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
Acknowledgement is made of Applicant’s claim to priority from US Applications 63/138,341; 63/138,323; 63/138,337; 63/138,321; 63/138,320; 63/138,318; 63/138,314; 63/138,312 and 63/138,310 filed 01/15/2021, from US Applications 63/138,316 filed 01/20/2021 and from US Application 63/148,527 filed 02/11/2021.
Application Status
Claims status: Claim 13 is cancelled. Claims 1-12 and 14-22 are pending. Claims 1-12 and 14-22 are under examination.
Any objection or rejection not reiterated herein has been overcome by Applicant’s amendments. Applicant’s amendments and arguments have been thoroughly reviewed, but are not persuasive to place the claims in condition for allowance for the reasons that follow.
Information Disclosure Statement
The information disclosure statements (IDSs) submitted on 10/08/2025 and 11/18/2025 were filed after the mailing date of the Office Action on 08/18/2025. The submissions are in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statements are being considered by the examiner.
The following rejections are maintained from Office Action dated 08/18/2025:
Claim Rejections - 35 USC § 103
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 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:
A. Determining the scope and contents of the prior art.
B. Ascertaining the differences between the prior art and the claims at issue.
C. Resolving the level of ordinary skill in the pertinent art.
D. Considering objective evidence present in the application indicating obviousness or non-obviousness.
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.
Claims 1, 3-4, 6-12, 14-15 and 17-22 are rejected under 35 U.S.C. §103 as being unpatentable over Wolf (Wolf, A. et al. WO 2019/073049 A1, published April 18, 2019), Naik (Naik, P. et al. “ Nucleic Acid amplification on paper substrates”. Paper Microfluidics (2019), pp: 115-146), and Bst DNA polymerase-NEB pdf and WarmStart® RTx Reverse Transcriptase-NEB pdf (both downloaded from Internet on 12/01/2024; previously cited).
Regarding claim 1, Wolf teaches solid-phase LAMP that can successfully and specifically amplify and detect small amounts of DNA (see “[S]hort conclusion” ¶, page 43 of 96, lines 17-19). Wolf teaches a Solid phase, see page 10, lines 18-35 and page 11, lines 1-9; page 23, lines 4-14; page 32, lines 1-8).
Wolf also teaches exact amounts of reagents in Tables 3 to 5 (pages 41 to 42), see below:
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In Table 3 as example, Wolf teaches:
A couple of primers (primer E (F3) and primer F (B3) specific to target sequences of interest. (see page 9, line 8).
a DNA polymerase, i.e. WarmStart® Bst 2.0® DNA polymerase from New England Biolabs (NEB).
A re-solubilization agent, i.e. water.
Betaine.
Wolf is silent on ammonium sulfate. Examiner interprets that Wolf’ reagents do not include ammonium sulfate.
In the Specification, Applicant defines “hygroscopic agents” as “include glycerol, ethanol. Methanol, calcium chloride, potassium chloride, calcium sulfate and combinations thereof” (see [0015]). And according to Applicant’s definition of “substantially free” mentioned above, it “refers to the complete or nearly complete extent of degree of an action, characteristic, property, state, structure, item, or result” (see instant Specification ¶ [0090]).
Regarding the amount of hygroscopic agents, Wolf add a hygroscopic agent, i.e. glycerol, in the LAMP mix since the stock solution of the DNA polymerase contains glycerol. Wolf add the enzyme at a final concentration of 8 Units in 25 µl of reaction (see Table 3, page 41 of 96). This dilution uses the enzyme at a final effective concentration of 0.32 U/µl, since the total is 8 units of enzyme. The stock solution being at 120 U/µl, according to Bst 2.0® DNA polymerase-New England Biolabs pdf, only 0.067 µl was needed, therefore, 0.034 µl of glycerol were carried over, in a total volume of 25 µl. There is 0.14% of a hygroscopic agent in the composition taught by Wolf, which is less than what is required by claim 1.
Wolf does not teach a reverse transcriptase, since Wolf teaches a LAMP reaction. However, Naik teaches RT-LAMP and reverse transcriptase for RT-LAMP are commercially available as evidenced by WarmStart® RTx Reverse Transcriptase-NEB pdf.
Naik teaches Loop-Mediated Isothermal Amplification (LAMP) assays on page 120, section 8.3.1 and Figure 8.3. Naik teaches that RT-LAMP reactions are well known (see page 121, lines 6-11). Naik teaches that solid-phase LAMP are known and used in the prior art (see Figures 8.4 and 8.5).
