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 .
The Examiner assigned to this case has changed. Please note the Examiner' s contact information at the close of this Office action.
Election of Species
After further consideration, the requirement for election of species mailed on 8/27/2025 has been withdrawn, and a new Election of Species is set forth herein.
The requirement for election between the species of the inventions previously set forth as Species "1", "2", and "3", drawn respectively to different concentrations of the extract, different chemicals components, and functional groups present in the extract is herein withdrawn. After reviewing the Applicant’s response, the disclosure, and the relevant art, these species (the concentrations, chemical constituents of the propolis extract, and functional groups thereof) are found to be obvious variants, based on the current record.
This application contains claims directed to the following patentably distinct species:
i (previously Species 4). The distinct species of the liquid comprising: a solvent selected from the group consisting of water, alcohol, glycol, glycerol, glycerin, glyceryl ether, diglycerol, and vegetable oil, as in claim 7; a mixture of propylene glycol and vegetable glycerol, as recited in Claim 8; and a mixture of propylene glycol and water, as recited in claim 10.
ii (previously Species 5). The distinct species of the one or more additives recited in claims 11-17, including the distinct species of fixative agents (as in claim 12).
iii (previously Species 6). The distinct species of the stain, further applied to the fixed tissue sample, from among those recited in claim 20.
The species are independent or distinct because the species of the invention set forth above comprise distinct fixative compositions, having different additives or different solvents. The use of different solvents for the extract would predictably result in different chemical properties and distinct functional results. Further, the addition of the different additives, such as one or more of the fixative agents, salts, buffering agents, crosslinking agent, or emulsifying agents would result in mutually exclusive compositions having distinct properties. It is not clear from the present record that the designated species of the inventions are interchangeable. In regards to the selection of different stains, each species of the stain amounts to distinct staining methods requiring different and mutually exclusive techniques and distinct outcomes. In addition, these species are not obvious variants of each other based on the current record.
Applicant is required under 35 U.S.C. 121 to elect a single disclosed species, or a single grouping of patentably indistinct species, for prosecution on the merits to which the claims shall be restricted if no generic claim is finally held to be allowable. Currently, claims 1-7, 9, and 11-20 are generic.
There is a serious search and/or examination burden for the patentably distinct species as set forth above because at least the following reasons apply:
the species of the solvents and additives have acquired a separate status in their art due to their divergent subject matter (e.g. different chemical and functional properties);
the species of the invention (the distinct solvents, additives, and stains) require a different field of search (e.g., searching different electronic resources, or employing different search strategies or search queries); and
the species of the invention (the distinct solvents, additives, and stains) are likely to raise different non-prior art issues under 35 U.S.C. § 101 and/or 35 U.S.C. § 112(a).
Applicant is advised that the reply to this requirement to be complete must include (i) an election of a species to be examined even though the requirement may be traversed (37 CFR 1.143) and (ii) identification of the claims encompassing the elected species or grouping of patentably indistinct species, including any claims subsequently added. An argument that a claim is allowable or that all claims are generic is considered nonresponsive unless accompanied by an election.
The election may be made with or without traverse. To preserve a right to petition, the election must be made with traverse. If the reply does not distinctly and specifically point out supposed errors in the election of species requirement, the election shall be treated as an election without traverse. Traversal must be presented at the time of election in order to be considered timely. Failure to timely traverse the requirement will result in the loss of right to petition under 37 CFR 1.144. If claims are added after the election, applicant must indicate which of these claims are readable on the elected species or grouping of patentably indistinct species.
Should applicant traverse on the ground that the species, or groupings of patentably indistinct species from which election is required, are not patentably distinct, applicant should submit evidence or identify such evidence now of record showing them to be obvious variants or clearly admit on the record that this is the case. In either instance, if the examiner finds one of the species unpatentable over the prior art, the evidence or admission may be used in a rejection under 35 U.S.C. 103 or pre-AIA 35 U.S.C. 103(a) of the other species.
Upon the allowance of a generic claim, applicant will be entitled to consideration of claims to additional species which depend from or otherwise require all the limitations of an allowable generic claim as provided by 37 CFR 1.141.
