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 .
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and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114.
Remarks
The claims and remarks filed on 02/27/2026 have been entered and considered. The text of those sections of Title 35, U.S. Code not included in this action can be found in a prior Office action. The rejections and/or objections presented herein are the only rejections and/or objections currently outstanding. Any previously presented objections or rejections that are not presented in this Office Action are withdrawn.
Claims 1-2, 4-7, 9, 11-15, 17-19, 23-24, 30, 49, and 51-52 are pending.
Claims 3, 8, 10, 16, 20-22, 25-29, 31-48, and 50 are canceled.
Claims 9 and 49 are withdrawn.
Claims 1-2, 4-7, 11-15, 17-19, 23-24, 30, and 51-52 have been examined on the merits.
Priority
This application, U.S. Application number 18/043568, is a national stage entry of International Application Number PCT/US21/48526 filed on 08/31/2021, which claims priority under 35 U.S.C. 119(e) from U.S. Provisional Applications of No. 63218103 filed on 07/02/2021, No. 63140592 filed on 01/22/2021, No. 63119451 filed on 11/30/2020, and No. 63072656 filed on 08/31/2020.
Information Disclosure Statement
The information disclosure statement (IDS) submitted on 02/27/2026 is acknowledged. The submission is in compliance with the provisions of 37 CFR 1.97., and has been considered by the examiner.
Rejections - Withdrawn
The rejection of Claims 1, 2, 5-6, 11-15, 17-19, 23-24, 30, and 51-52 under 35 U.S.C. 103 over Jahn et al. in view of Al-Kady et al. is withdrawn in favor of the rejections listed below.
The rejection of Claims 4 and 7 under 35 U.S.C. 103 over Jahn et al. in view of Al-Kady et al., Zhang et al., and Nalder et al. is withdrawn in favor of the rejections listed below.
Claim Rejections - 35 USC § 103
Claims 1, 5-6, 17, 19, 30, and 52 are rejected under 35 U.S.C. 103 as being unpatentable over Jacks et al. (Analytical Biochemistry, 1967, 21:279-285, of record), as evidenced by the Wikipedia printout of Caprylic Acid (retrieved on 11/18/2025 from URL of https://en.wikipedia.org/wiki/Caprylic_acid), and by Jahn et al. (Pharm. Res., 37(6): 118, pages 1-13, published on June 3, 2020, cited in IDS).
Jacks et al. teach a method/assay for determining lipolytic activity of lipase in a sample, which is based on hydrolysis of fatty acyl esters of 4-methylumbelliferone (as a fluorogenic substrate of lipase) to yield a fluorescent product, i.e. free 4-methylumbelliferone (free 4MU) (title, page 279/para 2), which is the same fluorescent product as the “4- methylumbelliferone (MU)” recited in step (b) of instant claim 1, wherein the method comprises steps: (a) forming a reaction mixture by adding a sample containing a lipase, a reaction buffer, and 0.6 mmol of the fluorogenic substrate (i.e. a fatty acyl ester of 4-methylumbelliferone); (b) monitoring the conversion of the fluorogenic substrate from a non-fluorescent state to the fluorescent product 4MU to measure a fluorescent signal of the reaction mixture; and (c) quantifying the enzymatic activity of the ligase based on the conversion of the fluorogenic substrate (the para spanning pages 280 and 281, page 281/paras 3 and 6, page 283/second para, page 284/last 2 paras, Tables 2-3, Fig. 2); wherein seven fatty acyl esters of 4MU having different carbon chain lengths of fatty acyl are respectively applied as the fluorogenic substrate, specifically these fatty acyl esters are esters of butyryl (MU-C4, 4-carbon length), hexanoyl (MU-C6, 6-carbon length), heptanoyl (MU-C7, 7-carbon length), octanoyl (MU-C8, 8 carbon length), nonanoyl (MU-C9, 9-carbon length), palmitoyl (MU-C16, 16-carbon length), and oleoyl (MU-C18, 18-carbon length), derived respectively from hexanoic, heptanoic, octanoic, nonanoic, palmitic, and oleic fatty acids (Table 1, page 280/para 2, page 282/para 1, page 284/para 2/lines 3-4); and wherein five different lipase samples are respectively applied in the method/assay, which respectively comprise porcine lipase, wheat lipase, steapsin lipase, Castor bean lipase, and Peanut lipase (table 2, page 279/last line – page 280/line 5). Jacks et al. further demonstrated that MU-C8 (octanoyl ester) is a fluorogenic substrate most favorably hydrolyzed by castor bean lipase and it is also effectively hydrolyzed by all 5 lipases tested, while MU-C6 (hexanoyl ester), MU-C7 (heptanoyl ester), or MU-C9 (nonanoyl ester) is most favorably hydrolyzed by other 4 lipases tested (Table 2, page 285/para 2). Jacks et al. further demonstrated that MU-C8 (octanoyl ester) is a fluorogenic substrate more favorably hydrolyzed by all 5 lipases tested when compared to the fatty acyl esters having long carbon chains, such as MU-C18 (oleoyl ester) and MU-C16 (palmitoyl easter) (see Table 2).