Naik teaches the requirement for reverse transcriptase for RT-LAMP (see page 121, lines 10-11).
Incidentally, Naik also teaches that “[D]ried reagents need to be thoroughly rehydrated prior to DNA amplification. This is achieved by applying rehydration buffer [i.e. re-solubilization agent, e.g. water] or the liquid sample on the paper substrate” (see page 142, lines 3-5).
The WarmStart® RTx Reverse Transcriptase-NEB pdf teaches a specific reverse transcriptase adapted for RT-LAMP reaction. Regarding the amount of hygroscopic agent, i.e. glycerol carried over when using this reagent, WarmStart® RTx Reverse Transcriptase-NEB pdf also teaches glycerol as a component of the storage buffer for the RTx enzyme (50% glycerol; see pdf). However, the enzyme is also offered in a glycerol-free format.
It would have been prima facie obvious to a person of ordinary skill in the art, before the effective filing date, to have combined the teachings of Wolf with the teachings of Naik and transform the solid-phase LAMP of Wolf into an RT-LAMP. One with ordinary skills in the art motivated in performing a LAMP reaction which includes detection of RNA viruses, would have performed this modification with a reasonable expectation of success based on results obtained by Wolf and those reviewed by Naik and arrived at the claimed invention.
Regarding claim 3, Wolf is silent on volatile agents in the LAMP reaction. Naik teaches the use of folds in the paper solid phase to ensure rehydration (re-solubilization) of reagents by the sample (see page 123, lines 2-5; and section 8.5.3., lines 1-7). Therefore, there was no use of volatile agents such as ethanol to resuspend reagents. Naik teaches the use of the sample or DNA amplification buffer to rehydrate the reagents deposited on the paper substrate (see page 142, lines 3-5).
Instant Disclosure’s definition of “substantially free” is described above , and it “refers to the complete or nearly complete extent of degree of an action, characteristic, property, state, structure, item, or result” (see instant Specification ¶ [0090]).
Naik is silent on volatile agents in the RT-LAMP reaction. Examiner interprets that there was no volatile agents added to the composition for RT-LAMP reaction. Therefore, the combination of references teaches a composition “substantially free of volatiles agents” since no volatiles agents are used. The obviousness of the combination of references teachings the claimed invention’s elements are discussed above.
Regarding claims 4 and 6, in the Specification, Applicant defines “hygroscopic agents” as “include glycerol, ethanol, Methanol, calcium chloride, potassium chloride, calcium sulfate and combinations thereof” (see [0015]). The instant Specification’s definition of “substantially free of hygroscopic agents”. Instant Disclosure’s definition of “substantially free” is described above , and it “refers to the complete or nearly complete extent of degree of an action, characteristic, property, state, structure, item, or result” (see instant Specification ¶ [0090]).
Wolf teaches the use of the WarmStart® Bst 2.0® DNA polymerase from New England Biolabs (NEB), at a final concentration of 8 Units in 25 µl of reaction (see Table 3, page 41 of 96). This dilution uses the enzyme at a final effective concentration of 0.32 U/µl, since the total is 8 units of enzyme. The stock solution being at 120 U/µl, according to Bst 2.0® DNA polymerase-New England Biolabs pdf, only 0.067 µl was needed, therefore, 0.034 µl of glycerol were carried over, in a total volume of 25 µl. There is 0.14% of a hygroscopic agent in the composition taught by Wolf.
The WarmStart® RTx Reverse Transcriptase-NEB pdf also teaches glycerol as a component of the storage buffer for the RTx enzyme (50% glycerol; see pdf). However, the enzyme is offered in a glycerol-free format.
Wolf teaches a concentration of hygroscopic agents that is closer to zero and much less than 1%.
Wolf and Naik do not teach ethanol, methanol, calcium chloride nor calcium sulfate in the RT-LAMP reaction.
WarmStart® Bst 2.0® DNA polymerase from New England Biolabs pdf teach glycerol only in enzyme stock solution.
Therefore, Examiner interprets that Wolf and Naik do not add these reagents in the reactions.
Regarding claims 7 and 8, Triton™ X-100 is taught by Wolf is considered a non-ionic surfactant. Wolf teaches that the primers are mixed in a solution comprising this surfactant (See page 39 of 96, line 31). Wolf also teaches the use of BSA (bovine serum albumin) as a blocking agent to treat the surface of the slide (see page 40 of 96, line 4). Naik teaches that “surface passivation of paper substrates” can be done using BSA (see page 136, line 3). Naik also teaches BSA as a stabilizing agent in the LAMP reaction (see page 124, line 5).