During a telephone conversation with Stefan U. Koschmieder, on 2/5/2026, a provisional election of species was made to prosecute the species of "a mixture of propylene glycol and vegetable glycerol" for the solvent (as recited in claims 7 and 8), "formalin" as the fixative agent provided as an additive (as in claims 11 and 12), and "a hematoxylin and eosin stain" for the additional staining method (as recited in claim 20).
Affirmation of this election must be made by applicant in replying to this Office action.
Claim 10 is withdrawn from further consideration by the examiner, see 37 CFR 1.142(b), as being drawn to a non-elected species of the invention.
Information Disclosure Statement
The information disclosure statement (IDS) submitted on 5/9/2023 is acknowledged. The submission is in compliance with the provisions of 37 CFR 1.97. Accordingly, the information disclosure statement is being considered by the examiner.
Specification
The disclosure is objected to because of the following informalities:
The application uses in many places - including the title, specification, and claims - the verb “fixating” to describe a process of performing fixation on tissues or cells (i.e. for fixation of a biological sample). This term is not used in the art for this process. The correct verb for biological fixation- a process through which tissues are preserved for staining, imaging, or evaluation- is either “performing fixation” or “fixing” (the past tense/participle is “fixed”). The term “fixate” or “fixating”, as recited herein, is a verb meaning “to direct one’s eyes toward” and is used to describe a view point, or the focusing of a lens/camera. Thus, the term “fixating” should be replaced with “fixing” (i.e. “A method of fixating a tissue sample” should instead say “A method of fixing a tissue sample”).
The title of the invention also contains the term “Fixating” which is not the proper term/verb for the techniques performed herein. As described above, the verb for this process of preserving tissues should be “fixing”. The following title is suggested: “Method of Fixing Tissue Samples Using a Composition Containing Propolis Extract”.
The specification recites in multiple locations, including page 3, line 23 and page 12, line 3, the phrase “are B vitamin”. This appears to be an obvious error for “a B vitamin”. It is noted that the phrase “one or more vitamins B” is used on page 17 to describe components of the propolis extract. From the disclosure, and the references incorporated therein (see page 12), it is clear that a propolis extract includes one or more B vitamins. See MPEP § 2163.07 regarding obvious errors or grammatical corrections in an application: “An amendment to correct an obvious error does not constitute new matter where one skilled in the art would not only recognize the existence of error in the specification, but also the appropriate correction. In re Oda, 443 F.2d 1200, 170 USPQ 268 (CCPA 1971).”
Appropriate correction is required.
Claim Objections
Claims 1-2, 5, 18 and 20 are objected to because of the following informalities:
As described above, the term “fixating”, in claim 1, line 1, should be replaced with “fixing” (“fixating a tissue sample” should recite “fixing a tissue sample”).
Similarly, the term “fixated” as used in claim 1, line 3, should be changed to “fixed”. This is the proper English language term for these processes and descriptions. The term also appears in claims 18 (line 1) and 20 (line 2).
Claims 1, 2, and 5 each recite the term “wt. %”, which includes an abbreviation and a period, which should not appear in the body of the claim. It is suggested, for improved clarity, to amend each use of “wt. %” to “percent weight”.
Appropriate correction is required.
Claim Rejections - 35 USC § 112(b)
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 applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention.
Claim 1 recites, in lines 7-10, the limitation “wherein the pre-treatment composition comprises a mixture of proteins, nucleic acids, carbohydrates, lipids, minerals, and vitamins; wherein the mixture of nucleic acids, carbohydrates, lipids, minerals, and vitamins in the pre-treatment composition is the same as that in the post-treatment composition”.
As an initial matter, there is insufficient antecedent basis for this limitation in the claim because “the mixture” of lines 9-10 is not the same as the mixture described in lines 7-8, because “proteins” is omitted. It is unclear whether the proteins of the mixture are purposefully omitted or if this is an error, however, the resulting claim language is indefinite, because one cannot determine accurately the meaning of the claim.
Further, there is no manner for one to determine whether this limitation has been fulfilled, because the phrase “is the same as that...” is a subjective phrase, and no specific assays or measurements have been set forth in the claims or the disclosure for which this sameness is to be determined. It is unclear whether the term “the same” in reference to only the identity of the materials that comprise the pre-treatment and post-treatment compositions or if this limitation also requires that the concentrations of each are the same. Is the “sameness” determined via extensive HPLC-Mass spec analysis, through lipidomics, metabolomics, genomics, et cetera? Further, it is unlikely that such analytical measurements can possibly be performed pre-treatment and post-treatment since the treatment itself should interfere with the accuracy of said methods.
