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 .
Election/Restrictions
Applicant’s election without traverse of 1, 11-14, 40, 48, 53-55, 57-59 in the reply filed on 07/10/2026 is acknowledged.
Claims 15, 21, and 25-29 withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to nonelected inventions, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 07/10/2026.
Claim Interpretation
In claim 1, the preamble recites a method that is “suitable for” protein footprinting. The phrase “suitable for” is subjective, not a term of art, nor specially defined in the specification; in the interest of compact prosecution, examiner interprets “suitable for” to mean “capable of being used for.”
Claim Rejections - 35 USC § 112
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.
Claim 11, 53, 55, and 56-59 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 11 recites the limitation "the presence of the protein of interest" in line 2. There is insufficient antecedent basis for this limitation in the claim.
Claim 11 recites both “the reaction solution” and “the presence of the protein of interest.” As claimed, claim 1 already defines the reaction solution as containing the protein of interest, so there is a lack of clarity as to the intended scope of “the presence of the protein of interest.” In the interest of compact prosecution, Examiner interprets this to mean “anything containing the protein of interest.”
Claim 11 introduces an additional step of adding a quenching agent, but it is unclear when this is meant to occur relative to the steps introduced in claim 1. In the interest of compact prosecution, examiner interprets the step of claim 11 to occur during and/or after step b) of claim 1.
A broad range or limitation together with a narrow range or limitation that falls within the broad range or limitation (in the same claim) may be considered indefinite if the resulting claim does not clearly set forth the metes and bounds of the patent protection desired. See MPEP § 2173.05(c). In the present instance, claim 53 recites:
the broad recitation “a concentration of at least 1 µM”, and the claim also recites “at least 50 M” and “at least 5 mM” which are the narrower statements of the range/limitation. The claim(s) are considered indefinite because there is a question or doubt as to whether the feature introduced by such narrower language is (a) merely exemplary of the remainder of the claim, and therefore not required, or (b) a required feature of the claims. In the interest of compact prosecution, Examiner will consider concentration to be “a concentration of at least 1 µM.”
the broad recitation “a length of time of less than 1 ms”, and the claim also recites “less than 500 µs” and “less than 1 µs” which are the narrower statements of the range/limitation. The claim(s) are considered indefinite because there is a question or doubt as to whether the feature introduced by such narrower language is (a) merely exemplary of the remainder of the claim, and therefore not required, or (b) a required feature of the claims. In the interest of compact prosecution, Examiner will consider a length of time to be “a length of time of less than 1 ms.”
The term “excess” in claim 57 is a relative term which renders the claim indefinite. The term “excess” is not defined by the claim, the specification does not provide a standard for ascertaining the requisite degree, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. It is impossible to know exactly how much of the “hydroxyl radical precursors” would need to be present to constitute “excess quantity”. In the interest of compact prosecution, examiner interprets “excess quantity” to mean “more than anything else in the reaction solution.”
Claim Rejections - 35 USC § 102
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.
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.
Claim(s) 1, 11, 13, 40, 55, 57-59 is/are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Cheng et al. (Angew Chem Int Ed Engl, Vol. 56, No. 45, pgs. 14007–14010, 06 November 2017, as cited in the IDS submitted on 08/08/2023), as evidenced by the provided Supplemental Information.
Regarding claim 1, Cheng et al. teaches a one-pot method of multiplex protein labeling that is suitable for protein footprinting (abstract) (We describe a novel, laser-initiated radical trifluoromethylation for protein footprinting and establish its broad residue coverage), the method comprising:
generating hydroxyl radicals in a reaction solution (pg. 3 second paragraph) (The key to pulsed formation of •CF3 is the hydroxyl radical, formed by photolysis of HO-OH (pathway a)), the reaction solution comprising hydroxyl radical precursors (pg. 3 first paragraph) (laser irradiation in the presence of H2O2), trifluoromethyl radical precursors (pg. 3 first paragraph) (laser irradiation in the presence of… the water-soluble salt, NaSO2CF3), and a protein of interest (pg. 3 first paragraph) (neuropeptide Y 18-36 (NPY18-36)… exposing NPY18-36 to…), the hydroxyl radicals generating trifluoromethyl radicals from the trifluoromethyl radical precursors (pg. 3 second paragraph) (The key to pulsed formation of •CF3 is the hydroxyl radical… •OH oxidizes CF3SO2− to CF3SO2•, which rapidly collapses to generate CF3•); Examiner is interpreting the laser as a hydroxyl/trifluoromethyl precursor because it is required for the generation of hydroxyl/trifluoromethyl radicals (pg. 2 first paragraph) (found no detectable CF3-products in the absence of… laser irradiation). (See also pg. 2 of supplemental information of Cheng, section 2.1.1., which shows that water is used for the generation of hydroxyl radicals) (protein solution was then added H2O2 (5 μL, 100 mM in water) and CF3SO2Na (5 μL, 100 mM in water).”
