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 .
Continued Examination Under 37 CFR 1.114
A request for continued examination under 37 CFR 1.114, including the fee set forth in 37 CFR 1.17(e), was filed in this application after final rejection. Since this application is eligible for continued examination under 37 CFR 1.114, and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on August 4, 2025 has been entered.
DETAILED ACTION
This application is a US national phase of PCT/JP2020/033556, filed September 4, 2020, with foreign application JP2019-161824, filed September 5, 2019.
Applicant's amendments filed May 28, 2025 and August 4, 2025 have been acknowledged. Claims 3-5, 11, 13-22, and 26-27 are canceled and claims 35-36 are newly added, and claim 1 is amended. The previous objection and 112(b) rejection in the Final office action mailed February 28, 2025 are withdrawn due to Applicant’s amendment to the claims.
Claims 1-2, 6-10, 12, 23-25, and 28-36 are pending. Claims 7-10 and 32-34 remain withdrawn. Claims 1-2, 6, 12, 23-25, 28-31, and 35-36 are examined.
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS.—Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
The following is a quotation of pre-AIA 35 U.S.C. 112, fourth paragraph:
Subject to the following paragraph [i.e., the fifth paragraph of pre-AIA 35 U.S.C. 112], a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claim 2 is rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends.
Claim 2 recites the use of the substrate in independent claim 1, i.e. “for directly measuring tryptase activity in a blood sample”, however, this limitation does not further limit the claim structurally or functionally, only reciting its intended use, therefore is improper. Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
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, 2, 6, 12, 23-25, 28-31, 35 and 36 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 the limitation “fluorescence characteristics of color development characteristics" in the last two lines. There is insufficient antecedent basis for this limitation in the claim. Applicant may consider changing the word “of” to “or” in this phrase to obviate this rejection. Claims 2, 6, 12, 23-25, 28-31, 35 and 36 are rejected for depending from claim 1.
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, 2, and 23-24 are rejected under 35 U.S.C. 103 as being unpatentable over Af Ekenstam et al. (US 4,137,225, cited in PTO-892 mailed 10/21/2024, hereinafter “Af”) in view of Niles et al. (US 2002/0197661 A1, cited in PTO-892 mailed 10/21/2024, hereinafter “Niles”), Weissleder et al. (NATURE BIOTECHNOLOGY VOL 17 APRIL 1999, cited in PTO-892 mailed 2/28/2025, hereinafter “Weissleder”), and Choksakulnimitr et al. (Journal of Controlled Release 34 (1995) 233-241, cited in PTO-892 mailed 2/28/2025, hereinafter “Chok”).
Af teaches diagnostically active chromogenic substrate with high specificity to serine proteases having the formula: H-D-A1-A2-A3-NH-R or salts thereof, wherein A1 is Gly, Ala, Val, Leu, Ile, Pip, Pro, or Aze; A2 is Gly, Ala, Val, Leu, Ile, Pip, Pro, Aze, or Phe; A3 is Arg, Lys, or Orn, and R is a chromophoric group (abstract). Af teaches the substrates are suitable for quantitative determination of the serine proteases, which split in the peptide chain on the carboxyl side of arginine or lysine (col. 1, lines 6-11). Af teaches among the best substrates of this type are those having a benzoylated N-terminal end and a chromophoric group coupled to the C-terminal end (col. 1, lines 25-28). Af teaches tripeptide bonded dye label in the formula: Cbo-D-Ala-Ala-Arg(NO2)-pNA (col. 7, Table 1). Af teaches for the synthesis of the novel chromogenic enzyme substrates conventional protective groups and coupling methods are used, such as the α-amino protective group it is of advantage to use carbobenzoxy or t-butyloxy carbonyl or some group related thereto such as for instance p-metoxy, p-nitro or p-metoxyphenylazo-carbobenzoxy (col. 2, lines 17-24). Af teaches the carbobenzoxy protective group (Cbo-) is attached to the tripeptide N-terminally (col. 3, lines 57-59).