Finally, there is likely extensive chemical modification of at least the proteins, lipids, carbohydrates, and amino acids due to the disclosed cross-linking ability of the propolis composition and the functional groups (esters, hydrocarbons, alcohols, and carboxylic acids) which comprise the propolis extract (see specification on page 16). There is no reason why such reactive chemicals, stated in the specification to crosslink proteins, would not also react with chemical groups on lipids, carbohydrates, and on nucleic acids. This is further evidenced by the examples and embodiments, which do not provide any measurement of the pre-treated composition nor does it teach any means by which one can determine what is present in the pre-treated composition. The phrase does not adequately set forth the metes and bounds of the claimed subject matter and thus, the claim is indefinite.
For the purposes of comparing the claimed invention to the prior art, this limitation has been interpreted under the B.R.I. of the specification as a desired result to be achieved, and afforded the appropriate patentable weight. A method providing essentially the same composition as so instantly claimed, comprising a propolis extract of 0.1 to 50 % w/w, will be deemed to fulfill this functional limitation of a desired result.
Claim 1 also recites the phrase “the cellular structural integrity of the tissue sample maintains after the treating”. This phrase is a relative and subjective phrase which renders the claim indefinite. The phrase “the cellular structural integrity of the tissue sample maintains” is not defined by the claim, the specification does not provide a standard for ascertaining the meaning of the phrase, and one of ordinary skill in the art would not be reasonably apprised of the scope of the invention. There is no objective manner for one to determine what the structure “maintains”. The phrase has been interpreted to mean either “the cellular structural of the tissue sample maintains its integrity after the treating” or “the cellular structural of the tissue sample is intact after the treating”. Correction is required.
Claim 3 recites, in line 3, the alternative substance of “are B vitamin”. The resulting language is indefinite as the meaning of “are B vitamin” cannot be determined. The claim has been interpreted to mean that the extract may further comprise a B vitamin (i.e. the same as “a C vitamin”). From the disclosure in the specification, it is evident that an extract of propolis may contain one or more B vitamins (see the description on page 12 and on page 17, lines 19-24).
Claim 18 recites the term “enhanced”, which is a relative term that renders the claim indefinite. The term “enhanced” 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. Claim 18 recites a “fixated tissue having enhanced preservation and fixation ability compared to a tissue sample treated with formalin”. This limitation renders the claim scope indefinite as there is no manner for one to determine the meaning of “enhanced preservation and fixation ability”. Further, the tissue cannot have “enhanced preservation and fixation ability” because the tissue is being preserved or fixed by the propolis-containing composition. These must be abilities or properties of the propolis-containing composition. Therefore the resulting claim is indefinite.
All other claims depend directly or indirectly from the rejected claims and are, therefore, also rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, for the reasons set forth above.
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.
Claims 1-4, 6-7, 9, 11-17, and 19 are rejected under 35 U.S.C. 103 as being unpatentable over Bottalico (FR-3073125-B1, on IDS filed 5/9/2023- citations are in reference to the English language translation provided by Applicant) in view of Hesselbart et al. (US PGPub No. 20070003591).
Bottalico teaches a composition for preserving tissues or organs - particularly embalming- which comprises, inter alia, an aqueous-alcoholic propolis extract (Abstract). Bottalico teaches that the composition comprises a solution of 30-60 wt %, preferably 40-50 wt %, aqueous alcoholic propolis extract (pg 2, under “Preferred Composition”; and claim 1).
Bottalico teaches that such alternative fixation solutions (i.e. for tissue embalming) are preferably to chemical fixatives containing strong concentrations of formaldehyde or glutaraldehyde, which are carcinogenic and there exists a need to identify and develop healthier and more ecological compositions (pg. 4, last paragraph – pg. 5 first paragraph).
Bottalico teaches that propolis is a substance obtained from beehives, comprising resin and waxes (pg. 5: ‘by “propolis", we mean a series of resinous, gummy and balsamic substances, of viscous consistency collected on certain parts (buds and bark mainly) of plants (certain trees mainly like willows, birches, poplars) by worker bees. which bring them back to the hive and modify them by the contribution of some of their secretions (wax and salivary secretions)”.