b) maintaining a hydroxyl radical concentration for a first length of time sufficient to both:
i) react a first portion of the hydroxyl radicals with the trifluoromethyl radical precursors in the reaction solution to maintain a trifluoromethyl radical concentration for a second length of time sufficient to react with the protein of interest (pg. 3 second paragraph) (•OH oxidizes CF3SO2− to CF3SO2•, which rapidly collapses to generate CF3•… •CF3 then reacts with the protein (pathway b) by addition of CF3); and
ii) react a second portion of the hydroxyl radicals with the protein of interest, thereby labeling the protein of interest with both hydroxyl substituents and trifluoromethyl substituents (pg. 2 second paragraph) (In a minor pathway… ·OH abstracts H• from the protein to produce a protein radical that… reacts with HO•… to generate oxygen-oxidization).
Regarding claim 11, Cheng teaches the method of claim 1 as rejected above, (pg. 2 first paragraph and pg. 3 first paragraph of supplemental information) (Given that the concentration of CF3SO2− is ~ 300 times that of •OH, the •OH is readily quenched by the oxidation and All samples were collected in vials containing 500 nM catalase and 7 mM methionine, protecting protein from further oxidation by H2O2).
Regarding claim 13, Cheng teaches the method of claim 1 as rejected above, wherein generating hydroxyl radicals comprises irradiating the hydroxyl radical precursor or the hydroxyl radical precursors with photolytic radiation (pg. 3 first paragraph) (laser irradiation in the presence of H2O2… photolysis of HO-OH).
Regarding claim 40, Cheng teaches the method of claim 1 as rejected above, wherein labeling the protein of interest comprises labeling at least 13 distinct amino acids of the 20 most abundant amino acids found in proteins (abstract and pg. 2 first paragraph) (•CF3 reacts with 18 of 20 common amino acids and •OH reacts with all amino acids).
Regarding claim 55, Cheng teaches the method of claim 1 as rejected above wherein the hydroxyl radical precursor comprises water or hydrogen peroxide or the hydroxyl radical precursors comprise water or hydrogen peroxide (pg. 2 of supplemental information of Cheng, section 2.1.1.) (protein solution was then added H2O2 (5 μL, 100 mM in water)).
Regarding claim 57, Cheng teaches the method of claim 1 as rejected above, wherein the hydroxyl radical precursor or the hydroxyl radical precursors are present in excess quantity (pg. 2 of supplemental information of Cheng, section 2.1.1.) (protein solution was then added H2O2 (5 μL, 100 mM in water) and CF3SO2Na (5 μL, 100 mM in water) sequentially to make a solution of 10 μM NPY 18-36, 10 mM H2O2, 10 mM CF3SO2Na in PBS buffer). Water is considered to be a hydroxyl radical precursor and a trifluoromethyl radical precursor, and the reaction solution is an aqueous media; there is excess water compared to other components of the solution. Therefore, Chen teaches the hydroxyl radical precursor or the hydroxyl radical precursors are present in excess quantity.
Regarding claim 58, Cheng teaches the method of claim 1 as rejected above, wherein the trifluoromethyl radical precursor comprises or the trifluoromethyl radical precursors comprise a precursor selected from the group consisting of Langlois reagent, umemato-tetrafluoroborate, umemato-trifluoromethanesulfonate, zinc trifluoromethanesulfonate, ethyl trifluoromethanesulfonate, 4,4,4,4',4',4'-hexafluoro-DL-valine, or a combination thereof (pg. 3 first paragraph) (trifluoromethylation of neuropeptide… by exposing NPY18-36 to laser irradiation in the presence of H2O2 and the water-soluble salt, NaSO2CF3). Note that NaSO2CF3 is Langlois reagent.