Regarding the fluorogenic substrate “4-methylumbelliferyl caprylate (MU-C8)” recited in the claim 1, it is noted that the octanoyl ester of 4-methylumbelliferone/4MU (MU-C8, an ester formed by octanoic acid and 4MU) taught by Jack et al. is a synonym of the claimed 4-methylumbelliferyl caprylate, as evidenced by the attached Wikipedia printout, which teaches that caprylic acid is known as octanoic acid and is a fatty acid having a 8-carbon chain; and esters of octanoic acid are known as caprylates, and the name of acyl group of octanoic acid is octanoyl or capryloyl (see page 1: para 1; and also the chemical structure and description in right panel of page 1). Regarding the limitations “hydrolase”, “esterases”, “carboxylic ester hydrolases”, “carboxylesterases” and “lipases” recited in the claims 1, 5, 6, and/or 52, the lipases taught by Jack et al. read on these claimed enzymes, as evidenced by the specification (page 16/last para/lines 3-4, and page 35, last para, lines 4-6), which discloses that lipase enzymes belong to a subclass of esterase enzymes, and carboxylic ester hydrolases belong to the hydrolase sub-subclass, and lipases are known carboxylic ester hydrolases; and as supported by Jacks et al., who teach lipase is one kind of carboxylesterases (see page 279, para 1, line 2). Regarding the limitations about the sample comprising HCPs recited in the claims 1, 5-6, and 52, the structure of HCPs in the claims is only defined with hydrolase enzymes (including esterases, carboxylic ester hydrolases, carboxylesterases and/or lipases). The sample containing lipases taught by Jacks et al. comprises all the claimed hydrolase enzymes which define the HCPs, thus meeting the structures of the HCPs and the claimed limitations.
The method of Jack et al. differs from the method of the claim 1 in that Jack et al. do not teach the container used for the enzymatic assay is a microplate. However, it is a common practice in the prior art to use a microplate for enzymatic assay, as evidenced by Jahn et al., whose teachings are described below (see page 8). It is an obvious design choice to use a microplate as a container in the method of Jack et al. for holding reaction mixtures and performing the enzymatic assay of lipases. Examiner notes that the adjustment of particular working conditions of enzymatic assay is deemed merely a matter of design choice, judicious selection, and routine optimization which is well within the purview of the skilled artisan having the cited reference as a guide. Thus, the claims would have been obvious over the teachings of Jack et al.
Regarding Claim 17, Jack et al. teach adding 0.6 mmol of the fluorogenic substrate into a reaction mixture of 6 ml, which is equivalent to 0.1 mM concentration of the fluorescent substrate, reading on the claimed range “about 0.1-5 mM”.
Regarding Claim 19, Jack et al. teach exposing the sample to an excitation wavelength 330 nm and detecting at an emission wavelength 450 nm (page 281, para 4), which read on the claimed ranges “300-400 nm” and “400-500 nm”, respectively.
Regarding Claim 30, Jack et al. (page 281, lines 4-5) teach performing a negative control by forming a reaction mixture containing only the buffer and fluorogenic ester substrate, and performing the reaction in the absence of lipase enzyme under condition of the assay, and Jack et al. indicate that fluorogenic esters did not hydrolyze in the absence of the enzyme (i.e. no fluorescent 4MU is converted from the fluorogenic esters in the negative control). Given Jack et al. conducted the negative control and detected background fluorescent signal in the buffer and abstract, it would have been obvious to subtract such background fluorescent signal (if background signal is detected) from the fluorescent signal of regular assay for accurately quantifying the lipase enzymatic activity.
Therefore, the invention as a whole would have been prima facie obvious to a person of ordinary skill in the art at the time the invention was made.