The obviousness of the combination of references over the claimed elements is described above.
Regarding claims 9-12, Wolf teaches influenza virus and norovirus as viral pathogens (see page 27 of 96, line 3; page 43 of 96, lines 21-32).
Naik teaches viral pathogens detected using RT-LAMP (see page 124, lines 14-44), including bovine herpesvirus-1, influenza virus (H1N1), human papillomavirus and HIV-1.
The obviousness of the combination of references over the claimed elements are discussed above.
Regarding claims 14 and 15, Wolf teaches “water” as a non-discoloration agent (see Table 3). Naik teaches non-discoloration additives that can be a sugar, a buffer or a combination of both; Naik teaches the use of trehalose as a stabilizing agent that help in maintaining the structure and function of the enzymes in the dried form (see page 141, section 8.5.3., lines 9-14).
The obviousness of the combination of references over the claimed elements are discussed above.
Regarding claims 17, 21 and 22, the combination of references Wolf, Naik, and NEB Warmstart® reagents pdfs renders obvious elements in the composition of claim 1. Wolf also teaches that some polymerase enzymes has strand displacement activity at moderate temperatures, around 20-37⁰C, while others are active at elevated temperatures around 65⁰C (see page 24 of 96, lines 30-34).
Wolf teaches coupling the SP-primer onto the solid support to enable amplification in the solid support ( see page 23 of 96, lines 25-27). Wolf further teaches that the SP-LAMP (solid phase-LAMP) method includes not only a sample and suitable primers for amplification thereof, but also LAMP amplification mixture that is mixed with the sample and the primer set . The amplification mixture comprises all the building blocks necessary for nucleotide amplification and suitable polymerase enzymes to facilitate the amplification (see page 24 of 96, lines 16-20).
Naik also teach methods for LAMP analysis on a solid phase medium, depositing a biological sample onto the solid phase medium and heating the assembly to an isothermal temperature sufficient to facilitate a LAMP reaction (see section 8.3.1., pages 120-125).
The obviousness of the combination of references over the claimed elements are discussed above.
Regarding claims 18 and 19, Wolf teaches that the term “sample” refers to any biological solution, entity or subject, including biological fluids (see page 9 of 96, lines 6-14). Wolf further teaches that the sample can be selected from blood, sputum, swabs and feces (see page 26 of 96, lines 15-20).
Naik teaches that the biological sample in a RT-LAMP assay can be oral fluids, i.e. saliva (page 121, lines 9 and 14), human blood (page 121, line 20).
The obviousness of the combination of references over the claimed elements are discussed above.
Regarding claim 20, Wolf teaches influenza virus and norovirus as viral pathogens (see page 27 of 96, line 3; page 43 of 96, lines 21-32).
Naik teaches viral pathogens detected using RT-LAMP (see page 124, lines 14-44), including bovine herpesvirus-1, influenza virus, human papillomavirus and HIV-1.
The obviousness of the combination of references over the claimed elements are discussed above.
Claims 2 and 16 are rejected under 35 U.S.C. §103 as being unpatentable over Wolf (Wolf, A. et al. WO 2019/073049 A1, published April 18, 2019), Naik (Naik, P. et al. “ Nucleic Acid amplification on paper substrates”. Paper Microfluidics (2019), pp: 115-146), and Bst DNA polymerase-NEB pdf and WarmStart® RTx Reverse Transcriptase-NEB pdf (both downloaded from Internet on 12/01/2024; previously cited), as applied to claim 1, and in further view of Ahn (Ahn, S.J. et al. “Rapid and simple colorimetric detection of multiple influenza viruses infecting humans using a reverse transcriptional loop-mediated isothermal amplification (RT-LAMP) diagnostic platform”. BMC Infectious Diseases, Vol. 19 (2019), p: 676).
The rejection of claim 1 is described above. The combination of Wolf, Naik and Bst DNA polymerase-NEB pdf and WarmStart® RTx Reverse Transcriptase-NEB pdf renders elements of claim 1 obvious. Wolf teaches betaine which is also an antioxidant. However Naik does not teach an antioxidant directly in the RT-LAMP reaction. Therefore, the combination of references does not teach another antioxidant besides Betaine.
Regarding claims 2 and 16 reciting “further comprising an antioxidant” (claim 2) and “an indicator” (claim 16), Ahn teaches the use of Phenol Red, which is both an antioxidant and an indicator in the RT-LAMP reaction (see page 3 of 12, right column, second paragraph).