Bottalico teaches performing an extraction of the crude propolis using an ethanol/water extraction solvent to prepare a solution (pg. 5, last full paragraph; and see pg. 7: “The composition according to the invention is in liquid form”).
Regarding claim 7, Bottalico also teaches that the composition may further comprise penetrating agents which include ethylene glycol, propylene glycol, glycerol and glycerin (pg. 8, second paragraph). Bottalico further teaches adding a solubilizing agent such as Eumulgin® CO455 (BASF), which comprises a mixture of hydrogenated castor oil, water and propylene glycol (pg. 9, last paragraph).
Regarding claims 11 and 12, Bottalico teaches that the compositions of the invention can also be used in combination with formalin or formaldehyde, which is useful to considerably reduce the doses of formaldehyde/formalin used (paragraph spanning pgs. 12-13).
Regarding claims 11, 13, and 14, Bottalico teaches that the composition may also include surfactants including sodium lauryl sulfate, sodium dodecyl sulfate, esters of sucrose fatty acids, such as sucrose oleate, sorbitan esters, ethers of fatty alcohols and polyethylene glycol (PEG), such as PEG oleyl ethers, phospholipids, such as lecithin, and soaps (pg. 8, 7th paragraph).
Regarding claims 11, and 15-17, Bottalico teaches that the composition may additionally contain sodium citrate, heparin, sodium oxalate, EDTA, borates such as sodium borate, sodium tetraborate or pyroborate sodium, magnesium sulfate, sodium chloride, sodium sulfate and sodium phosphate. These reagents such as sodium chloride, sodium citrate, and sodium phosphate are well-known salts and/or buffering agents in the art.
However, Bottalico does not explicitly disclose that the propolis extract has two or more functional groups including an ester group, a hydrocarbon group, an alcohol group, and a carboxylic acid group, that chemically crosslink the proteins in the tissue composition.
Hesselbart et al. (US PGPub No. 20070003591), drawn to the use of propolis as a coating material for medical implants, teaches applications of propolis and describes the chemical constituents that provide propolis with its functional properties (Abstract; [0009]-[0028]).
Hesselbart teaches that propolis extract comprises resin and wax, and is recited from bees ([0009]). Hesselbart teaches that propolis includes: aliphatic acids and their esters ([0012]); carboxylic acids including long chain fatty acids behenic, palmitic, stearic, myristinic acid, as well as short-chain acids including succinic, angelic and butyric acid and their esters ([0013]); aromatic acids and their esters, including cinnamon acid, methyl salicylate, caffeic acid, vanillic acid and p-cumaric acid benzaldehyde ([0014]-[0015]); hydroquinone and cinnamon alcohols ([0016]-[0017]); aldehydes including vanillin, protocatechualdehyde(3,4-dihydroxybenzldehde), p-; hydroxybenzaldehyde ([0018]-[0019]).
Hesselbart also teaches that the propolis includes numerous hydrocarbons that consist of a complex mixture of n-alkanes which have an odd number of C atoms in the range including C27H56, C29H60, C31H64, and C33H68 ([0028]).
Therefore, to one of ordinary skill in the art, before the effective filing date of the claimed invention, it would have been obvious from the teachings of Bottalico to use a composition comprising a propolis extract for the fixation (i.e. preservation) of biological tissues with a reasonable expectation of success, and it would have been obvious, in view of the combined teachings of Bottalico and Hesselbart that the propolis extract comprises chemicals with one or more of an ester group, a hydrocarbon group, an alcohol group, and a carboxylic acid group, chemicals which chemically crosslink proteins in a tissue composition.
One of ordinary skill in the art would have been motivated from the teachings of Bottalico, teaching that a propolis extract is used for the embalming and preservation of tissues and whole bodies, to modify the methods for the preservation of tissue specimens.