Claims 1, 2, 5-6, 11-15, 17-19, 23-24, 30, and 51-52 are rejected under 35 U.S.C. 103 as being unpatentable over Jahn et al. (Pharm. Res., 37(6): 118, pages 1-13, published on June 3, 2020, cited in IDS) in view of Jacks et al. (Analytical Biochemistry, 1967, 21:279-285, of record) and Nakao et al. (US 2010/0075380, cited in IDS).
Jahn et al. teach that polysorbates are critical stabilizers in biopharmaceutical protein formulations, but they degrade due to hydrolysis catalyzed by lipases present in host cell proteins (HCPs) in drug substance (DS) and drug product (DP) during storage, which leads to a comprised quality of final biopharmaceutical products; and the purpose of their study is to develop an assay to detect lipolytic activity in biopharmaceutical
DS and DP formulations (suitable for assessing a lipase activity towards polysorbate degradation in the formulations) (abstract; page 2: left col/para 1/lines 1-4 and para 2/last 6 lines, and right col/para 2/lines 1-5). Jahn et al. further teach a method for determining an enzymatic activity of HCPs in a sample comprising lipases, which is based on hydrolysis of lipase substrate 4-methylumbelliferyl oleate (4MuO) to yield a fluorescent product 4-methylumbelliferone (4Mu, reading on the “MU” in claim 1) (abstract/method, Fig. 1), wherein the method comprises steps: (a) forming a reaction mixture in a microplate by adding a sample containing CCHF (cell culture harvest fluid) containing a mixture of lipases as parts of the HCPs, a HCMB reaction buffer, and a solution of 100 uM 4-methylumbelliferyl oleate (4MuO) as a fluorogenic substrate to a 96-well microplate, wherein a lipase PPL is used as control (abstract/method; page 3/right col/para 2 and left col/para 1/lines 3-6 from bottom; page 1/Abbreviation section; page 3/left col/last para – page 4/left col/para 2); (b) monitoring the conversion of 4MuO to the fluorescent product 4Mu (MU) by measuring fluorescence signals of sample/reaction mixture with excitation and emission wavelengths respectively at 330 nm and 495 nm; and (c) quantifying the enzymatic activity of the ligases/HCPs based on the conversion of the fluorogenic substrate, wherein a 4Mu calibration curve, prepared from reference standard concentrations of 4Mu, is used for quantifying the enzymatic activity, where amounts of the conversion product 4Mu stand for the quantified enzymatic activities (page 4: left col/paras 2 and 3/fluorescent analysis, and right col/para 3; page 3/left col: first and last para/lines 4 - last line; Figs. 7 and 10). It is noted that the 4-methylumbelliferyl oleate (4MuO) taught by Jahn et al. is a 4-methylumbelliferone carboxylate easter having a carbon chain length 18, as evidenced by the disclosure of the specification (Fig. 1, page 6/para 3) as well as Jahn et al. (Fig. 1). Regarding the limitations “hydrolase”, “esterases”, “carboxylic ester hydrolases”, “carboxylesterases” and “lipases” recited in the claims 1, 5, 6, and/or 52, the lipases taught by Jahn et al. read on these claimed enzymes, as evidenced by the specification (page 16/last para/lines 3-4, and page 35, last para, lines 4-6), which discloses that lipase enzymes belong to a subclass of esterase enzymes, and carboxylic ester hydrolases belong to the hydrolase sub-subclass, and lipases are known as carboxylic ester hydrolases; and as evidenced by Jacks et al., who teach lipase is one kind of carboxylesterases, as indicated above.
The method of Jahn et al. differs from the method of the claim 1 in that the fluorogenic substrate used in the method of Jahn et al. is 4-methylumbelliferyl oleate (i.e. 4MuO or MU-C18, carboxylate ester of 4-MU having an 18-carbon chain), but not 4-methylumbelliferyl caprylate (i.e. MU-C8, carboxylate ester of 4-MU having an 8-carbon chain).
The teachings of Jacks et al. are described above.
Nakao et al. teach an assay for determining enzymatic activities of lipases (LIP32 and LIP40) expressed in E. coli by using 4-methyl unberyferylcaprylate (MU-C8) or 4-methyl unberyferyloleate (MU-C18) as a fluorogenic substrate subjected to hydrolyzing by the LIP32 or LIP40 lipase, for releasing a fluorescent product for detecting the lipase activities, wherein the assay comprises steps: forming a reaction mixture comprising 0.1 mM MU-C8 or MU-C18, a reaction buffer, and a sample containing a lipase; monitoring conversion of MU-C8 or MU-C18 to the fluorogenic substrate and measuring change in fluorescence intensity; and quantifying the lipase activities (paras 0069, 0071; Examples: paras 0174, 0186/last 6 lines, 0187-189, 0195, 0207-208, 01218-219, and Tables 5-7). It is noted that MU-C8 and MU-C18 used in the assay of Nakao et al. read on the MU-C8 in the instant claim 1 and the 4 MuO of Jahn et al., respectively. It is further noted that the lipases of Nakao et al. read on the “hydrolase”, “esterases”, “carboxylic ester hydrolases”, “carboxylesterases” and “lipases” recited in the claims 1, 5, 6, and/or 52, for the reasons indicated above.
