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 .
Information Disclosure Statement
The information disclosure statement(s) (IDS) filed 26 March 2025 is considered, initialed, and attached hereto.
Claim Status
Claims 1-20 are pending and under examination.
Drawings
The drawings are objected to because the views in pages 1 and 2 are numbered “Fig. 1A-1B” and “Fig. 1A-1B continued”. The numbering of views must comply with 37 C.F.R. 1.84(u):
(u) Numbering of views.
(1) The different views must be numbered in consecutive Arabic numerals, starting with 1, independent of the numbering of the sheets and, if possible, in the order in which they appear on the drawing sheet(s). Partial views intended to form one complete view, on one or several sheets, must be identified by the same number followed by a capital letter. View numbers must be preceded by the abbreviation "FIG." Where only a single view is used in an application to illustrate the claimed invention, it must not be numbered and the abbreviation "FIG." must not appear.
(2) Numbers and letters identifying the views must be simple and clear and must not be used in association with brackets, circles, or inverted commas. The view numbers must be larger than the numbers used for reference characters.
Corrected drawing sheets in compliance with 37 CFR 1.121(d) are required in reply to the Office action to avoid abandonment of the application. Any amended replacement drawing sheet should include all of the figures appearing on the immediate prior version of the sheet, even if only one figure is being amended. The figure or figure number of an amended drawing should not be labeled as “amended.” If a drawing figure is to be canceled, the appropriate figure must be removed from the replacement sheet, and where necessary, the remaining figures must be renumbered and appropriate changes made to the brief description of the several views of the drawings for consistency. Additional replacement sheets may be necessary to show the renumbering of the remaining figures. Each drawing sheet submitted after the filing date of an application must be labeled in the top margin as either “Replacement Sheet” or “New Sheet” pursuant to 37 CFR 1.121(d). If the changes are not accepted by the examiner, the applicant will be notified and informed of any required corrective action in the next Office action. The objection to the drawings will not be held in abeyance.
Specification
The disclosure is objected to because of the following informalities: the word “amplification” is misspelled as “amplifaction” on multiple lines of page 12 of the specification.
Appropriate correction is required.
Claim Objections
Claim 20 is objected to because of the following informalities: “the intercalating-dye” in line 1 should read as “the fluorescent intercalating-dye”. Appropriate correction is required.
Claim Interpretation
Claim 2 recites the phrase “equivalent sample”. The instant specification provides a limiting definition of equivalent sample that is used for the purpose of claim interpretation: “As used herein, “equivalent” samples are samples from the same source or having the same origin and utilized at the same concentration or quantity in the LAMP reaction” (page 5 line 28+).
Claims 3, 5-6, 14, and 18 recite limitations preceded by the word “optionally”, denoting that the limitation following it is optional. MPEP §2111.04 states: “Claim scope is not limited by claim language that suggests or makes optional but does not require steps to be performed, or by claim language that does not limit a claim to a particular structure”. As such, the broadest reasonable interpretation of the claim scope of claims 3, 5-6, 14, and 18 does not include the limitations following the word “optionally”.
Claim 5 recites “wherein the plurality of LAMP reactions are in a plurality of cells”. The word “cells” may be understood by one of ordinary skill in the art to refer to either biological cells (such as a neuron, fibroblast, red blood cell, etc.) or to physically separate locations that the reactions may take place. As the contents of biological cells are physically sequestered from each other, they are a narrower interpretation. Therefore, claim 5 is given the broadest reasonable interpretation by interpreting “cells” to refer to physically separate locations.
Claims 9-13 recite limitations regarding the amount of an additive (DMSO, ethanol, glycerol, glycogen, or Gu-HCl, respectively) in a LAMP reaction between about [first value] to [second value]. The word “about” is interpreted as modifying both the first and second values in this claim, not just the first value.
Claim Rejections - 35 USC § 112(b) - Indefiniteness
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 15 and 17-20 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 15 recites the limitation "the LAMP primer set" in line 1. There is insufficient antecedent basis for this limitation in the claim, since claim 15 depends upon claim 1 which recites a plurality of LAMP reactions, each comprising a LAMP primer set, and therefore a plurality of LAMP primer sets (see also “different LAMP primer sets” in line 9 of claim 1).
