.g., p. 4, under “3.3 Antigen-bead coupling”, col 2, para. 1). It would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to perform routine optimization of the components in the claimed invention to make and use the claimed invention. As noted in In re Aller, 105 USPQ 233 at 235, more particularly, where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Routine optimization is not considered inventive and no evidence has been presented that arriving at the claimed incubation protocols were anything other than routine, that the properties of the incubation protocols from the optimization have any unexpected properties, or that the results should be considered unexpected in any way as compared to the closest prior art. Optimization of parameters is a routine practice that would be obvious for the artisan to employ. See MPEP § 2144.05. The artisan would have had a reasonable expectation of success based on the cumulative disclosures discussed above.
Claim 14 is rejected under 35 U.S.C. 103 as being unpatentable over Salema, et al. (cited above) in view of Jung, et al. (cited above), a ResearchGate comment from S Ashraf Imam (cited above), Zhao (cited above), Yan (cited above), and Richman (cited above) as evidenced by the material safety data sheet of Agarose beads from Gold Biotechnology (cited above) and the product data sheet of PBS buffer from Biosolve Chemical (cited above), as applied to claims 1, 12, and 18-19 above, and further in view of Bronge, et al. (“Sensitive detection of antigen-specific T-cells using bead-bound antigen for in vitro re-stimulation”, published 2019, cited in PTO-892 dated 08/14/2025).
Salema, Jung, Imam, Zhao, Yan, and Richman teach as set forth above, but fail to teach blocking binding sites on the beads which have not reacted with the antigen, as in claim 14.
However, Bronge teaches blocking binding sites on the beads which have not reacted with the antigen, as in claim 14 (see, e.g., p. 1638, under “Antigen bead coupling”, para. 1).
Salema, Jung, Imam, Zhao, Yan, Richman, and Bronge are analogous to the field of the claimed invention because they are all in the field of microbiology. One of ordinary skill in the art before the effective filing date of the application would have found it obvious to add the blocking protocol of Bronge to methods of Salema, Jung, Imam, Zhao, Yan, and Richman. Blocking after functionalization steps is common in the art and Bronge teaches that the protocol provides the advantage of blocking “still reactive carboxylic groups” to prevent non-specific surface attachment and improve assay sensitivity and specificity. An artisan would have understood that still reactive functionalization groups would lead to non-specific binding to the beads without a blocking step, and would have a reasonable expectation of success based on the cumulative disclosures.
Claims 15 and 16 are rejected under 35 U.S.C. 103 as being unpatentable over Salema, et al. (cited above) in view of Jung, et al. (cited above), a ResearchGate comment from S Ashraf Imam (cited above), Zhao (cited above), Yan (cited above), and Richman (cited above) as evidenced by the material safety data sheet of Agarose beads from Gold Biotechnology (cited above) and the product data sheet of PBS buffer from Biosolve Chemical (cited above), as applied to claims 1, 12, and 18-19 above, and further in view of Turner, et al. (“Optimized expression of soluble cyclomaltodextrinase of thermophilic origin in Escherichia coli by using a soluble fusion-tag and by tuning of inducer concentration”, published 2005, cited in PTO-892 dated 08/14/2025).
Selema, Jung, Imam, Zhao, Yan, and Richman teach as set forth above. Jung further teaches that the microorganisms of the expression library are treated with the protein expression inducer, IPTG, for 5 hours at a concentration of 1 mM, similar to claims 15 and 16 (see, e.g., p. 42, under “Culture Conditions” to under “Preparation of Spheroplasts and FC Analysis”, para. 1).
With respect to claims 15 and 16, Selema, Jung, Imam, Zhao, Yan, and Richman fail to teach the microorganisms from the expression library are treated with a protein expression inducer for 2 to 4 hours and the protein expression inducer is isopropyl-β-D-1-tiogalactopyranoside (IPTG) at a concentration between 20 µM to 100 µM.
However, Turner teaches the importance of optimizing the expression conditions, such as IPTG concentrations (see, e.g., p. 54, under “Abstract”). Turner teaches optimizing the induction time with time points of 1, 2, 3, and 4 h induction with 50 µM IPTG, as in claims 15 and 16 (see, e.g., p. 57, under “Fig. 3.”).
Therefore, it would have been obvious to one of ordinary skill in the art before the effectively filing date of the instant application to modify the method of Selema as modified by Imam, Zhao, Yan, and Richman to utilize IPTG for 2 to 4 hours at a concentration between 20 µM to 100 µM as taught by Jung and Turner. While Jung and Turner do not explicitly identify IPTG induction for 2 to 4 hours at a concentration of 20 to 100 µM as the optimal induction conditions, Turner does teach the importance of optimizing the conditions of induction. It would have been prima facie obvious for one of ordinary skill in the art before the effective filing date of the claimed invention to perform routine optimization of the components in the claimed invention to make and use the claimed invention. As noted in In re Aller, 105 USPQ 233 at 235, more particularly, where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. Routine optimization is not considered inventive and no evidence has been presented that arriving at the claimed induction protocol were anything other than routine, that the properties of the induction protocol from the optimization have any unexpected properties, or that the results should be considered unexpected in any way as compared to the closest prior art. Optimization of parameters is a routine practice that would be obvious for the artisan to employ. See MPEP § 2144.05. The artisan would have had a reasonable expectation of success based on the cumulative disclosures discussed above.