Therefore, it would have been obvious to one with ordinary skills in the art, before the effective filing date, to have modified the composition for RT-LAMP taught by Wolf modified by Naik, and add the indicator and antioxidant phenol red to the composition as taught by Ahn. One motivated in a rapid colorimetric test using convenient reagents for a result directly observable by naked eyes could performed this modification. One motivated in reagents already optimized and compatible with the DNA polymerase and the reverse transcriptase from NEB, would be interested in the WarmStart® Colorimetric LAMP 2X Master Mix containing phenol red used by Ahn (see page 3 of 12, right column, second paragraph). One with ordinary skills in the art could performed this modification with a reasonable expectation of success, since Ahn also teaches an RT-LAMP using WarmStart® reagents with success, and would arrived at the claimed invention.
Claim 5 is rejected under 35 U.S.C. §103 as being unpatentable over Wolf (Wolf, A. et al. WO 2019/073049 A1, published April 18, 2019), Naik (Naik, P. et al. “ Nucleic Acid amplification on paper substrates”. Paper Microfluidics (2019), pp: 115-146), and Bst DNA polymerase-NEB pdf and WarmStart® RTx Reverse Transcriptase-NEB pdf (both downloaded from Internet on 12/01/2024; previously cited), as applied to claim 1, and in further view of Lin (Lin, X. et al. “Influence of water evaporation/absorption on the stability of glycerol-water marbles”. RSC Advances, Vol. 9 (2019), p: 34465).
The rejection of claim 1 is described above. The combination of references Wolf, Naik and Bst DNA polymerase-NEB pdf and WarmStart® RTx Reverse Transcriptase-NEB pdf renders elements of claim 1 obvious, however, it does not teach elements of claim 5, i.e. “the composition is substantially free of agents that absorb more than about 10 wt% when stored between about 40% and about 90% relative humidity at 25⁰ C”.
Regarding claim 5, the references combined, i.e. Wolf, Naik, modified by the NEB WarmStart® reagents pdfs, provide for a composition that is substantially free of hygroscopic agents (i.e. glycerol), in which the concentration of hygroscopic agent is less than 1 % of total volume, see above. However, the combination does not teach characteristics of glycerol, i.e. the only hygroscopic agent capable of absorbing water, in various conditions such as 40% to about 90% of relative humidity when stored at 25 ⁰C.
However, Lin teaches glycerol-water marbles and their stability in different environmental conditions, i.e. Relative Humidity (RH) from 26% to 82% (see Figure 3). In this figure 3, it is clear that the stability of the glycerol-water marbles depends on the original concentration/amount of glycerol in the marble. The pure water marble (0% of glycerol) decreased within 52 minutes at low RH indicating that almost no water was left in the marble, the water marble lost weight until complete collapse. However, higher RH, there was slower evaporation and better resistance to evaporation. When adding glycerol at 33.3%, there is weight loss in the marble, i.e. evaporation, with evaporation resistance at higher RH, but no mass gain. Li teaches mass gain for marbles containing 50 to 100% glycerol at 82% RH. There is mass gain when glycerol is at 71.4% or higher at 76% RH or higher. All measurements were performed in an air-conditioned laboratory at 297.2 +/- 1.0 K (which is approximately 24-25 ⁰C) (see page 2, left column, “Characterization” paragraph).
Therefore, since the amount of glycerol in the LAMP reaction is less than 0.15%, at RH from 40% to 82 %, the hygroscopic agent, glycerol, will not be able to absorb water and gain weight in an air-conditioned laboratory at 297.2 +/- 1.0 K (24-25 ⁰C).
It would have been obvious to one with ordinary skills in the art, before the effective filing date, to have combined the teachings of Wolf, Naik, and Bst DNA polymerase-NEB pdf and WarmStart® RTx Reverse Transcriptase-NEB pdf, and measured the ability of the total hygroscopic agents’ amount in the mixture to absorb water, under the conditions taught by Lin, and tried and maintained the concentration of hygroscopic agents close to zero, as these are the appropriate conditions to ensure that there is no water absorption under storage conditions. Water absorption would modify the osmolarity of other dried components deposited onto the solid phase’s surface and modify their reactivity with samples. Naik teaches that the presence of stabilizing agents such as glycerol that also increases the fluid viscosity may contribute to inhomogeneous dispersion of assay components and indirectly affects detection signal intensity (see section 8.5.3., page 142). One with ordinary skills in the art motivated in optimum storage conditions under 40% to 90% RH at 25 ⁰C, and maintaining the integrity and the reproducibility of the assays, would have tried and minimized the amount of hygroscopic agents capable of absorbing water as suggested by the teachings of Lin, with a reasonable expectation of success and arrived at the claimed invention.