Regarding claim 59, Cheng teaches the method of claim 1 as rejected above, wherein the trifluoromethyl radical precursor comprises Langlois reagent or the trifluoromethyl radical precursors comprise Langlois reagent (pg. 3 first paragraph) (trifluoromethylation of neuropeptide… by exposing NPY18-36 to laser irradiation in the presence of H2O2 and the water-soluble salt, NaSO2CF3). Note that NaSO2CF3 is Langlois reagent.
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.
The factual inquiries for establishing a background for determining obviousness under 35 U.S.C. 103 are summarized as follows:
1. Determining the scope and contents of the prior art.
2. Ascertaining the differences between the prior art and the claims at issue.
3. Resolving the level of ordinary skill in the pertinent art.
4. Considering objective evidence present in the application indicating obviousness or nonobviousness.
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.
Claim(s) 12 is/are rejected under 35 U.S.C. 103 as being unpatentable over Cheng et al. (Angew Chem Int Ed Engl, Vol. 56, No. 45, pgs. 14007–14010, 06 November 2017, as cited in the IDS submitted on 08/08/2023), as evidenced by the provided Supplemental Information, as applied to claim 1 above, further in view of Kiselar et al. (Annual Review of Biophysics, Vol. 47, pgs. 315-333, 14 March 2018, as cited in the IDS submitted on 08/08/2023).
Regarding claim 12, Cheng teaches the method of claim 1 as rejected above. Cheng teaches generating hydroxyl radicals via photolysis (pg. 3 first paragraph of Cheng) (laser irradiation in the presence of H2O2… photolysis of HO-OH).
Cheng does not teach wherein generating hydroxyl radicals comprises irradiating the hydroxyl radical precursor or the hydroxyl radical precursors with ionizing radiation.
In the analogous art of hydroxyl radical protein labelling, Kiselar teaches of irradiating a hydroxyl radical precursor with ionizing radiation (pg. 319 first paragraph of Kiselar) (Hydroxyl radicals can also be generated in the protein’s interior when ordered waters are activated by radiolysis, leading to stable modifications reflective of internal protein structure). Kiselar teaches that radiolysis is more effective than photolysis because it enables the labelling (‘oxidation’) of the protein interior (pg. 319 second paragraph of Kiselar) (The internal oxidations seen with radiolysis are not observed in photolysis, as the hydrogen peroxide does not easily enter the protein interior).
It would have been obvious to a person having ordinary skill in the art before the effective filing date of the instant application to substitute the photolytic generation of hydroxyl radicals as taught by Cheng with the method of radiolytically generating hydroxyl radicals taught by Kiselar because it would lead to labelling the protein interior to more thoroughly label the protein with a reasonable expectation of success (see pg. 3 first paragraph of Cheng; pg. 319 first and second paragraph of Kiselar). See MPEP 2143(I)(B).
Claim(s) 14 is/are rejected under 35 U.S.C. 103 as being unpatentable over Cheng et al. (Angew Chem Int Ed Engl, Vol. 56, No. 45, pgs. 14007–14010, 06 November 2017, as cited in the IDS submitted on 08/08/2023), as evidenced by the provided Supplemental Information, as applied to claim 1 above, further in view of Monroe et al. (Analytical Chemistry, Vol. 85, pgs. 6185-6189, 18 June 2013).
Regarding claim 14, Cheng teaches the method of claim 1 as rejected above. Cheng teaches generating hydroxyl radicals via photolysis (pg. 3 first paragraph of Cheng) (laser irradiation in the presence of H2O2… photolysis of HO-OH).
Cheng does not teach wherein generating hydroxyl radicals comprises electrochemically generating hydroxyl radicals.
In the analogous art of labelling proteins of interest with hydroxyl radicals, Monroe teaches electrochemically generating hydroxyl radicals (abstract of Monroe) (we develop the electro-Fenton reaction as a means to generate hydroxyl radicals for structural footprinting mass spectrometry experiment). Monroe teaches that electrochemically generating hydroxyl radicals is cheaper than using photolysis and improves the control of the timing and extent of the radical generation process (abstract of Monroe) (laser photolysis of H2O2… adds significant costs and/or complexity to the experiments… the electro-Fenton reaction… while reducing the costs and complexity of initiating such experiments… enables control of the timing and extent of the radical generation process, while reducing the complexity).