Similarly, Niles teaches synthetic polypeptide substrates for detecting tryptase enzymes and teaches the sample for measuring the tryptase activity can include whole blood, serum, inter alia (abstract, para 14). Niles also teaches an isolated polypeptide comprising in amino to carboxy order P4-P3-P2-P1, wherein P4 is Proline (“P”), P3 is Arginine (“R”) or Lysine (“K”), P2 is any amino acid, and P1 is K or R, and further wherein a fluorogenic leaving group comprising 7-amino-4-carbamoylmethyl- coumarin (ACC) is bound via an amide bond to P4-P3-P2-P1 at a carboxy terminus of P4-P3-P2-P1, wherein P4 is acetylated and amino terminally blocked (para 12, 14). Niles teaches mast cells express at least four distinct tryptase genes: α, βI, βII, and βIII, wherein βI and βII have very similar P4 to P1 substrate preferences (para 16). This shared preference for peptide substrates likely extends to a shared preference for physiological substrates, which the optimal sequence for β-tryptase, P4=P, P3=R or K, P2=any amino acid, and P1=K or R, was found in the macromolecular substrates taught by Niles (para 16). Niles teaches if the sample has any tryptase activity, such activity will produce a detectable fluorescent moiety, the fluorescence of the sample is then measured to determine whether it undergoes a detectable change in fluorescence, the detectable change being an indication of the activity of the enzymatically-active B-tryptase in the sample (para 14).
Neither Af nor Niles teach the tripeptide-X is N-terminally linked to a poly(L-lysine).
However, Weissleder teaches in vivo imaging of tumors with protease-activated near-infrared fluorescent probes (NIRF) (title). Weissleder teaches NIRF probes were bound to a long circulating graft copolymer consisting of poly-L-lysine and sterically protected by methoxypolyethylene glycol succinate (abstract, pg. 375, col. 2, para 2). Weissleder teaches the construction of synthetic graft copolymer consisting of poly-L-lysine (PL) which contained Cy5.5 fluorophore (pg. 375, col. 2, para 2). The probe was synthesized by covalently attaching methoxypolyethelene glycol succinate (5 kDa) to poly-L-lysine (35.5 kDa) resulting in PL-MPEG (pg. 377, col. 2, para 3). Weissleder teaches the compound was treated with trypsin to determine whether copolymer bound Cy could be released enzymatically (pg. 375, col. 2, para 3). Weissleder discloses internalization of the copolymer into tumor cells occurs by fluid-phase endocytosis, which is often upregulated in rapidly proliferating cells and have elevated proteolytic enzymes allowing the intracellular release of the NIRF probes (pg. 375, col. 1, para 2). Weissleder teaches that cysteine protease (cathepsin B, H, and L) inhibitors such as E64 and leupeptin completely inhibited NIRF generation; likewise, trypsin inhibitors (tosyl-l-lysyl chloromethyl ketone) and trypsin-like serine protease inhibitor (3,4-dichloroisocoumarin) also inhibited NIRF generation, demonstrating the NIRF signal generation was a result of lysosomal cysteine/serine protease activity. (pg. 376, col. 2, para 1).
Af, Niles, and Weissleder do not teach the poly(L-lysine) has a MW between 5000-15,000.
However, Chok teaches in vitro cytotoxicity of macromolecules in different cell culture systems (title). Chok teaches poly(L-lysine) hydrobromide with MW 8000 that have been assessed for their cytotoxicity on brain microvessel endothelial cells (BMECs) and Mɸ (macrophages) (pg. 234, sec. 2.1 & 2.4). Chok teaches severe LDH-release induced by poly-L-lysine (MW 39,800) in both cultured cells, and moderate LDH-release induced by poly-L-lysine MW 8000 in BMECs but only a little LDH-release in Mɸ, and that larger cationic macromolecules induce higher cytotoxicity (pg. 235, col. 2, para 4).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to modify the diagnostically cleavable substrate comprising the tripeptide formula Cbo-Ala-Ala-Arg(NO2)-pNA taught by Af with a poly (L-lysine) protective group as taught by Weissleder on the N-terminal side of the tripeptide, and utilize a low molecular weight poly (L-lysine) protective group that would confer less cytotoxicity as taught by Chok with a reasonable expectation of success. Furthermore, it would have been prima facie obvious to modify the substrate taught by Af, with a cleavable peptide sequence used to measure tryptase enzymatic activity in a blood sample, with the formula P4-Lys-P2-Arg and a fluorogenic leaving group attached to the C-terminal end of the peptide as taught by Niles. Niles teaches the P2 position can be any amino acid, thus it would have been obvious to try any amino acid at that position to arrive at the tripeptide formula recited in claims 1 and 23. Af and Niles are considered analogous art, and both teach serine proteases hydrolyze substrates at the C-terminal end of a peptide with an Arg-X linked to a fluorochrome at the P1 position. The Arg-X link is sensitive to tryptase cleavage as taught by Niles (para 10). Additionally, one of ordinary skill in the art would have been motivated to combine the tripeptide fluorochrome conjugate with a macromolecule, poly (L-lysine), which can advantageously serve as a delivery for an in vivo or in vitro diagnostic test as taught by Weissleder and Chok.