Claims 17-20 are indefinite because of the recitation “at least about” in claim 17 line 3 and claim 19 lines 4-5, which claim 18 inherits by depending upon claim 17 and claim 20 inherits by depend upon claim 19. The phrase “at least” typically indicates a minimum point; however, the phrase is controverted by the term “about”, which implies that values above and below the indicated amount are permitted. Therefore, the juxtaposition of these two terms makes it unclear what minimum values are encompassed by the claim.
Claim Rejections - 35 USC § 102
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.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claims 1-8 and 13-15 are rejected under 35 U.S.C. 102(a)(1) and (a)(2) as being anticipated by Tanner et al. (U.S. Patent Application Publications Cite No 1 in IDS filed 26 March 2025)(US 2021/0285064, published 16 September 2021, effectively filed 12 March 2020), herein Tanner.
Regarding claim 1, Tanner teaches a method of performing a plurality of LAMP assays in parallel at a single temperature (“a method for determining whether a target nucleic acid is present in a sample” [0016]; “the method involves isothermal amplification of the target nucleic acid in a LAMP reaction […] In one embodiment, the method uses two, three, four, or five sets of target-specific primers in a multiplexed reaction (e.g. multiplexed LAMP)” [0017]), the method comprising:
providing a plurality of LAMP reactions (“multiplexed LAMP” [0017]), wherein each LAMP reaction comprises a LAMP primer set, a sample comprising nucleic acids (“bringing an aliquot of the sample into contact with a master mix as described herein to form a reaction mixture wherein the reaction mix additionally includes sets of primers that are specific for the target nucleic acid” [0016]), and a temperature-resistant strand displacing DNA polymerase (“a master mix comprising: a DNA polymerase suitable for isothermal amplification of DNA” [0003]; “Suitable DNA polymerases are known in the art and include strand displacing DNA polymerases preferably mesophilic DNA polymerases such as Bst polymerase or variants thereof” [0004]), and wherein at least one LAMP reaction comprises at least one LAMP-temperature shifting agent allowing for more efficient amplification at a set temperature compared to the same LAMP reaction in the absence of the at least one LAMP-temperature shifting agent (“the reaction mixture further comprises a molecule comprising C—(NH2)2NH; such as guanidine hydrochloride” [0025]);
performing the plurality of LAMP reactions at the set temperature, wherein at least two of the LAMP reactions comprise different LAMP primer sets (“a heating block or water bath suitable for heating a reaction tube, plate, or paper, or a plurality of the same to a temperature suitable for isothermal amplification” [0015]; “the method involves isothermal amplification of the target nucleic acid in a LAMP reaction […] the method uses two, three, four, or five sets of target-specific primers” [0016]); and
detecting LAMP reaction products produced by the plurality of LAMP reactions (“Determination as to whether the target nucleic acid is present in the sample may then proceed by detecting a change in the spectral properties, color, or fluorescence of the reaction mixture” [0016]).
Regarding claim 2, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein each LAMP reaction comprises an equivalent sample (“bringing an aliquot of the sample into contact with a master mix as described herein to form a reaction mixture wherein the reaction mix additionally includes sets of primers that are specific for the target nucleic acid” [0016], as the same sample is being contacted to the sets of primers, the sample is tautologically equivalent to itself).
Regarding claim 3, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein each LAMP reaction in step (a) further comprises deoxyribonucleotides (“a master mix comprising: […] dNTPs (dATP, dGTP, dCTP, and dTTP)” [0003]) and a divalent metal ion suitable for DNA synthesis (“standard amplification buffer containing Tris-HCl, pH 8.8 at 25° C.; (NH4)2SO4; KCl; MgSO4” [0313]” MgSO4 contains Mg as a divalent metal ion; while this standard amplification buffer is not explicitly called for in the method, the method is taught as having a buffer “In embodiments where the master mix is in solution (e.g. following reconstitution), the master mix is in a weakly buffered solution” [0014] and throughout the reference amplification reactions are performed in the 1x ThermoPol buffer (NEB) or 1x Isothermal Amplification Buffer (NEB), which both contain MgSO4, so one of ordinary skill in the art would instantly understand from the disclosure that these are buffers that may be used in the amplification reaction of the method and that they contain a divalent metal ion), and optionally wherein the plurality of LAMP reactions further comprises a reverse transcriptase (“The master mix may comprise a reverse transcriptase” [0010]).