The aqueous-alcoholic propolis extract of Bottalico appears to be substantially overlapping in composition with the alcohol extracted propolis composition of Hesselbart and that of the instant invention (pg. 11, last paragraph of the instant specification stating that ethanol is used for the extract and it dissolves the primary biologically active components of propolis). Therefore, although Bottalico fails to explicitly describe the chemical constituents of propolis and the associated functional properties, the fact that such chemicals and properties are present would have been evident to one of ordinary skill in the art prior to the instant invention (as shown at least in Hesselbart, published in 2007). Thus, the application of a propolis extract for fixing or preserving tissue, as taught in Bottalico would intrinsically use the properties of the propolis to achieve the desired result of crosslinking amino acid residues in the tissue proteins.
Known embalming fluids such as formaldehyde and formalin are used as fixatives (cross-linkers) for tissue specimens, and both the instant invention and that of Bottalico are applying the propolis extract as a substitute for formalin, which has toxic side-effects known to the art. See MPEP § 2143.I.C for the obviousness of using a known technique to improve similar devices, methods, or products in the same way “One of ordinary skill in the art would have been capable of applying this known method of enhancement to a "base" device (method, or product) in the prior art and the results would have been predictable to one of ordinary skill in the art. "It's enough … to show that there was a known problem … in the art, that [another reference] … helped address that issue, and that combining the teachings of [the two references] wasn't beyond the skill of an ordinary artisan. Nothing more is required to show a motivation to combine under KSR." See Intel Corp. v. PACT XPP Schweiz AG, 61 F.4th 1373, 1380-81, 2023 USPQ2d 297 (Fed. Cir. 2023)”. In the instant case, Bottalico teaches propolis extract compositions for preserving tissues, replacing formalin as a fixative, and thus using propolis extract for a fixative when seeking to fix tissue specimens (i.e. biopsies) would also have been prima facie obvious.
In regards to claim 2, Bottalico teaches that the composition comprises a solution of 30-60 wt % propolis extract, however, the determination of the effective concentration of a fixative would have been a matter of routine optimization and experimentation to one of ordinary skill in the art. One of ordinary skill would recognize that the concentration of a fixative should be regularly tested and optimized for any given tissue type. Thus, one would have reasonably arrived at the instantly claimed concentration, regardless of the preferred concentration taught in Bottalico. Further, there is no evidence in the specification that the concentration used in Bottalico would not be suitable for the instantly claimed methods (in fact it overlaps with the concentration of claim 1), nor is there evidence that the claimed concentration is critically important to the results of the fixation method. MPEP § 2144.05 describes that the determination of suitable or effective concentration of a known composition (or performing a known method) can be determined by one of ordinary skill in the art through the use of routine or manipulative experimentation to obtain optimal results, as these are variable parameters attainable within the art. Where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955).
In regards to claims 3 and 4, the compounds recited are known components of propolis extracts and derivatives thereof, from the teachings of Bottalico and Hesselbart cited above. Further, the providing of the propolis extract in Bottalico would necessarily include all of the natural occurring components found therein, regardless if they have been specifically identified or not (see MPEP § 2141.02). The instant specification does not describe that any the compounds of claim 4, are critical to the success of the invention. The specification also provides evidence itself that the compounds of claim 4 and functions thereof are inherently present in propolis extracts (see pg. 13, lines 4-14). In Ex parte Novitski, discussing the reference of “Dart”, it was noted that the Applicant’s disclosure appreciating the function upon usage thereof is further evidence of the presence of the function.
Regarding claim 6, as described above, the claimed crosslinking properties of the resin and wax of propolis amount to natural properties that are also applied with the propolis extract in Bottalico. The reference does not explicitly describe these properties; however, the result of fixing/preserving tissues is described in the cited art. One of ordinary skill with knowledge of the chemical constituents of propolis extract, as taught in Hesselbart, would predict that the application of the propolis containing composition of Bottalico results in the bonding and crosslinking of proteins via at least a first functional group (e.g. the hydrocarbons in the wax, as taught in Hesselbart) and one or more second functional groups, such as the esters and carboxylic acids found in the resin, said functional groups having known binding properties.
In regards to claims 7 and 9, Bottalico teaches that the composition can comprise ethylene glycol, propylene glycol, glycerol and glycerin, which are known solvents/vehicles in the art. The selection of these would have been obvious to one of ordinary skill in the art. Further, Bottalico specifically teaches that the composition used is a solution, and additionally, the production of emulsions, capsule suspensions, and dispersions are all obvious known methods of preparing a solution to one of ordinary skill in the art.