It would have been obvious to a person having ordinary skill in the art to substitute the photolytic generation of hydroxyl radicals as taught by Cheng with the method of electrochemically generating hydroxyl radicals taught by Monroe because it would lead to reducing costs of the labelling experiment while improving the control of the timing and extent of the radical generation process of the labelling method with a reasonable expectation of success (see pg. 3 first paragraph of Cheng; abstract of Monroe). See MPEP 2143(I)(B).
Claim(s) 48 is/are rejected under 35 U.S.C. 103 as being unpatentable over Cheng et al. (Angew Chem Int Ed Engl, Vol. 56, No. 45, pgs. 14007–14010, 06 November 2017, as cited in the IDS submitted on 08/08/2023), as evidenced by the provided Supplemental Information, as applied to claim 1 above, further in view of Barth et al. (Journal of Proteomics, Vol. 222, No. 103793, pg. 1-11, 27 April 2020).
Regarding claim 48, Cheng teaches the method of claim 1 as rejected above. Cheng teaches evaluating the extent of protein labelling by observing the percentage of labelled residues within different regions of a protein of interest, labelling up to 89% of the evaluated protein regions (pg. 4 second paragraph of Cheng) (CF3-modification… dramatically different modification extents for regions 80–96 (hMb 6.6% vs aMb 89.0%)).
Cheng does not teach wherein labeling the protein of interest comprises labeling at least 50% of total surface accessible residues within the protein of interest.
In the analogous art of labelling proteins, Barth teaches of examining the extent of surface labelling (pg. 2 left column sixth paragraph) (Here we examine protein surface labelling… following an MS-based workflow). Barth teaches labeling at least 50% of total surface accessible residues within the protein of interest; see table S2, which shows at least 50% of total surface accessible residues within the protein of interest are labelled:
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Barth teaches that quantifying the percentage of labelled surface-accessible residues provides a complete picture of the extent of labelling while accounting for structural changes due to “over-labelling” (pg. 2 left column sixth paragraph) (we assess labelling-induced structural changes that might occur due to “over-labelling”), as well as characterizing the relationship between labelling reagent concentration and the percentage of labelled surface accessible residues (pg. 10 left column second paragraph of Barth) (Following a quantitative workflow, solvent accessible amino acids of two model proteins could be identified by an increase in labelling percentage for increasing amounts of labelling reagents).
It would have been obvious to a person having ordinary skill in the art before the effective filing date of the instant application to modify the regional protein analysis of Cheng to instead evaluate the percentage of surface-accessible residues that are labelled as taught by Barth because it would lead to characterizing the extent of labelling of all surface-accessible residues, accounting for structural changes due to “over-labelling,” and characterizing the relationship between labelling reagent concentration and the percentage of labelled surface accessible residues with a reasonable expectation of success (pg. 4 second paragraph of Cheng; see pg. 2 left column sixth paragraph, pg. 10 left column second paragraph, Table S2, of Barth).
Claim(s) 53-54 is/are rejected under 35 U.S.C. 103 as being unpatentable over Cheng et al. (Angew Chem Int Ed Engl, Vol. 56, No. 45, pgs. 14007–14010, 06 November 2017, as cited in the IDS submitted on 08/08/2023), as evidenced by the provided Supplemental Information, as applied to claim 1 above, further in view of Niu et al. (J. Am. Soc. Mass. Spectrom., Vol. 26, pgs. 843-846, 25 February 2015).
Regarding claim 53, Cheng teaches the method of claim 1 as rejected above. Cheng teaches that the generation of hydroxyl radicals occurs in, presumably, the time required to pulse the sample with 15 mJ of energy from a 248 nm laser light source (pg. 3 of supplemental information of Cheng) (The sample solution was irradiated by a 248 nm KrF excimer laser (15mJ/pulse… Presumably every portion of sample solution are only subjected to one laser shot).
Cheng is silent to wherein labeling the protein of interest comprises a time-generation profile of hydroxyl radicals including a concentration of at least 1 µM, at least 50 µM, or at least 5 mM within a length of time of less than 1 ms, less than 500 µs or less than 1 µs.