Regarding claim 4, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein the set temperature in step (b) is controlled by a water bath, heating element, and/or thermal cycler (“a heating block or water bath suitable for heating a reaction tube, plate, or paper, or a plurality of the same to a temperature suitable for isothermal amplification” [0015]).
Regarding claim 5, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein the plurality of LAMP reactions are in a plurality of cells and wherein the cells are optionally within a plate, linked test tubes or a microfluidic chip (“a heating block or water bath suitable for heating a reaction tube, plate, or paper, or a plurality of the same to a temperature suitable for isothermal amplification” [0015]; “In one embodiment, the method comprises analyzing multiple samples. For example, the method may use a reaction container that has multiple compartments each for analyzing a separate sample” [0027]).
Regarding claim 6, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein step (c) comprises detecting light absorption, light emission upon excitation, or light scattering before and after step (b); and optionally detecting a change in color, turbidity, or fluorescence after step (b) compared to before step (b) (“Determination as to whether the target nucleic acid is present in the sample may then proceed by detecting a change in the spectral properties, color, or fluorescence of the reaction mixture” [0016]).
Regarding claim 7, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein the temperature-resistant strand displacing DNA polymerase is a B. stearothermophilus polymerase (“Suitable DNA polymerases are known in the art and include strand displacing DNA polymerases preferably mesophilic DNA polymerases such as Bst polymerase” [0004], Bst polymerase is B. stearothermophilus polymerase).
Regarding claim 8, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein the LAMP-temperature shifting agent is one or more of DMSO, ethanol, glycerol, glycogen, and Gu-HCl (“the reaction mixture further comprises a molecule comprising C—(NH2)2NH; such as guanidine hydrochloride” [0025]).
Regarding claim 13, Tanner teaches the method of claim 8 (see 35 U.S.C. 102 rejection of claim 8 above), wherein Gu-HCl is present in the LAMP reaction between about 5 to 100 millimolar (“The C—(NH2)2NH containing molecule can be added to the reaction at a concentration of up to 60 mM, such as in the range of 20 mM-40 mM (e.g. about 20 mM, 30 mM, or 40 mM)” [0025]; FIG. 18 and 19 using 40mM Guanidine HCl).
Regarding claim 14, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein the sample is from a subject; and optionally wherein the sample from the subject is a nasopharyngeal swab or a saliva swab (“The sample may be a clinical sample, such as a sample of a body fluid (e.g. blood, sputum, saliva, mucous, lymph, sweat, urine, feces, etc.) or a sample taken from a swab such as a nasal, oral, or buccal swab, which may be from a human or other mammalian subject” [0018]).
Regarding claim 15, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein the LAMP primer set is specific for SARS-CoV-2 (“the master mix or reaction mixture comprises at least one set of primers specific for a target SARs-CoV-2 nucleic acid” [0020]).
Therefore, claims 1-8 and 13-15 are anticipated by the teachings of Tanner.
Claims 19-20 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Quyen et al. (Non-Patent Literature Document Cite No 4 in IDS filed 26 March 2025)(“Classification of Multiple DNA Dyes Based on Inhibition Effects on Real-Time Loop-Mediated Isothermal Amplification (LAMP): Prospect for Point of Care Setting” Front Microbiol 10: 2234 (2019)), herein Quyen, and as evidenced by Eischeid (“SYTO dyes and EvaGreen outperform SYBR Green in real-time PCR” BMC Res Notes 4: 263 (2011)) and Monis et al. (“Comparison of SYTO9 and SYBR Green I for real-time polymerase chain reaction and investigation of the effect of dye concentration on amplification and DNA melting curve analysis” Anal Biochem 340(1), pages 24-34 (2005)), herein Monis.
Regarding claim 19, Quyen teaches a method for increasing an end-point signal-to-noise ratio of a fluorescent intercalating-dye based LAMP assay (“Of the nine dyes tested SYTO 9 showed the highest SNR” page 4 right column paragraph 3), the method comprising:
performing one or more LAMP reactions with a fluorescent intercalating-dye (“These real-time LAMP assays were performed at 5 µM dye concentration in the presence of 2 ng of genomic S. Enteritidis. Of the nine dyes tested, SYTO 9” page 4 right column paragraph 3);
heating the one or more LAMP reactions at a temperature of at least about 80 °C for at least about 30 seconds (“The reactions were performed at 65°C for 60 min and the reactions were then terminated by heating to 90°C for 10 min” page 3 left column paragraph 2); and
detecting a LAMP reaction product by detecting the fluorescent intercalating-dye (“The fluorescent signal was recorded every minute of amplification” page 3 left column paragraph 2).