In regards to claims 11-17, it would have been obvious in view of the teachings of to use the propolis extract with additives, including formalin, which would have been useful to ensure the solution is fixed while considerably reducing the dose of formalin needed. Bottalico also teaches that the composition can include which include surfactants, fatty acid sulfates (sodium lauryl sulfate, sodium dodecyl sulfate), ethers of fatty alcohols, polyethylene glycol (PEG), salts such as sodium citrate, sodium borate, magnesium sulfate, sodium chloride, sodium sulfate, sodium phosphate, and/or EDTA. Thus the providing of these would have been reasonable to one of ordinary skill in the art, as these are known compounds with important chemical properties and one would have been motivated to select these for the properties they impart to the composition (i.e. surfactants add penetrating or emulsifying abilities, sodium phosphate has buffering properties, alkali metal halides such as sodium chloride affect the pH and solubility of the composition, EDTA chelates metal ions).
Regarding claim 19, Bottalico does not explicitly teach that the volume of tissue sample to the composition is in the range of 1:1 or 1:20, however Bottalico does teach that for preservation of bodies, the volume of the liquid composition used can be determined by the practitioner. One having ordinary skill in the art of tissue preservation would recognize that the fixative should be in a sufficient volume to cover the tissue, thus at least at 1:1 ratio by volume. Generally, in regard to the amounts, ratios, and volumes, it is noted that differences in experimental parameters such as volume and concentration of compounds in a solution will not support the patentability of subject matter encompassed by the prior art unless there is evidence indicating such parameter is critical. In the instant case, there are no teachings of the criticality of a specific concentration for this ratio in the instant disclosure. Where the general conditions of a claim are disclosed in the prior art, it is not inventive to discover the optimum or workable ranges by routine experimentation. In re Aller, 220 F.2d 454, 456, 105 USPQ 233, 235 (CCPA 1955). Thus, the instantly claimed volume ratios would be a matter of routine optimization and judicious selection, absence any evidence of their criticality.
From the teachings of the cited references, it is apparent that there would have been a reasonable expectation of success in combining the teachings therein to arrive at the claimed invention because Bottalico teaches using propolis extract as a tissue fixative or preservative, and Hesselbart discloses a number of compounds in the extract which will contribute to crosslinking.
Therefore, the invention as a whole would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date, as evidenced by the cited references, especially in the absence of evidence to the contrary.
Claims 1-7, 9, 11-17, and 19 are rejected under 35 U.S.C. 103 as being unpatentable over Bottalico (FR-3073125-B1) and Hesselbart et al. (US PGPub No. 20070003591) as applied to claims 1-4, 6-7, 9, 11-17, and 19 above, and further in view of Fetissova et al. (US PGPub No. 20070140990), to include the rejection of claim 5.
Bottalico and Hesselbart make obvious the methods for fixing tissues using a composition that comprises propolis extract, for all of the reasons described above.
However, the combination of Bottalico and Hesselbart does not teach that the propolis extract comprises resin at a concentration of 40 to 80 wt.%; one or more of the oil and wax at a concentration of 20 to 60 wt. %; and pollen at a concentration of 1 to 20 wt. %; each wt. % based on a total weight of the propolis extract, as in claim 5.
Fetissova et al. (US PGPub No. 20070140990 teaches an active oral compositions comprising a propolis extract, wherein the “propolis extract" or “an extract of propolis" encompass all suitable resin products produced by species and sub-species of the family Apidne, including bees of the genus Apis, and synthetic or semi-synthetic equivalents of such natural extracts or active components contained therein ([0019]). Fetissova teaches that on a weight basis propolis contains about 45 to about 55% resins and balsams (including for example, flavonoids, phenolic acids, and esters); about 25 to about 35% waxes and fatty acids; about 10% essential oils; about 5% pollen (including proteins and protein-derived amino acids); and about 5% of other organic compounds and minerals (including trace minerals, vitamins, ketones, lactones, quinones, steroids, benzoic acid and esters, and sugars) .