In the analogous art of labelling proteins of interest via radical chemistry, Niu teaches a time-generation profile of hydroxyl radicals including a concentration of at least 50 µM (actual concentration ~1 mM) within a length of time of less than 500 µs; see figure 2 of Niu:
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Niu teaches that characterizing the in situ concentration of OH radicals and the time required to generate them allows for the optimization of the reaction conditions and the adjustment of the extent of labelling that occurs on the protein of interest (pg. 846 left column ‘conclusion’ paragraph) (The approach permits optimization of FPOP conditions and adjustment of experimental parameters to realize tunable extents of modifications).
It would have been obvious to a person having ordinary skill in the art before the effective filing date of the instant application to combine the hydroxyl radical generation method of Cheng with the time-generation profile as taught by Niu because it would lead to optimization of the reaction conditions and the adjustment of the extent of labelling that occurs on the protein of interest with a reasonable expectation of success (see pg. 3 of supplemental information of Cheng; figure 2, pg. 846 left column ‘conclusion’ paragraph of Niu).
Regarding claim 54, Cheng teaches the method of claim 1 as rejected above. Cheng teaches that the generation of hydroxyl radicals occurs in, presumably, the time required to pulse the sample with 15 mJ of energy from a 248 nm laser light source (pg. 3 of supplemental information of Cheng) (The sample solution was irradiated by a 248 nm KrF excimer laser (15mJ/pulse… Presumably every portion of sample solution are only subjected to one laser shot).
Cheng is silent to wherein labeling the protein of interest comprises a time-generation profile of hydroxyl radicals including a concentration of between 1 µM and 5 mM within a time range of between 10 µs to 50 ms.
In the analogous art of labelling proteins of interest via radical chemistry, Niu teaches a time-generation profile of hydroxyl radicals including a concentration of between 1 µM and 5 mM (actual concentration ~1 mM); see figure 2 of Niu:
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Niu teaches that characterizing the in situ concentration of OH radicals and the time required to generate them allows for the optimization of the reaction conditions and the adjustment of the extent of labelling that occurs on the protein of interest (pg. 846 left column ‘conclusion’ paragraph) (The approach permits optimization of FPOP conditions and adjustment of experimental parameters to realize tunable extents of modifications).
It would have been obvious to a person having ordinary skill in the art to combine the hydroxyl radical generation method of Cheng with the time-generation profile as taught by Niu because it would lead to optimization of the reaction conditions and the adjustment of the extent of labelling that occurs on the protein of interest with a reasonable expectation of success (see ; figure 2, pg. 846 left column ‘conclusion’ paragraph of Niu).
Modified Cheng (Cheng in view of Niu) is silent to wherein labeling the protein of interest comprises a time-generation profile of hydroxyl radicals including a time range of between 10 µs to 50 ms.
Niu teaches that the timing of the hydroxy radical concentration observed within the time-generation profile may be extended depending on the nature and concentration of what is present in the reaction solution (figure 2 caption of Niu) (Lifetime of •OH (~100 ns to more than 10 μs) depends on the nature and concentration of the scavenger). Niu teaches that the time range for observing the hydroxyl radicals may be increased to 100 µs by adding alanine to the reaction solution (pg. 846 left column second paragraph) (we can adjust the [•OH] available for footprinting by varying either the nature or concentration of the scavenger… lifetime is approximately… 100 μs with alanine as scavengers). Niu teaches that extending the timing of hydroxyl radical generation is advantageous because it enables labeling over a wider time scale and investigating various folding/unfolding events (pg. 846 left column second paragraph) (Adjusting the lifetime of •OH enables labeling over a wider time scale, making FPOP a flexible tool to investigate various folding/unfolding events).
It would have been obvious to a person having ordinary skill in the art to modify the time-generation profile of Modified Cheng (Cheng in view of Niu) to extend the lifetime of hydroxyl radicals to 100 µs as taught by Niu because it would enable labeling over a wider time scale and investigating various folding/unfolding events with a reasonable expectation of success (see pg. 3 of supplemental information of Cheng; see figure 2 and figure 2 caption, pg. 846 left column ‘conclusion’ paragraph, pg. 846 left column second paragraph of Niu).
Conclusion
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/H.D.C./Examiner, Art Unit 1758
/MARIS R KESSEL/Supervisory Patent Examiner, Art Unit 1758
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