The SYTO 9 dye taught in the method of Quyen is an intercalating dye, as evidenced by Monis (““the intercalating dye SYTO9” Abstract).
Regarding claim 20, Quyen teaches the method of claim 19 (see 35 U.S.C. 102 rejection of claim 19 above), wherein the fluorescent intercalating-dye is a cyanine dye. The SYTO 9 dye taught by Quyen is a cyanine dye, as evidenced by Eischeid (“The SYTO dyes constitute a large family of commercially available cyanine dyes” page 1 right column paragraph 2).
Therefore, claims 19-20 are anticipated by the teachings of Quyen.
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 9 is rejected under 35 U.S.C. 103 as being unpatentable over Tanner et al. (U.S. Patent Application Publications Cite No 1 in IDS filed 26 March 2025)(US 2021/0285064, published 16 September 2021, effectively filed 12 March 2020), herein Tanner, as applied to claims 1-8 and 13-15 above, and in view of Bamford (US 2016/0289752, published 6 October 2016, effectively filed 31 March 2015).
Regarding claim 9, Tanner teaches the method of claim 8 (see 35 U.S.C. 102 rejection of claim 8 above). However, Tanner does not teach the method wherein DMSO is present in the LAMP reaction between about 1 to 5% volume per volume (v/v). This deficiency is made up for in the teachings of Bamford.
Regarding claim 9, Bamford teaches a LAMP assay wherein the sensitivity is increased by enhancing the reaction rate and reducing the likelihood of false-positives when DMSO is present in the LAMP reaction between about 1 to 5% v/v (“The present invention improves upon the conventional LAMP method to enhance the reaction rate and to reduce the likelihood of false-positives” [0005]; “The present invention relates to a novel buffer formulation for reducing reaction time compared to conventional LAMP buffer” [0006]; “In one embodiment, the buffer further comprises an enhancer. In one embodiment, the enhancer is 2%-4% DMSO” [0008]).
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to perform the simple substitution of DMSO taught by Bamford for guanine hydrochloride as the additional reagent to enhance sensitivity of the LAMP assay in the method of Tanner (“additional reagents to enhance sensitivity of the assay such as a guanidine salt” Tanner [0016]). (MPEP §2143 I. B.). One of ordinary skill in the art could have performed this substitution and would have found the results of this substitution predictable because both DMSO and guanine hydrochloride are taught as being used in LAMP assays in order to increase sensitivity. Therefore, the invention as a whole of claim 9 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Claim 10 is rejected under 35 U.S.C. 103 as being unpatentable over Tanner et al. (U.S. Patent Application Publications Cite No 1 in IDS filed 26 March 2025)(US 2021/0285064, published 16 September 2021, effectively filed 12 March 2020), herein Tanner, and in view of Bamford (US 2016/0289752, published 6 October 2016, effectively filed 31 March 2015) as applied to claim 9 above, and further in view of Piskur et al. (“Aggregated DNA in ethanol solution” FEBS Lett 375(3), pages 174-178 (1995)), herein Piskur, and Oscorbin et al. (“Derivatives of Bst-like Gss-polymerase with improved processivity and inhibitor tolerance” Nucleic Acids Res 45(16), pages 9595-9610 (2017)), herein Oscorbin.
Regarding claim 10, Tanner teaches the method of claim 8 (see 35 U.S.C. 102 rejection of claim 8 above) and, in combination with the teachings of Bamford, teach a modified version of method of claim 8 wherein DMSO is used as an additive in the LAMP reaction that both increases sensitivity of the reaction (see 35 U.S.C. 103 rejection of claim 9 above) and lowers the melting temperature (“dimethyl sulfoxide (DMSO) or formamide can be used as the regulator for melting temperature (Tm) […] these regulators for melting temperature act for lowering melting temperature” Bamford [0090]). However, neither Tanner nor Bamford teaches the method wherein ethanol is present in the LAMP reaction between about 1.5 to 3% v/v. This deficiency is made up for in the teachings of Piskur and the teachings of Oscorbin.