Response to Arguments
Applicant's arguments filed 11/18/2025 have been fully considered but they are not persuasive.
Applicant states in Remarks on page 6 of 9 that “It would not be appropriate for the Office to take official notice of facts without citing a prior art reference where the facts asserted to be well known are not beyond dispute, or are not capable of instant and unquestionable demonstration as being well-known.”
In response, Examiner used the terms “well known” because Naik is reviewing the state of the art and stated :
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Therefore, Examiner used the terms “well-known” since the reference refers to the use of paper as a solid phase as “obvious”, detailed in a reference dating from 2011.
Applicant further argues : “The mere fact that the prior art may be modified in the manner suggested by the Examiner does not make the modification obvious unless the prior art suggested the desirability of the modification."
In response, Wolf teaches on page 23 that paper is an alternative for the solid support:
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Since Naik teaches that the use of paper is an “obvious choice”, one of ordinary skills in the art, under affordability restraints and for mass production purposes, would pick paper as the solid phase.
Applicant also argues on pages 6 and 7 of 9, that “Claim 1 recites inter alia: "...wherein the composition is substantially free of ammonium sulfate..." The Office at p. 6 of the Action states "...that Wolf' reagents do not include ammonium sulfate..." in what is, essentially, a de facto taking of notice that fails to consider the reference as a whole” and “This is important in the context of the pH sensitive nature of the indicators taught and claimed by Applicant. The Office factually errs in asserting that a supposed silence of Wolff regarding ammonium sulfate means that ammonium sulfate is not present.”.
In response, Applicant is reminded that claim 1 is drawn to a composition with a list of reagents. The method of using is not the purpose of the claim.
Examiner stated that Wolf is silent on Ammonium sulfate. Wolf is indeed silent on having ammonium sulfate in the PCR composition. It is not taking “Official notice” when stating that a reference is silent on a substance.
Therefore, the compounds listed are taught or are rendered obvious by the combination of references. The pH of the composition is not claimed in claim 1. In response to applicant's argument that the references fail to show certain features of the invention, it is noted that the features upon which applicant relies (i.e., sulfuric acid formation, pH) are not recited in the rejected claim(s). Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
Applicant also argues that “Further, although Naik does discuss a high-level overview of LAMP reactions and various substrates, nowhere does Naik teach or discuss anything to do with pH sensitivity or optimizing any substrates for use with a pH indicator as taught and claimed by Applicant. Instead, all one of ordinary skill in the art is left with is the basics of LAMP as taught at a high-level overview with multiple references pointing to a source that explicitly teaches the usage of ammonium sulfate.”
In response, Applicant is reminded that claims 1-12 and 14-16 are drawn to a composition. There is no claim that is drawn to optimum pH or to a method of avoiding raising the pH on a solid phase support. Although the claims are interpreted in light of the specification, limitations from the specification are not read into the claims. See In re Van Geuns, 988 F.2d 1181, 26 USPQ2d 1057 (Fed. Cir. 1993).
As a person of ordinary skills can obtain glycerol-free buffers for WarmStart® RTx reaction, from the alternatives offered on the website, it is also possible to further optimize conditions for LAMP, finding other alternative buffers for the enzymes through routine optimization.
Examiner would also like to point out the following on New England Biolabs™ ‘s website at WarmStart® RTx Reverse Transcriptase | NEB, “FAQs & Troubleshooting”, question no. 6:
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And following the link:
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Then, searching for composition of Buffers on the website:
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Searching for citations and literature on the same website, they show that NEBuffers 1.1, 2.1, 3.1 and CutSmart™ Buffer have been available since before April 2020. Therefore, one with ordinary skills in the art, before the effective filing date, could have substituted the reaction’s buffer with any of NEBuffer 1.1, 2.1, 3.1 and CutSmart™ and optimize to obtain the claimed composition and the claimed technique. And In response to applicant's arguments against the references individually, one cannot show non-obviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986).
The rejections are maintained.
Conclusion
No claim is allowed
THIS ACTION IS MADE FINAL. Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
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/A.D./Examiner, Art Unit 1636
/NANCY J LEITH/Primary Examiner, Art Unit 1636