It would have been obvious to one of ordinary skill in the art to use 4-methylumbelliferyl caprylate (MU-C8) as the fluorogenic substrate in the method of Jahn et al. for detecting and determining lipase/lipolytic activities of biopharmaceutical formulations, as taught by Jacks et al. and Nakao et al. One of ordinary skill in the art would have been motivated to do so, because the 4-methylumbelliferyl caprylate (MU-C8) is a fluorogenic substrate well known in the art for determining lipase activities in an enzymatic assay, as supported by Jacks et al. and Nakao et al. In addition, Nakao et al. demonstrate that both MU-C8 (recited in the claims) and MU-C18 (used by Jahn et al.) are fluorogenic substrates suitable for effectively determining lipase activities. Furthermore, Jacks et al. demonstrate that MU-C8 is a fluorogenic substrate most favorably hydrolyzed by certain lipases and it is more favorably hydrolyzed by all the lipases tested when compared to the fatty acyl esters having long carbon chains, such as MU-C18. As such, applying MU-C8 as a fluorogenic substrate in the method of Jahn et al. would increase the sensitivity of detecting lipase activities of biopharmaceutical formulations. One of ordinary skill in the art has a reasonable expectation of success at modifying the method of Jahn et al., because the methods for applying MU-C8 as a fluorogenic substrate for determining lipase activities has been well established in the art, as supported by Jacks et al. and Nakao et al. Furthermore, the fluorescent product (i.e. 4-methylumbelliferone, 4-MU) released from lipase-catalyzed hydrolysis of MU-C8 is the same as that released from MU-C18, thus being readily detected by the fluorescent detection method of Jahn et al. for estimating hydrolase activity of HCPs.
Regarding Claim 2, Jahn et al. teach the sample comprises HCPs comprising a mixture of lipases. As such, the enzymatic activity determined by the method of Jahn et al. represents the collective activity of two or more HCPs in the sample, thus meeting the claimed limitation.
Regarding Claims 11-15, Jahn et al. teach the CCHF sample comprises a recombinant protein: a monoclonal antibody mAb1 produced by a CHO host cell line (page 3/left col/para 1/last 6 lines), wherein the monoclonal antibody mAb1 is an IgG 1 monoclonal antibody (page 10, left col, lines 9-10).
Regarding Claim 17, Jahn et al. further teach the 4MuO substrate at 100 mM is added to the assay mixture at 5% (v/v) (page 3/right col: para 2/lines 6-7), which can be converted to a final concentration 5 mM. Although the concentration taught by Jahn et al. does not match the claimed concentration range, it is considered that the concentration of Jahn et al. can be readily modified through routine optimization for improving the detection of lipolytic lipase activities. Furthermore, Jack et al. and Nakao et al. teach adding MU-C8 at concentrations in the claimed range, as indicated above. Thus, the claimed concentration would have been obvious over Jahn et al. in the absence of any showing of unexpected results or criticality. See MPEP 2144.05.
Regarding Claim 19, Jahn et al. teach exposing the sample to an excitation wavelength 330 nm and detecting at an emission wavelength 495 nm, which read on the claimed ranges “300-400 nm” and “400-500 nm”, respectively.
Regarding Claim 23, Jahn et al. teach their method is suitable for assessing a lipase activity that causes polysorbate degradation in biopharmaceutical products, as indicated above. Further, Jahn et al. specifically assess/measure levels of lipase activity in samples as well as the polysorbate degradation in the samples, which caused/contributed by the lipase activity (see Figs. 9 and 10).