Claims 6, 12, and 28-31 are rejected under 35 U.S.C. 103 as being unpatentable over Af, Niles, Weissleder, and Chok as applied to claim 1 above, and further in view of Chi (PhD dissertation, 2014, Kyushu Institute of Technology).
Af, Niles, and Weissleder teach fluorochromes/chromophores fluorescing when enzymatically cleaved by serine proteases, but do not teach the specific fluorescent dye labels recited in claims 6, 12, and 28-31.
However, Chi teaches application of peptide 4-methylcoumaryl-7-amide (MCA) as a novel fluorescent substrate (title). Chi teaches fluorescent substrates are a good tool for measuring protease activity, and describes the development of MCA was synthesized as Boc-Val-Pro-Arg-MCA, Boc-Ile-Glu-Gly-Arg-MCA and Boc-Ser-Gly-Arg-MCA and were found to be specific substrates for individual enzymes, and is widely used commercially for enzymatic assay of cathepsin B (pg. 1, 8). Chi teaches a method for detecting histone methyltransferases (HMTs), wherein the design sequence of peptide-MCA as an abbreviation expressed as R-XK-D (R represent a hydrogen atom or a protecting group for an amino terminus; X represents a peptide consisting of zero or one or more amino acid residues; K represents a lysine residue; D represents one dye label linked via an amide bond to the carbonyl terminus of a lysine residue) (pg. 56). Chi teaches D should be one dye label linked via an amide bond to the carbonyl terminus of the K (lysine residue) and whose fluorescence property is changed by the cleavage of the amide bond can be selected as 4-methyl-coumaryl-7-amide (MCA), 7-amino-4-chloromethyl-coumarin (CMCA), 2-aminonaphthalene-6-sulfonic acid (ANS), 7-amino-4-trifluoro-methylcoumarin (FMCA), and 2-amino-7-methylpurine-6-thiol (AMP), which meets the limitations in claims 6, 12, and 28-31 (pg. 57).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing of the claimed invention to modify the substrate structures of Cbo-Ala-Ala-Arg(NO2)-pNA or P4-Lys-P2-Arg and a fluorogenic leaving group attached to the C-terminal end of the peptide with an N-terminally linked poly(L-lysine) having a MW between 5-15 kDa, as taught by Af and Niles as modified by Weissleder and Chok, by replacing the chlorophore pNA, with MCA, ANS, CMCA, or AMP on the C-terminal end of the tripeptide, as taught by Chi with a reasonable expectation of success. Af, Niles, and Chi are considered to be analogous art to the claimed invention because the references are in the same field of endeavor of constructing substrates with a C-terminal Arg linked to a fluorochrome. Therefore, it would have been obvious to someone of ordinary skill in the art to have a simple substitution with the MCA, ANS, CMCA, or AMP as disclosed by Chi, because these fluorochromes are known to fluoresce when cleaved by a serine protease and would have obtained the predictable result of detecting serine protease activity, providing an accurate and detailed measurement of serine protease activity (i.e. tryptase activity).
Claim 25 is rejected under 35 U.S.C. 103 as being unpatentable over Af, Niles, Weissleder, and Chok as applied to claim 1 above, and further in view of Poreba et al. (Amino Acids (2014) 46:931–943, cited in PTO-892 mailed 2/28/2025, hereinafter “Poreba”), as evidenced by Hashemi (Université de Montréal, PhD Dissertation, 2006, pgs. 1-257, cited in PTO-892 mailed 2/28/2025).
Af teaches the general formula of a chromogenic substrate as H-D-A1-A2-A3-NH-R, wherein A1 can be Ala, 2-azetidine carboxylic acid (Aze), inter alia; A2 can be Ala, inter alia; and A3 can be Arg, inter alia (col. 9, claim 1). As evidenced by Hashemi, Aze is closely related structurally to 2-aminobutyric acid (Abu), wherein Aze is derived from Abu (pg. 16, sec. 1.3.2.1).
Af, Weissleder, and Niles do not teach the tripeptide comprises 2-aminobutyric acid (Abu), specifically the sequence Abu-Ala-Arg-X.