Response to Arguments
The applicant’s arguments filed 02/17/2026 have been fully considered. However, they have not been found persuasive.
35 U.S.C. 103 Rejections
The applicant begins the discussion of the 35 U.S.C. 103 rejections by discussing the beneficial features of the method of claim 1.
Beneficial feature 1 is the selective removal of non-specific binders. The centrifugation of claim 1 separates the antigen-coated bead bound microorganisms expressing the antigen-specific VHH from the unbound microorganisms. However, Jung (cited above) teaches, “Three hundred microliters of the spheroplasts and 50 mL of the 1% (wt/vol) digoxigenin immobilized beads were mixed in a glass test tube and incubated at room temperature for 2 h with shaking and washed in 1 mL of PBS to remove unbound spheroplasts by centrifugation at 3,000 rpm for 3 min” (see, p. 42, col. 2, para. 1, emphasis added). Jung teaches separating bound microorganisms from unbound microorganisms by centrifugation. Therefore, the beneficial feature 1 is known in the art, as demonstrated by Jung.
The second beneficial feature is the phenomenon that allows for the first beneficial feature. The spatial separation of bound and unbound microorganisms by centrifugation allows for the selective removal of non-specific binders. Therefore, Jung also teaches the second beneficial feature.
The third beneficial feature is the elimination of expensive instrumentations. The separation process using centrifugation can remove the need for expensive instrumentation, such as a cell sorter. Jung inherently teaches the third beneficial feature by teaching the use of centrifugation for separating bound and unbound microorganisms. A person of ordinary skill in the art would have understood that Jung’s method provides the benefit of eliminating the need for expensive instrumentations.
The fourth beneficial feature is the speed of the separation process, which is faster than cell sorter-based isolation. Again, Jung inherently teaches the fourth beneficial feature by teaching the use of centrifugation for separating bound and unbound microorganisms. A person of ordinary skill in the art would have understood that Jung’s method provides the benefit of increasing the speed of the separation process.
The supposed fifth beneficial feature is the successful application of the claimed invention to research in the art. However, further research in the art, does not show novelty or non-obviousness. The examples provided are authored (or at least co-authored) by the inventors. The inventors’ use of their own claimed invention does not impact the patentability of the claims.
The applicant continues by arguing that a person of ordinary skill in the art would not have expected the beneficial features of the method of claim 1. However, Jung teaches the beneficial features of claim 1 by teaching the separation of bound and unbound microorganism by centrifugation. A person of ordinary skill in the art would have been well aware of the cost and speed of using a cell-sorter based isolation method, and they would have understood that the centrifugation method of Jung would not require the resources associated with the cell-sorter based isolation method.
The applicant continues by arguing that Salema (cited above) does not teach the density gradient after centrifugation separation of the claimed invention. However, Jung, not Salema, is relied upon for the disclosure of centrifugation for isolation of microorganisms bound to beads. The applicant argues that Jung discloses the use of flow cytometry for isolation of spheroplasts expressing a single-chain antibody fragment. However, Jung teaches, “Three hundred microliters of the spheroplasts and 50 mL of the 1% (wt/vol) digoxigenin immobilized beads were mixed in a glass test tube and incubated at room temperature for 2 h with shaking and washed in 1 mL of PBS to remove unbound spheroplasts by centrifugation at 3,000 rpm for 3 min. FC analysis was performed using a BD FACSort” (see, p. 42, col. 2, para. 1, emphasis added). Jung clearly teaches isolating bound spheroplasts by removing unbound spheroplasts via centrifugation and then analyzing the bound spheroplasts using flow cytometry.
The applicant continues on p. 10 of the applicant’s remarks filed 02/17/2026 by arguing that PBS is not recited in the claims nor disclosed in the present application as a medium for forming a density gradient after centrifugation. However, the 35 U.S.C. 103 rejection was written in response to the claims filed 11/14/2022. PBS was encompassed by the broadest reasonable interpretation of “an inert medium with a density greater than or equal to 1 g/mL”, which is the language of the claims filed 11/14/2022. The amended claims are rejected in the new 35 U.S.C. 103 rejections above.
The applicant continues by arguing that Zhao (cited above) and Imam (cited above) do not teach or suggest the separation of microorganisms bound to beads by centrifugation. However, Zhao is relied upon for teaching washing with PBS and Imam is relied upon for teaching centrifugation conditions, as discussed above. The applicant continues that a person of ordinary skill in the art would not have expected the upper fraction, which contains microorganisms not bound to beads, and the bead sediment, which contains microorganisms bound to beads, can be separated. However, as discussed above, Jung clearly teaches that centrifugation can be used to separate microorganisms bound to beads from microorganisms not bound to beads.
The same argument is used for Yan (cited above). However, Jung teaches the separation by centrifugation and Yan is relied upon for teaching that magnetic activated cell sorting and density gradient centrifugation are functional equivalents.
The rest of the applicant’s arguments are a discussion of how none of the cited references teach or suggest the separation of microorganisms through an inert media that is capable of forming a density gradient centrifugation. However, as discussed above, Jung teaches the separation by centrifugation and the other references are teaching additional limitations of the claimed invention.
Conclusion
No claims are allowed.
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/MICHAEL CAMERON SVEIVEN/ Examiner, Art Unit 1678
/GREGORY S EMCH/ Supervisory Patent Examiner, Art Unit 1678
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