Regarding claim 10, Oscorbin teaches that the Bst-like polymerase Gss-polymerase and fusion proteins made with it (His-Gss, Gss-His, DBD-Gss, Sto-Gss, and Gss-Sto) can be used in place of Bst polymerase in a LAMP assay and that they have the advantage of being particularly resistant to inhibition by ethanol with only minor changes in time-to-threshold below 5% ethanol concentration (“we cloned and characterized the Bst-like polymerase Gss-polymerase” page 9596 left column paragraph 3; “we examined the influence of heparin, NaCl, EDTA, ethanol, urea and whole blood on the ability of fused proteins to act as DNA polymerase in qLAMP (Figure 11)” page 9606 left column paragraph 1; Figure 11D).
Regarding claim 10, Piskur teaches that low concentrations, including those between 1.5 and 3% v/v, of ethanol are can be used to reduce the melting temperature of DNA (“When ethanol is added at low ethanol concentrations, NaDNA still remains in the B form, but Tm of DNA decreases” page 175 left column paragraph 2; Fig. 1 encompassing a range including 1.5-3% and teaching that they induce a small decrease in melting temperature).
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to perform the simple substitution of the Gss polymerase or the fusions of Gss polymerase taught by Oscorbin for the polymerase suitable for LAMP reactions exemplified by Bst polymerase in the method of the combination of Tanner and Bamford (MPEP §2143 I. B.). One of ordinary skill in the art could have performed this substitution and would have found the results of this substitution predictable because Oscorbin demonstrates the suitability of the Gss polymerase and its fusions in a LAMP reaction and the method of Tanner that it substitutes into a LAMP reaction. Therefore, the combination of Tanner, Bamford, and Oscorbin would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to perform the simple substitution of ethanol as an additive to reduce the melting temperature of DNA as taught by Piskur for DMSO in the method of the combination of Tanner, Bamford, and Oscorbin since DMSO effects the LAMP reaction of the method by reducing the melting temperature of DNA (MPEP §2143 I. B.). One of ordinary skill in the art could have performed this substitution and would have found the results of this substitution predictable because both ethanol and DMSO are taught to be able to reduce the melting temperature of DNA and because the polymerases taught by Oscorbin for use in the LAMP reaction are minimally inhibited by the low concentrations of ethanol that reduce the melting temperature (so the use of ethanol does not make the method non-functional). Therefore, the invention as a whole of claim 10 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Claim 11 is rejected under 35 U.S.C. 103 as being unpatentable over Tanner et al. (U.S. Patent Application Publications Cite No 1 in IDS filed 26 March 2025)(US 2021/0285064, published 16 September 2021, effectively filed 12 March 2020), herein Tanner, and in view of Bamford (US 2016/0289752, published 6 October 2016, effectively filed 31 March 2015) as applied to claim 9 above, and further in view Fuller et al. (US 5,500,339, issued 19 March 1996), herein Fuller.
Regarding claim 11, Tanner teaches the method of claim 8 (see 35 U.S.C. 102 rejection of claim 8 above) and, in combination with the teachings of Bamford, teach a modified version of method of claim 8 wherein DMSO is used as an additive in the LAMP reaction that both increases sensitivity of the reaction (see 35 U.S.C. 103 rejection of claim 9 above) and lowers the melting temperature (“dimethyl sulfoxide (DMSO) or formamide can be used as the regulator for melting temperature (Tm) […] these regulators for melting temperature act for lowering melting temperature” Bamford [0090]). However, neither Tanner nor Bamford teaches the method wherein glycerol is present in the LAMP reaction between about 0.5 to 5% v/v. This deficiency is made up for in the teachings of Fuller.
Regarding claim 11, Fuller teaches that a 1% solution of glycerol lowers the melting temperature of DNA by about 0.4° C (“a 1% solution of glycerol will lower the melting temperature (Tm) of DNA by about 0.4° C” col 1 lines 29-31).
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to perform the simple substitution of glycerol as an additive to reduce the melting temperature of DNA as taught by Fuller for DMSO in the method of the combination of Tanner and Bamford since DMSO effects the LAMP reaction of the method by reducing the melting temperature of DNA (MPEP §2143 I. B.). One of ordinary skill in the art could have performed this substitution and would have found the results of this substitution predictable because both glycerol and DMSO are taught to be able to reduce the melting temperature of DNA and 1% glycerol would not be expected to prevent the LAMP reaction from functioning. Therefore, the invention as a whole of claim 11 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Claim 12 is rejected under 35 U.S.C. 103 as being unpatentable over Tanner et al. (U.S. Patent Application Publications Cite No 1 in IDS filed 26 March 2025)(US 2021/0285064, published 16 September 2021, effectively filed 12 March 2020), herein Tanner, as applied to claims 1-8 and 13-15 above, and in view of Ranoa et al. (Non-Patent Literature Document Cite No 3 in IDS filed 26 March 2025)(“Saliva-Based Molecular Testing for SARS-CoV-2 that Bypasses RNA Extraction” bioRxiv (2020)), herein Ranoa.