However, Poreba teaches unnatural AA increase activity and specificity of synthetic substrates for human and malarial cathepsin C (title). Poreba teaches mammalian cathepsin C is primarily responsible for the removal of N-terminal dipeptides and activation of several serine proteases in inflammatory or immune cells (abstract). Poreba teaches optimal cathepsin C substrates as NH2-Abu-Nle(OBzl)-ACC, and found Abu as best recognized by both cathepsin C and malarial cathepsin C analog, when screening AA’s (pg. 936, col. 2, para 3). Poreba teaches analysis of the kinetic parameters for human cathepsin C in the P2 position demonstrates a high preference for small aliphatic side chains, wherein Abu in the substrate exhibited the highest enzyme efficiency for an unnatural AA derivative (pg. 939, col. 2, para 1).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to modify the substrate structure of H-D-Aze-Ala-Arg-NH-R taught by Af and replace the Aze with Abu taught by Poreba, at the A1 position, as well as link a poly (L-lysine) with a MW 5000-15000 taught by Weissleder and Chok on the N-terminal side of the tripeptide, for the measurement of tryptase activity taught by Niles. One of ordinary skill in the art would have been motivated to substitute the Aze with Abu that has small aliphatic side chains that would be recognized by cathepsin C which in turn would activate serine proteases (i.e. tryptase) as taught by Poreba, which would then cleave the fluorophore of the tripeptide for advantageous tryptase measurement activity.
Claims 35-36 are rejected under 35 U.S.C. 103 as being unpatentable over Af, Weissleder, Chok, and Niles as applied to claim 1 above, and further in view of Law et al. (Chemistry & Biology, 2004, Vol. 11, 99–106, hereinafter “Law”).
As discussed above, Af and Niles teach chromogenic substrates specific for serine proteases and tryptase, and Weissleder teaches substrate-mediated cleavage of NIRF probes from structurally constrained biocompatible precursors, such substrates comprising PL and MPEG. Weissleder teaches NIRF probes that are sterically protected by methoxypolyethylene glycol succinate (abstract, pg. 375, col. 2, para 2). Weissleder does not explicitly teach the MPEG is a linker of the tripeptide and PL as recited in the claims.
However, Law teaches design, synthesis, and characterization of urokinase plasminogen-activator(uPA)-sensitive near-infrared reporter, wherein Law uses multiple uPA substrate motifs attached to a pegylated graft copolymer consisting of MPEG and PL (title, pg. 99, col. 2, para 2; Fig. 1). Law teaches uPA is a possible predictor of tumor disease outcome, and it is feasible to design and synthesize uPA-specific NIRF imaging probes to determine uPA activity in vivo (pg. 104, col. 1).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing date of the claimed invention to modify the diagnostically cleavable substrate comprising the tripeptide formula Cbo-Ala-Ala-Arg(NO2)-pNA taught by Af with a PL protective group as taught by Weissleder on the N-terminal side of the tripeptide, utlizing a low molecular weight poly (L-lysine) protective group that would confer less cytotoxicity as taught by Chok, and link the PL to the peptide substrate via a linker comprising PEG as taught by Law with a reasonable expectation of success. Furthermore, one of ordinary skill in the art would have been motivated to combine the tripeptide fluorochrome conjugate with a macromolecule, poly (L-lysine), which can advantageously serve as a delivery for an in vivo or in vitro diagnostic test, such as the NIRF probes taught by Weissleder and Law.
Response to Arguments
Applicant's arguments filed August 4, 2025 have been fully considered but they are not persuasive.
Regarding remarks directed to the 103 rejection, Applicant argues there is no motivation to modify MW of poly-L-lysine as taught by Choksakulnimitr because he substrate of the present claim is for measuring tryptase activity in a sample, such as a blood sample, collected from subjects, as described throughout the present specification. Specifically, the sample has been separated from the subject and therefore, it is not necessary to consider cytotoxic effect of the substrate due to the size of the molecular weight of the substrate.
In response to applicant's argument that there would be no motivation to use a low molecular weight PL as taught by Chok because the claimed substrate is to be used for measuring tryptase activity in a blood sample, a recitation of the intended use of the claimed invention must result in a structural difference between the claimed invention and the prior art in order to patentably distinguish the claimed invention from the prior art. If the prior art structure is capable of performing the intended use, then it meets the claim. Furthermore, a blood sample is broad to include blood that is separated from the body and recirculated back, which would provide motivation to modify the substrate to be less cytotoxic.