Thus to one of ordinary skill in the art, before the effective filing date of the claimed invention, it would have been obvious from the teachings of Bottalico and Hesselbart to perform a method using a propolis extract as claimed for the fixation (i.e. preservation) of biological tissues, and it would have been obvious, in view of the teachings of Fetissova that a natural propolis extract would intrinsically comprise resin at a concentration of 40 to 80 wt.% (e.g. about 45 to about 55% resins); one or more of the oil and wax at a concentration of 20 to 60 wt. % (e.g. about 25 to about 35% waxes and fatty acids); and pollen at a concentration of 1 to 20 wt. % (e.g. about 5% pollen).
One would have been motivated to provide a propolis extract having these claimed concentrations because they are taught in the art to be generally around the naturally occurring concentrations, and the prior art all pertains to applications of propolis extract, including Bottalico which teaches using this material for preserving/fixing biological tissues.
From the teachings of the applied references, it is apparent that there would have been a reasonable expectation of success in combining the teachings therein to arrive at the claimed invention because Bottalico teaches using propolis extract as a tissue fixative or preservative, whereas the teachings of Hesselbart and Fetissova render obvious the various components of the claimed compositions (derived from the same source) and concentrations thereof.
Therefore, the invention as a whole would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date, as evidenced by the cited references, especially in the absence of evidence to the contrary.
Claims 1-4, 6-9, 11-17, 19, and 20 are rejected under 35 U.S.C. 103 as being unpatentable over Bottalico (FR-3073125-B1) and Hesselbart et al. (US PGPub No. 20070003591) as applied to claims 1-4, 6-7, 9, 11-17, and 19 above, and further in view of Fan et al. (CN-108392462-A, a translation of which is provided with the accompanying PTO-892).
Bottalico and Hesselbart make obvious the methods for fixing tissues using a composition that comprises propolis extract, for all of the reasons described above.
However, the combination of Bottalico and Hesselbart does not teach that the liquid in the composition comprises propylene glycol to vegetable glycerol (i.e. glycerol that is derived from a plant), in a ratio (vol:vol) of 20:1 to 1:20.
Bottalico does teach that the composition may include propylene glycol, glycerol and/or glycerin (pg. 8, second paragraph). Bottalico further teaches adding a solubilizing agent such as Eumulgin® CO455 (BASF), which comprises a mixture of hydrogenated castor oil (oil from castor beans), water and propylene glycol (pg. 9, last paragraph).
Fan et al. (CN-108392462-A) teaches a composition that comprises resveratrol and propolis extract as main raw materials (Abstract, Title). Fan teaches that the solvent which can dissolve the composition including the propolis extract is a mixture of water, glycerol and “methyl propylene glycol” (i.e. propylene glycol which contains a methyl group) in a weight ratio of 1: 3.2: 2.0 (i.e. 1.6 to 1 falls which is within the 20:1 to 1:20 range claimed herein).
To one of ordinary skill in the art, before the effective filing date of the claimed invention, it would have been obvious to modify the method made obvious by the teachings of Bottalico and Hesselbart using a propolis extract for the fixation (i.e. preservation) of biological tissues such that the solvent (i.e. solubilizing agent) used in the composition includes both propylene glycol and glycerol at a ratio according to the instantly claimed amounts.
One would have been reasonably motivated to select propylene glycol and glycerol as part of the liquid composition because these are known solvents in the art, and they are explicitly taught for use together in Fan, with a composition comprising, inter alia, propolis extract. Further, selecting a glycerol that is derived from a plant or vegetable would be prima facie obvious, as there is no chemical difference in glycerol from plants compared to any other source of glycerol (i.e. all glycerol has the formula of C3H8O3 and share the same structure and properties). Thus, the selection of a solvent having both propylene glycol (as also used in Bottalico) and glycerol, including vegetable or plant-derived glycerol, would have been a matter of judicious selection to one of ordinary skill in the art, these solvents chosen from useful solvents known to the art. Further, the providing of these chemicals based on the ratios taught in Fan would have been prima facie obvious, and the arrival at any particular ratio would have been likewise obvious, as a matter of routine optimization to one of ordinary skill in the art. The instant application provides no evidence regarding the criticality of the selected ratio and solvent.
There would have been a reasonable expectation of success in combining the teachings therein to arrive at the claimed invention because Bottalico teaches using propolis extract as a tissue fixative or preservative and teaches including one or both of the claimed solvents (glycerol and propylene glycol) and Fan teaches a solvent for a propolis extract composition, wherein glycerol and propylene glycol are successfully used at a ratio of 3.2 : 2.0 or 1.6 : 1.