Regarding claim 12, Tanner teaches the method of claim 8 (see 35 U.S.C. 102 rejection of claim 8 above). However, Tanner does not teach the method wherein glycogen is present in the LAMP reaction between about 1 to 4 micrograms per microliter. This deficiency is made up for in the teachings of Ranoa.
Regarding claim 12, Ranoa teaches that glycogen is an RNA stabilizing agent and uses it in an RT-qPCR assay for SARS-CoV-2 detection (“various additives have been explored for their ability to enhance SARS-CoV-2 detection. Therefore […] RNA stabilizing agents, including […] glycogen […] were examined” page 9 paragraph 1; Supporting Figure 2, note that the amplification was not negatively impacted comparing glycogen vs no treatment) wherein the glycogen is present in the reaction at microgram per microliter (“Additives include […] glycogen (1 µg/µL)” page 22 paragraph 3).
In view of Ronoa’s teaching that glycogen is an RNA stabilizing agent and that the method of Tanner is taught such that RNA is one of the options for a target nucleic acid (“The target nucleic acid may be any DNA or RNA of interest” Tanner [0020]), one of ordinary skill in the art would be motivated to improve the method of Tanner by using the stabilization of glycogen taught by Ronoa to decrease any loss of target RNA in the method. One of ordinary skill in the art would have a reasonable expectation of success in this combination because Ronoa teaches that glycogen does not negatively affect amplification in an RT-qPCR-based method and therefore would be unlikely to negatively affect amplification in a LAMP-based method. Therefore, the invention as a whole of claim 12 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Claim 16 is rejected under 35 U.S.C. 103 as being unpatentable over Tanner et al. (U.S. Patent Application Publications Cite No 1 in IDS filed 26 March 2025)(US 2021/0285064, published 16 September 2021, effectively filed 12 March 2020), herein Tanner, as applied to claims 1-8 and 13-15 above, and in view of Sun et al. (US 2021/0292820, effectively filed 20 March 2020), herein Sun.
Regarding claim 16, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above), wherein the LAMP can differentiate between two or more viruses (“a plurality of different target nucleic acids may by analyzed by the methods described herein, for example, a plurality of sets of primers in the third tube may target sequences in the genome of SARS-CoV-2 and a different plurality of sets of primers in the third tube may target sequences in the genome of influenza virus. Different indicators associated with the oligonucleotide primers for influenza and coronavirus can provide different color or fluorescence endpoints corresponding to the presence of either or both viruses” [0143]). However, Tanner does not teach the method wherein the viruses differentiated are two more variants of SARS-CoV-2. This deficiency is made up for in the teachings of Sun.
Regarding claim 16, Sun teaches that it is advantageous for an assay to be able to differentiate SARS-CoV-2 variants in a sample because variants have different clinical outcomes and knowing the variant a patient has may enable tailoring care to the patient for that specific variant (“emerging evidence points to different clinical outcomes for individuals infected with SARS-CoV-2 that contain certain S gene mutations and the need to alter care pathways and methods of care based on the SARS-CoV-2 variant infecting a patient” [0007]) and teaches a method of differentiating SARS-CoV-2 variants that can be applied to LAMP assays (“performing an allelic discrimination plot analysis based on the obtained Ct values or RFUs to identify one or more SARS-CoV-2 mutations in the specimen; and (vi) determining the SARS-CoV-2 variant contained in the specimen based on the identified SARS-CoV-2 mutations” [0010] and rest of [0010]; “Reverse transcription loop-mediated isothermal amplification or reverse transcription isothermal amplification may be utilized instead of real time reverse transcription polymerase chain reaction” [0011]).
