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 .
Status of the Application
Applicant’s amendment and remarks submitted on 08/07/2026 are acknowledged.
Claims 1-20 are pending.
Claim 12 is amended.
Claims 1-20 have been examined on the merits.
Applicant’s election without traverse of the species “sodium azide” from the newly amended claim 12, in response to the species requirement set forth in the prior office action, is acknowledged. Claims 1-20 are under consideration by the Examiner, 37 CFR 1.142(b), as being drawn to the elected species.
Specification
The abstract of the specification is objected to, because it contains more than 150 words. Applicant is reminded of the proper language and format for an abstract of the disclosure. The abstract should be in narrative form and generally limited to a single paragraph on a separate sheet within the range of 50 to 150 words. It is important that the abstract not exceed 150 words in length since the space provided for the abstract on the computer tape used by the printer is limited. Appropriate correction is required.
Priority
This application, U.S. Application number 18/286315, is a national stage entry of International Application Number PCT/US2022/031008, filed on 05/26/2022, which claims for domestic priority under 35 U.S.C. 119(e) to provisional application No. 63/193,904 filed on 05/27/2021.
Information Disclosure Statement
The information disclosure statements (IDS) submitted on 09/10/2024 and 03/14/2025 are acknowledged. The submissions are in compliance with the provisions of 37 CFR 1.97., and have been considered by the examiner.
Drawings
The drawings submitted on 10/10/2023 have been reviewed and are accepted by the Examiner for examination purposes.
Claim Objections
Claims 15-16 are objected to because of the recitation of “DMSO”. Abbreviations should be spelled out at least once in the claims. Appropriate correction is required
Claim Rejections - 35 USC § 112(b), or 112, Second Paragraph
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 1-20 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 pre-AIA the applicant regards as the invention.
Claim 1 is indefinite due to the recitation of “incubating the biological tissue with a decolorizing solution … to substantially decolorize the biological tissue”. The recited term “substantially” is a relative term. This term is not defined in the specification with regard to what specific level of tissue decolorization can be considered as being substantially decolorized. In the absence of any benchmark or definition, it is unclear at what specific level of tissue decolorization a tissue can be considered as being substantially decolorized.
The remaining claims are rejected for depending from an indefinite 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.
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 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.
Claims 1-3, 5-8 and 19 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Miyawaki et al. (US 2017/0370810, 2017, cited in IDS).
Miyawaki et al. teach a clearing reagent (i.e. a decolorizing solution) as well as a method of making a biological tissue transparent (i.e. decolorizing a biological tissue) by incubating the tissue with a decolorizing solution comprising a surfactant for a sufficient time to decolonize the biological tissue, wherein the surfactant is Triton X-100 or Tween (abstract; Claims 1, 5-6, 9, and 11; para 0088/last 7 lines; Examples: page 16/paras 0185 and 0187, and Fig. 8); wherein the decolorizing solution comprising Triton X-100 has a pH of 7.7, 8.2 or 8.3 (Fig. 8) (Note: reading on the pH ranges in the instant claims 1, 2, and/or 3). It is noted that Triton X-100 or Tween taught by Miyawaki et al. reads on the “non-ionic surfactant” in the instant claim 1 as well as the further defined non-ionic surfactant in the instant claim 5, as evidenced by the specification of the instant application (see paragraph 0035 spanning pages 12 and 13). Thus, the method of Miyawaki et al. reads on the method of the instant claims 1-3 and 5.
Regarding the claim 6, the examples of Miyawaki et al. (para 0187) teach the tissue is incubated in the decolorizing solution (ScaleS4) for 12 hours.
Regarding the claim 7, the examples of Miyawaki et al. further teach the tissue is incubated in the decolorizing solution (ScaleS4) for 189 days (see para 0229, lines 7-11 from bottom), reading on the time range in the claim 7.
Regarding the claim 8, the examples of Miyawaki et al. teach the tissue is incubated at a temperature of 37oC (see para 0188, lines 1-2), reading on the temperature range in the claim 8.
Regarding the claim 19, the examples of Miyawaki et al. (para 0183) teach the biological tissue is a brain tissue, which reads on the “internal organ tissue” recited in the claim.
Therefore, in view of the teachings of Miyawaki et al., the method of Claims 1-3, 5-8 and 19 is anticipated by the method of Miyawaki et al.
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 of this title, 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.