Regarding Claims 18, 24, and 51, Jahn et al. further teach using their method to detect lipolytic activities in intermediate and final drug products, such as at downstream of purification process, for better and specifically reducing or depleting lipase HCPs from the products (abstract: conclusion/lines 1-4); and that the downstream purification process typically includes various chromatographic and (ultra)filtration steps to capture target therapeutic proteins expressed in mammalian or microbial host cells, and deplete HCPs (page 2: left col/last 6 lines, and right col/lines 1-3). Jahn et al. further teach that residual lipase activities in DS and DP due to sub-optimal depletion during the downstream purification process leads to hydrolysis of polysorbate and compromised quality of final biotherapeutic products; and that monitoring lipolytic activities should be carried out for improvement of downstream purification process and selecting purification process that overcomes lipase-related issues (page 2, right col, para 2). Given Jahn et al. expressively teach various chromatographic purification steps are typically applied for purifying target proteins in the downstream purification process for removing/depleting HCPs, it would have been obvious to practice the method of Jahn et al. to detect and monitor a lipolytic lipase activity of a chromatography purified pool sample for assessing effectiveness of chromatographic purification processes over removing/depleting HCPs from the biotherapeutic product that comprises the target protein, thus arriving at the claimed method of claim 18. Given Jahn et al. teach using their method for detecting/evaluating lipolytic activities during purification of intermediate and final drug products for selecting a protein purification process and improving removal of residual lipase HCPs, it would have been obvious to further comprise steps of comparing different purification processes and selecting an effective protein purification process in the method suggested by Jahn et al. for improving removal of lipase HCPs from intermediate and final products, thus arriving at the claimed method of claims 24 and 51.
Regarding Claim 30, Jahn et al. further teach forming a negative control (i.e. a sample negative control, and a control of non-enzymatic hydrolysis) without adding the lipase sample (the sample with HCPs) for applying the assay to biopharmaceutical formulations, followed by monitoring conversion of the fluorogenic substance in the negative control (page 4/right col/last para/lines 2-4 from bottom; page 10/left col/para 1: lines 2-5 and 7-8, Fig. 8). Jahn et al. are silent about subtracting an amount of fluorescent signal in the negative control from an amount of fluorescent signal in the assay reaction mixture containing the lipase sample, as recited in the step “(c)” of the claim. However, it would have been obvious to subtract an amount of fluorescent signal in the negative control from an amount of fluorescent signal in the assay reaction mixture that contains lipase sample, in the method suggested by Jahn et al. for accurately quantifying the lipolytic enzymatic activity of HCPs in biopharmaceutical formulations, because Jahn et al. teach that the 4MuO substrate generates 4Mu from non-enzymatic hydrolysis of 4MuO in the absence of lipase HCPs (see the sample negative and non-enzymatic hydrolysis controls in Fig. 8), and one of ordinary skill in the art would have recognized that fluorescent signal from 4Mu of negative control should be subtracted given it is not generated by lipase HCPs.
Regarding the claim 4, Jahn et al. do not teach that the reaction mixture comprises at least two different fluorogenic substrates. However, it would have been obvious to one of ordinary skill in the art to include one or more additional fluorogenic substrates, such as MU-C6, MU-C7, and/or MU-C9, in addition to the MU-C8 used in the method suggested by Jahn et al., Jacks et al., and Nakao et al. for improving sensitivity of detecting and determining lipase activities of HCPs and providing a better assessment for potential polysorbate degradation. This is because Jahn et al. teach the sample comprises HCPs comprising a mixture of different lipases; and Jack et al. demonstrate that different lipases have different sensitivity towards to 4-methylumbelliferone esters with different carbon chain lengths, and that fluorogenic substrates with different carbon chain lengths such as MU-C6, MU-C7, and MU-C9 (in addition to MU-C8) are more preferentially hydrolyzed by lipases. As such, combination of fluorogenic substrate MU-C8 with fluorogenic substrate having different carbon chain lengths (e.g. 6, 7, and/or 9 carbons) in the method suggested by Jahn et al. and other cited prior art would allow various different lipases in HCPs of drug substances (DS) or drug products (DP) to be more effectively detected in the assay, thus providing accurate estimation of hydrolytic enzymatic activities towards polysorbate degradation of DS and DP.
Therefore, the invention as a whole would have been prima facie obvious to a person of ordinary skill in the art at the time the invention was made.
Claims 4 and 7 are rejected under 35 U.S.C. 103 as being unpatentable over Jahn et al. (Pharm. Res., 37(6): 118, pages 1-13, published on June 3, 2020, cited in IDS) in view of Jacks et al. (Analytical Biochemistry, 1967, 21:279-285, of record) and Nakao et al. (US 2010/0075380, cited in IDS), as applied to Claims 1, 2, 4-6, 11-15, 17-19, 23-24, 30, and 51-52, further in view of Zhang et al. (Journal of Pharmaceutical Sciences, 2020, 109: 3300-3307, published on July 25, 2020, cited in IDS) and Nalder et al. (Biochimie, 2016, 128-129: 127-132, cited in IDS).
