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 Response of 19 Feb. 2026 has been entered.
Claims 3-13 are currently pending.
Election/Restrictions
Applicant’s election without traverse of the invention of Group II, claims 3, 4, 8, 9 and 12, in the reply filed on 3-13 is acknowledged.
Claims 5-7, 10 and 11 are withdrawn from further consideration pursuant to 37 CFR 1.142(b) as being drawn to a nonelected invention, there being no allowable generic or linking claim. Election was made without traverse in the reply filed on 19 Feb. 2026.
Claims 3, 4, 8, 9, 12 and 13 are considered here.
Allowable Subject Matter
Claims 3 and 13 are allowed.
The closest prior art is represented by US20100120085 to Hyman et al., which teaches a method of isolating/characterizing microorganisms from a liquid sample comprising mammalian cells and microorganisms (e.g., blood), comprising a step of selectively lysing the non-microorganism cells in the sample using a lysis solution (e.g., claim 1). Hyman teaches that the lysis solution can comprise one or more detergents and discloses a list that includes saponin (a non-denaturing lytic detergent) and bile salts (a denaturing lytic detergent) ([0044]). Hyman does not expressly teach or suggest the specific composition of saponin and cholate. Moreover, Hyman teaches that the "non-denaturing detergents and solubilizers are used at concentrations above their critical micelle concentration (CMC), while denaturing detergents may be added at concentrations below their CMC. For example, non-denaturing lytic detergents can be used at a concentration of about 0.010% to about 10%" ([0044]). While the range of saponin taught by Hyman overlaps the claimed range, Maslova et al., Crystallography Reports 63.3 (2018): 472-475, evidences that the CMC for sodium cholate is 13 mM (or ~0.56%), which is significantly below the claimed range. Consistent with this, Okazaki et al., Journal of Japan Oil Chemists' Society 29.10 (1980): 743-747, evidences that sodium cholate shows full hemolytic activity in the low-mM concentration range (Fig. 1 and related text). Thus, Hyman fails to teach the specific combination of saponin/cholate recited in the claims, and also fails to teach the claimed concentration of cholate (rather, Hyman suggests a concentration below the CMC of ~0.56%).
Claims 3 and 13 are also distinguishable from the combination of WO2019222862 in view of Mitra et al., Journal of agricultural and food chemistry 49.1 (2001): 384-394 which was cited in the EP Search Report (see IDS of 12 March 2026). WO2019222862 teaches a method of selectively lysing mammalian cells in a blood sample comprising microorganisms using a lysis solution comprising saponin (e.g., claim 1). WO2019222862 does not teach any combination with cholate or bile salts, but rather teaches that the lysis solution can further comprise a non-ionic surfactant (e.g., claim 20), which would not include cholate (an anionic surfactant). WO2019222862 further teaches a mechanism for selective lysis where "saponin, as a surfactant, may form a layer over the microbial cells without causing damage to the microbial cells" and "if a reagent were provided with both saponin and a high pH, the surfactant layer generated by saponin may act as a protective layer, shielding the microbial cells from the otherwise deleterious effect of the high pH environment" (p. 28, lines 8-17). There is no suggestion in WO2019222862 or the art that cholate would be effective in combination with saponin for such a mechanism. Mitra teaches use of a combination of saponin/cholate to solubilize purified cholesterol, but there is not suggestion for using such combination in a cell lysis method as claimed.
Claim Objections
Claim 9 is objected to because of the following informalities:
The "0.6-20" range for saponin should have a "%".
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.
Claims 4, 8, 9 and 12 are rejected under 35 U.S.C. 112(b) 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 4 recites a "method of separating and enriching microorganisms, the method comprising the method of claim 3 and further comprising, after the incubation, separating possible microorganisms from other components in a volume of the liquid sample and enriching the microorganisms." Since the claim recites "possible microorganisms", it is unclear whether the separating and enriching steps are required steps of the claimed method - e.g., in instances where the sample does not contain microorganisms.
Claim 8 recites a "method of selectively lysing mammalian eukaryotic cells in a liquid sample comprising mammalian eukaryotic cells and possibly bacterial cells, and thereafter visualizing any viable bacterial cells in the sample, the method comprising the method of claim 3." It is unclear how claim 8 further limits claim 3, as the method of claim 8 comprises "the method of claim 3". It is unclear whether the recitation "and thereafter visualizing any viable bacterial cells in the sample" is a required step of the claimed method or rather an intended result (e.g., carrying out the lysis so as to render the bacteria visualizable). Moreover, any visualizing would not occur where bacteria are absent, making it unclear how the method of claim 3 is further limited in such cases. The rejection can be overcome by amending the claim to expressly recite a step of visualizing bacteria in the sample (e.g., the method of claim 3, further comprising visualizing…).
Claim 9 recites "incubating the reference sample in the visualization buffer during illumination". There is insufficient antecedent basis for "the visualization buffer". The rejection can be overcome by amending as follows: "a visualization buffer".
Claim 9 further recites steps of isolating and visualizing "possible bacterial cells", making it unclear whether such steps are required steps of the claimed method in instances where the sample does not contain microorganisms.
Claim 12 recites "wherein the liquid sample is selected from the group consisting of a blood sample, urine, beverage, water, liquid, and liquidized food sample." The meaning of the second recitation of "liquid" is unclear. The rejection can be overcome by deleting said recitation from the list.
Conclusion
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/ROBERT J YAMASAKI/Primary Examiner, Art Unit 1657