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 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.
Claims 1, 2, 5, 8-14, and 17-19 are rejected under 35 U.S.C. 103 as being unpatentable over Ryeon et al. (KR 20010020628 A, 2001, Machine-translated English version is of record), as evidenced by the Wikipedia printout of Phosphate-buffered saline (downloaded from URL of: https://en.wikipedia.org/wiki/Phosphate-buffered_saline on 8/21/2026), and by the Wikipedia printout of Hydrogen peroxide (downloaded from URL of: https://en.wikipedia.org/wiki/Hydrogen_peroxide on 8/21/2026).
Regarding the instant claims 1, 5, 8-9, 11-13, and 17-18, Ryeon et al. teach a method of decolorizing a blood specimen, comprising a step of incubating the blood specimen with a decolorizing solution, wherein the solution comprises H2O2 (hydrogen peroxide) and NaN3 (an enzyme inhibitor for catalase), and further comprises a surfactant, specifically Tween 80 for a sufficient time to decolonize the biological tissue (Abstract; page 7/paras 3 and 4/lines 1-2 and 5-7; page 8/para 3; and page 6/paras 3-4), wherein the hydrogen peroxide is at a concentration of 1.5% and the incubation step is performed at room temperature (abstract/lines 5 and 7) (Note: it reads on the temperature range 4-55oC in the instant claim 8); and wherein the decolorizing solution comprises PBS buffer (abstract/line 5) (Note: it reads on the buffer recited in the instant claims 17-18). Examiner notes that the blood specimen taught by Ryeon et al. comprises blood cells which reads on the “biological tissue” recited in the instant claim 1, as evidenced by the specification of the instant application (see page 17/para 0045/line 4 from bottom). It is further noted that Tween 80 of Ryeon et al. reads on the “non-ionic surfactant” in the instant claim 1 as well as the further defined non-ionic surfactant in the instant claim 5, as evidenced by the specification of the instant application (see paragraph 0035 spanning pages 12 and 13).
Regarding the limitation of “at a pH of at least 7” in the claim 1 and the further limitation “above 7” in the claim 2, Ryeon et al. teach that the decolorizing solution comprises a PBS buffer and are silent about a specific pH of the solution. However, it would have been obvious to incubate the biological tissue in the PBS buffer-controlled decolorizing solution at a pH value above 7 in the method of Ryeon et al. for decolorizing the biological tissue, because it is well known in the art that PBS has a buffered pH range at a level of above 7, as evidenced by the Wikipedia printout of Phosphate-buffered saline, which teaches that Phosphate-buffered saline (PBS) is a buffer solution having a pH of ~ 7.4 and is commonly used in biological research (see page 1: para 1/lines 1-2). Thus, the teachings of Ryeon et al. render the claims 1-2, 5, 8-9, 11-13, and 17-18 to be obvious.
Regarding the claim 10, Ryeon et al. teach an amount of hydrogen peroxide is comprised in the solution at 1.5%. Ryeon et al. are silent about whether hydrogen peroxide (in a liquid form) is comprised in the solution at a level of 1.5 wt% or at a level of 1.5 vol%. However, even assuming that the 1.5% taught by Ryeon et al. is a volume percentage, the amount of Ryeon et al. still reads on the claimed range for the follow reasons. Specifically, the density of pure hydrogen peroxide is 1.45 g/cm3, i.e. 1.45 g/ml, as evidenced by the Wikipedia printout of hydrogen peroxide (see right-bottom corner of page 2). Given that water has a density of 1.0 g/ml, a volume percentage of 1.5% is equivalent to a weight percentage of 2.16% [1.5x1.45/(1.5x1.45 + 1x98.5) = 2.175/100.675 = 2.16 wt%], which reads on the claimed range of 0.2% - 7%.
Regarding the claim 19, the term “internal organ tissue” is not defined in the specification. In view of that all internal organs contain blood/blood cells, the blood taught by Ryeon et al. can be considered as an internal organ tissue, thus meeting the claimed limitation.
Regarding the claim 14, Ryeon et al. further teach that the catalase inhibitor, e.g. sodium azide is included at an amount in the range from 0.0001 to 0.1%, which encompasses the range recited in the claim, thus renders the claimed range to be obvious. See MPEP 2144.05, which states “in the case where the claimed ranges “overlap or lie inside ranges disclosed by the prior art” a prima facie case of obviousness exists.”