The teachings of Jahn et al., Jacks et al., and Nakao et al. are described above.
Regarding Claims 4 and 7, Jahn et al. do not teach the reaction mixture of the assay comprises at least two different fluorogenic substrates and an additional fluorogenic substrate has a carbon chain length of 8, 10, 12, 16, or 18.
Nalder et al. teach three new esters of 4-Hydroxy-N-propyl-1,8-naphthalimide (NAP) for fluorescence-based assay for activities of lipases and esterases, in which the NAP fluorophore is esterified with short (butyrate), medium (octanoate) and long (palmitate) chain fatty acids; and that a simple and rapid assay was developed and used to analyze a range of enzymes including lipases, esterases and phospholipases; and that clear differences are observed about hydrolysis of the various chain length esters, with lipases preferentially hydrolyzing the medium chain ester, whereas esterases reacted more favorably with the short ester (abstract, title). Nalder et al. further teach that these esters provide alternate substrates, which possess distinctly different excitation (447 nm) and emission (555 nm) optima (abstract/last 6 lines, page 128/left col/para 3).
Zhang et al. teach that polysorbates (PS) are surfactants commonly added in a
therapeutic product to protect proteins from denaturation, significant degradation of PS could lead to shortened drug shelf life, and a major root causes of PS degradation is the host cell protein (HCP) derived lipases and esterases (abstract/lines 1-4). Zhang et al. also teach that porcine liver esterase is able to specifically hydrolyze PS80, lipoprotein Lipase (LPL) has enzymatic activity on both PS20 and PS80, and lysosomal phospholipase A2 (LPLA2) can degrade both PS20 and PS80 at less than 1 ppm (page 3300, left col, last 5 lines). Zhang et al. further identified two esterase hydrolases (carboxylesterases) in a mAb drug substance, and PS80 becomes stable after the two hydrolases are depleted (2nd half of abstract).
It would have been obvious to one of ordinary skill in the art to include an additional fluorogenic substrate, specifically an ester of 4-Hydroxy-N-propyl-1,8-naphthalimide (NAP), in the method suggested by Jahn et al. and other cited prior art for detecting activities of additional esterase and phospholipase enzymes in biotherapeutic formulations (e.g. mAb drug) for better assessment of hydrolytic enzymatic activities towards polysorbate degradation, because it is known in the art that the ester of NAP is a fluorogenic substrate applied for a simple and rapid assay of a range of various enzymes including esterases and phospholipases (in addition to lipases assayed in the method suggested by Jahn et al.), and all these enzymes are HCPs-derived enzymes that degrade polysorbate (PS20 and PS80), as supported by Nalder et al. and Zhang et al. Furthermore, the ester of NAP possesses distinctly different excitation (447 nm) and emission (555 nm) optima, as taught by Nalder et al., which do not have confliction with the excitation (330 nm) and emission (495 nm) optima of 4MuO used by Jahn et al., thus being compatible with 4MuO for the fluorescence-based assay.
Regarding the carbon chain length recited in the claim 7, it is noted that the fluorogenic NAP substrates taught by Nalder et al. have a carbon chain length of 4, 8 or 16 (see Fig. 1), thus meeting the claimed limitation about the additional fluorogenic substrate having a carbon chain length of 8, 10, 12, 16, or 18.
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.
Response to Arguments
Applicant's arguments about the rejection of claims 1-2, 5-6, 11-15, 17-19, 23-24, 30 and 51-52 under 35 USC 103 over Jahn et al. in view of Al-Kady in the response filed on 02/27/2026 (pages 6-10) have been fully considered, but they are moot because the rejection has been withdrawn and the grand of the rejection in this office action is different from that in the previous office action.
Applicant's arguments about the rejection of claims 4 and 7 under 35 USC 103 over Jahn et al. in view of Al-Kady, Nakao et al., and Zhang et al. in the 02/27/2026 response (pages 10-11) are based on the arguments previously presented for the 103 rejection over Jahn et al. in view of Al-Kady in pages 6-10 of the response. These arguments have been fully considered, but they are moot for the reasons indicated above.
Overall, the conclusion of the obviousness of the claims 1-2, 4-7, 11-15, 17-19, 23-24, 30, and 51-52 has been established over the combined teachings of Jahn et al., Jahn et al., Jacks et al., Nalder et al., and/or Zhang et al. for all the reasons indicated above.
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