In view of Sun’s teaching of the advantage of an assay that can differentiate two or more variants of SARS-CoV-2 in enabling specific patient care based on the variant, one of ordinary skill in the art would be motivated to combine Tanner’s method of multiplex LAMP reactions with Sun’s method of differentiating SARS-CoV-2 variants to improve the method of multiplexed LAMP reactions with the ability to differentiate SARS-CoV-2 variants. One of ordinary skill in the art would have a reasonable expectation of success in this combination because Sun teaches that their method can be done using LAMP reactions as are used in the method of Tanner. Therefore, the invention as a whole of claim 16 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Claims 17-18 are rejected under 35 U.S.C. 103 as being unpatentable over Tanner et al. (U.S. Patent Application Publications Cite No 1 in IDS filed 26 March 2025)(US 2021/0285064, published 16 September 2021, effectively filed 12 March 2020), herein Tanner, as applied to claims 1-8 and 13-15 above, and in view of Quyen et al. (Non-Patent Literature Document Cite No 4 in IDS filed 26 March 2025)(“Classification of Multiple DNA Dyes Based on Inhibition Effects on Real-Time Loop-Mediated Isothermal Amplification (LAMP): Prospect for Point of Care Setting” Front Microbiol 10: 2234 (2019)), herein Quyen, and as evidenced by Eischeid (“SYTO dyes and EvaGreen outperform SYBR Green in real-time PCR” BMC Res Notes 4: 263 (2011)) and Monis et al. (“Comparison of SYTO9 and SYBR Green I for real-time polymerase chain reaction and investigation of the effect of dye concentration on amplification and DNA melting curve analysis” Anal Biochem 340(1), pages 24-34 (2005)), herein Monis.
Regarding claims 17 and 18, Tanner teaches the method of claim 1 (see 35 U.S.C. 102 rejection of claim 1 above) wherein step (c) comprises detecting an intercalating-dye (“Examples of suitable indicators include […] an intercalating molecule fused or linked via a linker group to an optically detectable dye moiety” [0044]). However, Tanner does not teach the method further comprising after step (b) and before step (c), a step (c’) comprising: (c’) heating the plurality of LAMP reactions to a temperature of at least about 80 °C or that the intercalating-dye is fluorescent. This deficiency is made up for in the teachings of Quyen.
Regarding claims 17 and 18, Quyen teaches a LAMP assay wherein after the LAMP reaction is terminated by heating it to 90°C (“the reactions were then terminated by heating to 90°C for 10 min” page 3 left column second paragraph). Furthermore, MPEP §2144.04 instructs that any order of steps is prima facie obvious in the absence of new or unexpected results: “In re Burhans, 154 F.2d 690, 69 USPQ 330 (CCPA 1946) (selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results)”. Therefore, it would be prima facie obvious to one of ordinary skill in the art for the detection step at the end of the method of Tanner to be done after the termination of the LAMP reaction by heating to a temperature of 90°C as taught by Quyen when combining the methods of Tanner and Quyen. Quyen also teaches that dyes such as SYTO 9, SYTO 82, SYTO 16, and SYTO 13 are advantageous for use in LAMP assays due to having no inhibitory effect and a high signal-to-noise ratio (SNR)(“SYTO 9, SYTO 82, SYTO 16, SYTO 13, and Miami Yellow were the best dyes with no inhibitory effect, low LOD and high SNR in the real-time LAMP reactions” Abstract), and Eischeid provides evidence that the SYTO dyes taught by Quyen are cyanine dyes (“The SYTO dyes constitute a large family of commercially available cyanine dyes” page 1 right column paragraph 2) and Monis provides evidence that SYTO 9 is an intercalating dye (“the intercalating dye SYTO9” Abstract).
It would have been obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to perform the simple substitution of the fluorescent intercalating dye SYTO9 taught by Quyen for the intercalating dye as an indicator in the method of Tanner (MPEP §2143 I. B.). One of ordinary skill in the art would be further motivated to make this substitution because Quyen teaches that the SYTO9 dye has the advantage of having no inhibitory effect on LAMP reactions and high SNR. One of ordinary skill in the art could have performed this substitution and would have found the results of this substitution predictable because the substitution of one dye for visualizing a result for another dye would not be expected to harm the reaction and because Quyen teaches that SYTO9 does not inhibit LAMP reactions. Therefore, the invention as a whole of claims 17-18 would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention.
Conclusion
Claims 1-20 are rejected.
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/JEFFREY BELLAH/Examiner, Art Unit 1683
/ANNE M. GUSSOW/Supervisory Patent Examiner, Art Unit 1683
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