Therefore, the invention as a whole would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention.
Claims 1-8 and 15-19 are rejected under 35 U.S.C. 103 as being unpatentable over Miyawaki et al. (US 2017/0370810, 2017, cited in IDS).
The teachings of Miyawaki et al. are described above.
Regarding the claim 4, Miyawaki et al. further teach that the decolorizing solution is preferably in a pH range of 6.0 to 9.0 (para 0108, line 7). It is noted that the pH 9.0 taught by Miyawaki et al. touches the low end of the claimed range of 9-11, thus rendering the claimed range to be obvious. See MPEP 2144.05.
Regarding the claims 15 and 16, Miyawaki et al. further teach adding DMSO to the clearing reagent (i.e. decolorizing solution) at an amount of 0.5% to 35% for benefits such as (i) improving permeability of the clearing reagent into a biological material and (ii) facilitating a clearing treatment over a tissue having a keratin surface (para 0106). In addition, Examples of Miyawaki et al. teach that a specific decolorizing solution (ScaleSS20) comprises 20% DMSO and 0.2% Triton X-100 and another specific decolorizing solution (ScaleSS40) comprises 17.78% DMSO and 0.18% Triton X-100 (para 0229/lines 1-7). Thus, the teachings of Miyawaki et al. render the limitations based on DMSO and its amount of 10-30% in the claims to be obvious.
Regarding the claims 17 and 18, Miyawaki et al. teach including a pH adjusting agent in the decolorizing solution (para 0102/line 6) and they do not expressively teach that the decolorizing solution in the Examples comprises a PBS buffer or Tris buffer.
However, Miyawaki et al. teach using a PBS buffer or Tris buffer to maintain pH at a level of above 7. See para 0232/lines 9-10 and last 2 lines: PBS for pH 7.5 to 8.0; and para 0158/lines 3-5 from bottom: buffered by tris hydrochloride and PBS buffered by phosphate. It would have been obvious to include a PBS or Tris buffer in the decolorizing solution in the method of Miyawaki et al. for decolorizing the biological tissue, because Miyawaki et al. expressively teach maintaining the pH of their decolorizing solution at a level about 7, as described above. Furthermore, it is a common practice in the art to add a PBS or Tris buffer for maintaining pH at a level above 7, as supported by Miyawaki et al.
Therefore, the invention as a whole would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention.
Claims 1-5, 8-14, and 17-19 are rejected under 35 U.S.C. 103 as being unpatentable over Ryeon et al. (KR 20010020628 A, 2001, Machine-translated English version is of record), as applied to Claims 1, 2, 5, 8-14, and 17-19, further in view of
Miyawaki et al. (US 2017/0370810, 2017, cited in IDS), as evidenced by Wikipedia printout of Phosphate-buffered saline (downloaded from URL of: https://en.wikipedia.org/wiki/Phosphate-buffered saline on 8/21/2026), and by Wikipedia printout of Hydrogen peroxide (downloaded from URL of: https://en.wikipedia.org/wiki/Hydrogen_peroxide on 8/21/2026).
The teachings of Ryeon et al. and Miyawaki et al. are described above
Regarding the claims 3 and 4, Ryeon et al. are silent about the pH of the decolorizing solution.
It would have been obvious to maintain the pH of the decolorizing solution at a level of 8 or 9 in the method of Ryeon et al. for decolorizing the biological tissue, thus arriving at the claimed method. This is because it is well known in the art that decolorization of a biological tissue in the presence of a nonionic surfactant (e.g. Tween) can be effectively carried out at a pH in the range of 8-9, as supported by Miyawaki et al.
Therefore, the invention as a whole would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention.
Claims 1-8 and 15-20 are rejected under 35 U.S.C. 103 as being unpatentable over Miyawaki et al. (US 2017/0370810, 2017, cited in IDS), as applied to Claims 1-8 and 15-19, further in view of Gradinaru et al. (US 2017/0199104, 2017, cited in IDS).
The teachings of Miyawaki et al. are described above.