Therefore, the invention as a whole would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date, as evidenced by the cited references, especially in the absence of evidence to the contrary.
Claims 1-4, 6-9, 11-17, 19, and 20 are rejected under 35 U.S.C. 103 as being unpatentable over Bottalico (FR-3073125-B1) and Hesselbart et al. (US PGPub No. 20070003591) as applied to claims 1-4, 6-7, 9, 11-17, and 19 above, and further in view of Özkan, N. et al. ("Honey as a substitute for formalin?." Biotechnic & Histochemistry 87.2 (2012): 148-153), to include the rejection of claim 20.
Bottalico and Hesselbart make obvious the methods for fixing tissues using a composition that comprises propolis extract, for all of the reasons described above.
However, the combination of Bottalico and Hesselbart does not teach to perform a stain on tissue after treating with a propolis extract containing composition.
Özkan teaches that tissues fixed in either honey or alcoholic formalin and 10% neutral buffered formalin (NBF) have similar histomorphology, and teaches that honey can be used as a safe alternative to formalin in histopathology (Title, Abstract). Specifically, Özkan teaches using a pine honey for tissue fixation and compared it to other routine fixatives for histopathology. To examine the tissue after fixation, Özkan teaches performing hematoxylin and eosin (H & E) staining as well as a trichrome stain for muscle and collagen to assess tissue morphology (Fig. 1, Fig. 2, and pg. 149, Staining). Özkan also teaches performing immunohistochemical stains, techniques known in the art for visualizing specific structures and proteins (see Figs. 3 and 4).
To one of ordinary skill in the art, before the effective filing date of the claimed invention, it would have been obvious from the teachings of Bottalico and Hesselbart to perform a method using a propolis extract as claimed for the fixation (i.e. preservation) of biological tissues with a reasonable expectation of success, and it would have been obvious in light of the teachings of Özkan to perform one or more of a H&E stain or trichrome stain, for the purpose of assessing the tissue structure after fixation.
One would have been motivated to examine the fixed tissue using one or more stains known to the art, including those taught in Özkan. Although Özkan teaches using such stains to assess the performance of honey, a similar but distinct compound that that so instantly claimed, it would have been advantageous when performing the similar fixation method suggested by Bottalico using propolis extract to further stain the tissue in a manner known in the art. See also MPEP § 2143.I.C, describing that it is obvious to use a known technique to improve similar devices, methods, or products in the same way: “One of ordinary skill in the art would have been capable of applying this known method of enhancement to a "base" device (method, or product) in the prior art and the results would have been predictable to one of ordinary skill in the art. In the instant case, the staining techniques taught in Özkan would have provided the predictable advantage of being able to assess the structure and integrity of the fixed tissue.
From the teachings of the applied references, it is apparent that there would have been a reasonable expectation of success in combining the teachings therein to arrive at the claimed invention because Bottalico teaches using propolis extract as a tissue fixative or preservative, and Özkan teaches that tissue stains (H&E stains) are known in the art for assessing structures and the quality of fixed tissue, and can be successfully used with various distinct fixatives.
Therefore, the invention as a whole would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date, as evidenced by the cited references, especially in the absence of evidence to the contrary.
Citation of Pertinent Art
The prior art made of record and not relied upon is considered pertinent to applicant's disclosure.
Mavri et al. “Chemical properties and antioxidant and antimicrobial activities of Slovenian propolis.” Chemistry & biodiversity vol. 9,8 (2012): 1545-58. doi:10.1002/cbdv.201100337, pertains to the characterization of the chemical constituents of propolis, or bee glue, which includes phenolic acids and flavonoids such as caffeic acid, ferulic acid, and luteolin (Abstract, Title, Table 3).
Alwahaibi, Nasar et al. “Effectiveness of neutral honey as a tissue fixative in histopathology.” bioRxiv 2021.04.27.437988; doi: 10.1101/2021.04.27.437988 (preprint), teaches the testing and use of neutral buffered honey and other types of honey fixatives to fix histological tissues (Abstract, Table 2).
Conclusion
No claims are allowable.
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/A.T.M./Examiner, Art Unit 1655
/ANAND U DESAI/Supervisory Patent Examiner, Art Unit 1655