Applicant also argues one of ordinary skill would not be motivated to choose a substrate having the amino acid sequence of Ala-Ala-Arg from among many substrates disclosed by Af. Similarly, Weissleder does not discriminate between tryptase and thrombin which are both belonging to serine proteases. Applicant argues With respect to Niles, it discloses that Ac-PRNK-ACC (SEQ. ID. NO: 12) is a specific substrate for tryptase in relation to other blood-borne serine proteases (paragraph [0108] and Fig. 5) and that the Ac-PRNK-CMK (SEQ. ID. NO: 14) inhibitor is largely ineffective against blood-borne serine proteases with proteolytic specificity divergent from tryptase, e.g., Factor Xa and thrombin (paragraph [0122], fig. 14). Therefore, one of ordinary skill in the art would not be motivated to modify the polypeptide (substrate) prepared by Niles.
In response to applicant’s argument that there is no teaching, suggestion, or motivation to combine the references, the examiner recognizes that obviousness may be established by combining or modifying the teachings of the prior art to produce the claimed invention where there is some teaching, suggestion, or motivation to do so found either in the references themselves or in the knowledge generally available to one of ordinary skill in the art. See In re Fine, 837 F.2d 1071, 5 USPQ2d 1596 (Fed. Cir. 1988), In re Jones, 958 F.2d 347, 21 USPQ2d 1941 (Fed. Cir. 1992), and KSR International Co. v. Teleflex, Inc., 550 U.S. 398, 82 USPQ2d 1385 (2007). In this case, Af teaches diagnostically active chromogenic substrates, specifically formula (2) in instant claim 1. Weissleder teaches the copolymer poly (L-lysine) as an effective delivery agent for NIRF probes that are protease activated, which is the same idea as the claimed invention, albeit without the specific tripeptide. Niles teaches the specific tripeptide formula 1 of claim 1, and in fact does show Ala in P2 has higher activity than other amino acids such as Gly (see Fig. 3b & 4b). Thus, one of ordinary skill in the art would be motivated to construct a diagnostic substrate for tryptase activity utilizing the teachings in Af, Niles, and Weissleder (and newly applied Law), therefore is obvious.
Pertinent Prior Art
Relevant to withdrawn claims 32-34, Beija teaches rhodamine dyes are widely used as fluorescent probes owing to their high absorption coefficient and broad fluorescence in the visible region of electromagnetic spectrum, high fluorescence quantum yield and photostability (abstract). Beija teaches several groups have made an effort to design new rhodamine derivatives to be used as latent fluorophores in studies of enzymatic activity (serine protease, caspase, esterase, DT diaphorase), of organometallic catalysis in living cells, in biomedical imaging or in in vivo detection of small molecules (thiols) (pg. 2412, col. 1, para 4). Beija teaches Rho 110 was modified in order to prepare synthetic fluorogenic amide substrates for assays of serine proteases and caspases, which meets the limitation of claims 32-33 (pg. 2412, col. 2, para 2). Beija teaches Rhodamine 101 (Rho 101) and Rhodamine B (Rho B) are among the most used rhodamines, and Rho B and Rho 6G are the less expensive dyes of this family and consequently they have been the most employed for further applications, which meets the limitation in claim 34 (pg. 2412, col. 1, para 3). Beija discloses that, due to these late developments on the synthetic methods for derivatization of rhodamine dyes, it is today possible to envisage attaching this dye to almost every molecule of interest, taking advantage of their outstanding photophysical properties (pg. 2432, col. 1, para 2).
Therefore, it would have been prima facie obvious to one of ordinary skill in the art prior to the effective filing of the claimed invention to modify the substrate structures of Cbo-Ala-Ala-Arg(NO2)-pNA or P4-Lys-P2-Arg and a fluorogenic leaving group attached to the C-terminal end of the peptide with an N-terminally linked poly(L-lysine) having a MW between 5-15 kDa, as taught by Af and Niles as modified by Weissleder and Chok, by replacing the fluorogenic leaving group (e.g. pNA), with rhodamine 110 or rho B fluorescent dyes on the C-terminal end of the tripeptide, as taught by Beija with a reasonable expectation of success. Beija discloses the broad applicability of rhodamine derivatives, and their use in serine protease substrates. Therefore, it would have been obvious to someone of ordinary skill in the art to have a simple substitution with the rhodamine 110 or Rho B as disclosed by Beija, because these fluorochromes are known to fluoresce when cleaved by a serine protease and would have obtained the predictable result of detecting serine protease activity, providing an accurate and detailed measurement of serine protease activity (i.e. tryptase activity).
Conclusion
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/LOUISE W HUMPHREY/Supervisory Patent Examiner, Art Unit 1657
/JESSICA EDWARDS/
Examiner, Art Unit 1657