Regarding the claim 20, Miyawaki et al. do not expressively teach the biological tissue is a heart tissue. However, Miyawaki et al. teach their method is appliable to an organ tissue of an animal or a multicellular organism, or ultra-deep tissues (paras 0112, and 0083: left col/line 2 from bottom), whose scope encompass a heart tissue. It would have been obvious to apply the method of Miyawaki et al. for decolorizing a heart tissue, because it is well known in the art that a decolonization solution comprising a non-ionic surfactant Triton X-100 is effective at decolorizing an internal organ tissue such as a heart tissue. In support, Gradinaru et al. teach a method for decolorizing a biological tissue, comprising: a step of incubating the biological tissue with a decolorizing solution comprising surfactant comprising a non-ionic surfactant Triton X-100 for a sufficient time to decolonize the biological tissue, wherein the biological tissue is heart, muscles, lungs, kidneys and like (claims 1 and 5, tables, and para 0033).
Therefore, the invention as a whole would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention.
Claims 1-20 are rejected under 35 U.S.C. 103 as being unpatentable over Miyawaki et al. (US 2017/0370810, 2017, cited in IDS) in view of Gradinaru et al. (US 2017/0199104, 2017, cited in IDS), as applied to Claims 1-8 and 15-20, further in view of Ryeon et al. (KR 20010020628 A, 2001, Machine-translated English version is of record) and Yu et al. (IScience 24: 1-21, 102178, published on Mar. 19, 2021, cited in IDS).
The teachings of Miyawaki et al. and Gradinaru et al. are described above.
Regarding the claims 9-14, Miyawaki et al. do not teach that the decolorizing solution comprises hydrogen peroxide and a catalase inhibitor sodium azide.
Yu et al. teach that tissue optical clearing technique provides an innovative approach for performing deep-tissue imaging by making tissue more transparent and reducing attenuation of light by using various optical cleaning reagents (abstract, page 1/last full para/lines 1-3). Yu et al. further teach optical cleaning reagents involved in tissue optical clearing: Triton X-100 for dissolving lipids of biological tissue thus to increase permeability (page 8/para 2/lines 2-3); DMSO for dehydration of biological tissue, thus reducing scattering of the tissue (page 10/last para – page 11/para 1); and hydrogen peroxide for decolorization of biological tissue through bleaching heme and melanin pigments in the tissue (page 14/last para – page 15/para 5, table 5).
The teachings of Ryeon et al. are described above. Ryeon et al. further teach that hydrogen peroxide is decomposed into water and oxygen by catalase in the biological tissue; and further adding an enzyme inhibitor to inhibit catalase and suppress the decomposition of hydrogen peroxide and the generation of oxygen, wherein sodium azide is used as the enzyme inhibitor for effectively inhibiting the catalase enzyme (page 6, paras 3-5).
It would have been obvious to modify the method suggested by Miyawaki et al. and Gradinaru et al. by including hydrogen peroxide and sodium azide in the decolorizing solution for improving optical clearing of the biological tissue, as taught by Yu et al. and Ryeon et al. A person of ordinary skill in the art would have been motivated to do so, because hydrogen peroxide bleaches pigments in the biological tissue, thus enhancing the transparency of the tissue, and it is well known in that art that hydrogen peroxide improves the tissue clearing, as supported by Yu et al. and Ryeon et al. Furthermore, it is well known in the art that sodium azide in the decolorizing solution acts as a catalase inhibitor, which would inhibit the catalase enzyme, thus protecting the action of hydrogen peroxide, as supported by Ryeon et al. One of ordinary skill in the art has a reasonable expectation of success at modifying the method suggested by Miyawaki et al. and Gradinaru et al., because it is expected that hydrogen peroxide further contributes to the clearing effect of Triton X-100 and DMSO, as supported by Yu et al.
Regarding the additional limitations about the amounts of hydrogen peroxide and sodium azide in the claims 10 and 14, these amounts are well known in the art, as supported by Ryeon et al. Thus, the claims would have been obvious over the combined teachings of the cited prior art.
Therefore, the invention as a whole would have been prima facie obvious to a person of ordinary skill in the art before the effective filing date of the claimed invention.
Double Patenting
The nonstatutory double patenting rejection is based on a judicially created doctrine grounded in public policy (a policy reflected in the statute) so as to prevent the unjustified or improper timewise extension of the “right to exclude” granted by a patent and to prevent possible harassment by multiple assignees. A nonstatutory obviousness-type double patenting rejection is appropriate where the conflicting claims are not identical, but at least one examined application claim is not patentably distinct from the reference claim(s) because the examined application claim is either anticipated by, or would have been obvious over, the reference claim(s). See, e.g., In re Berg, 140 F.3d 1428, 46 USPQ2d 1226 (Fed. Cir. 1998); In re Goodman, 11 F.3d 1046, 29 USPQ2d 2010 (Fed. Cir. 1993); In re Longi, 759 F.2d 887, 225 USPQ 645 (Fed. Cir. 1985); In re Van Ornum, 686 F.2d 937, 214 USPQ 761 (CCPA 1982); In re Vogel, 422 F.2d 438, 164 USPQ 619 (CCPA 1970); and In re Thorington, 418 F.2d 528, 163 USPQ 644 (CCPA 1969).
A timely filed terminal disclaimer in compliance with 37 CFR 1.321(c) or 1.321(d) may be used to overcome an actual or provisional rejection based on a nonstatutory double patenting ground provided the conflicting application or patent either is shown to be commonly owned with this application, or claims an invention made as a result of activities undertaken within the scope of a joint research agreement. Effective January 1, 1994, a registered attorney or agent of record may sign a terminal disclaimer. A terminal disclaimer signed by the assignee must fully comply with 37 CFR 3.73(b).
Claims 1-20 are provisionally rejected on the ground of nonstatutory obviousness-type double patenting as being unpatentable over claims 1-3, 5, 7-12, and 23-24 of copending Application No. 18/643510 in view of Miyawaki et al. (US 2017/0370810, 2017, cited in IDS) and Ryeon et al. (KR 20010020628 A, 2001, Machine-translated English version is of record).
The claims 1-3, 5, 7-12, and 23-24 of copending ‘510 Application are directed in part to a method for rendering biological tissue optically transparent for 3D light microscopy imaging, comprising: a step of pretreating a biological tissue with a blood decolorizing solution to make the biological tissue amenable for optical clearing, before the biological tissue is contacted with an optical clearing solution; wherein the decolorizing solution comprises an aqueous solution of hydrogen peroxide, a catalase inhibitor to decrease the ability of catalase in the blood to convert the hydrogen peroxide to water and oxygen; and the decolorizing solution comprises sodium azide and DMSO dimethyl sulfoxide (DMSO); wherein the decolorizing solution comprises an aqueous solution of hydrogen peroxide, sodium azide, and DMSO; and wherein said biological tissue is an internal organ tissue, specifically is heart tissue.
The claimed method of the ‘510 Application differs from the instantly claimed method in that The method of the ‘510 Application does not teach the decolorizing solution comprises a non-ionic surfactant (e.g. Triton X-100 or Tween).
The teachings of Miyawaki et al. and Ryeon et al. are described above.
It would have been obvious to further include a non-ionic surfactant (e.g. Triton X-100 or Tween) in the decolorizing solution in the method of the ‘510 Application for optically clearing the biological tissue because it is well known in the art that a non-ionic surfactant such as Triton X-100 or Tween is a clearing reagent that facilitates decolorization of biological tissue, as supported by Miyawaki et al. and Ryeon et al.
Regarding the additional limitations about pH range, temperature range, incubation time range, and amounts of hydrogen peroxide, DMSO, and sodium azide in Claims 1-4, 6-8, 10, 14, and 16, these are the features of decolorizing biological tissues well known in the art, as supported by Miyawaki et al. and Ryeon et al. Thus, the claims would have been obvious over the combined teachings of the cited prior art and the claims of the ‘510 Application.
Therefore, in view of the cited prior art, the method of Claims 1-20 of the instant application is deemed obvious over the method of claims 1-3, 5, 7-12, and 23-24 of copending Application No. 18/643510.
This is a provisional obviousness-type double patenting rejection because the conflicting claims have not in fact been patented.
Conclusion
No claim is in condition for allowance.
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Any inquiry concerning this communication or earlier communications from the examiner should be directed to Qing Xu, Ph.D., whose telephone number is (571) 272-3076. The examiner can normally be reached on Monday-Friday from 9:30 AM to 5:00 PM. If attempts to reach the examiner by telephone are unsuccessful, the examiner’s supervisor, Manjunath N. Rao, can be reached at (571) 272-0939. Any inquiry of a general nature or relating to the status of this application or proceeding should be directed to the receptionist whose telephone number is (571) 272-1600.
/Qing Xu/
Patent Examiner
Art Unit 1656
Prosecution Insights