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 .
DETAILED ACTION
Claims 1, 3-10, 12, and 14-16 are pending.
Response to Amendment
Applicant amended claim 1 and added the limitations “selectively” and “within 10 minutes after the adjusting of the hexose”, amended claims 6, 12, and 14-16 and canceled claims 2, 11, and 13. Applicant amended claims 12 and 14-16 to depend from claim 1 and added new limitations in claims 15-16.
The rejection of claim 6 under 35 U.S.C. 112(b) is withdrawn in view of the amendment.
The rejection of claims 15 and 16 under 35 U.S.C. 112(d) is withdrawn in view of the amendment.
The rejection of claims 11-13 and 15-16 under 35 U.S.C. 102(a)(1) is withdrawn in view of the amendment.
The rejection of claim 14 under 35 U.S.C. 103 is withdrawn in view of the amendment.
New rejection Necessitated by the amendment
Claim Rejections - 35 USC § 112
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.
Claim 15 is 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 15 recites the limitation " the acid-treated sample" in line 5. There is insufficient antecedent basis for this limitation in the claim.
Maintained Rejections
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.
Claims 1 and 3-10 remain rejected and claims 12 and 15-16 are rejected under 35 U.S.C. 103 as being unpatentable over Sunny (Food Microbiology 25.1 (2008): 183-189, of record in Office Correspondence mailed on 3/11/2026) in view of He (Methods and Applications in Fluorescence 5.2 (2017): 024002, of record in Office Correspondence mailed on 3/11/2026).
Regarding claims 1, 3-10, and 12 Sunny teaches a method of measuring the extrusion of intracellular accumulated cF upon energizing cells with glucose (an aldohexose) in food associated bacteria, specifically Lactobacillus rhamnosus (p.183 introduction). Sunny teaches that stationary growth phase cultures were harvested, washed twice in Ringer’s solution and that this step would remove saccharide as in instant claim 12. Sunny teaches the use of the fluorescent esterase substrate cFDA incubated for 10 minutes with cells, then washing the cells and adding the cell membrane damage reagent propidium iodide (PI) and further incubating for 10 minutes (p.186 par.6) (i.e., adding a reagent that exhibits fluorescence when degraded by an esterase and a reagent for determining cell membrane damage). Cells were further incubated with 20mM glucose for 40 min in order to measure the performance of cells in extruding intracellular accumulated cF (i.e., adjusting a hexose having an aldehyde group contained in the lactic acid bacteria containing beverage or food to 0.01 mM or more). Kinetic measurements were performed by withdrawing samples every 10 min for 40 min to monitor release of cF from glucose energized cells (p. 185 par. 7, Fig 5) (i.e., detecting lactic acid bacteria that excrete a fluorescent substance and have no cell membrane damage from the lactic acid bacteria-containing beverage or food within 10 minutes after the adjusting of the hexose).
Sunny does not teach the method for detecting live lactic acid bacteria in a beverage or food as in instant claim 1.
He teaches a method of quantifying live lactic acid bacteria in yogurt in figure 3 which is a lactic acid bacteria containing food and is also a milk product as claimed in instant claim 4. He uses flow cytometry and a live stain (SYTO9) combined with PI p.4 scheme 1. He teaches absolute quantification is achieved by the addition of 200 nm beads to the samples (p 3. par. 2.5) as in instant claim 9. He also teaches the above method for detection of live Lactobacillus casei (p.2 par. 2.2 and Figure 1), as in instant claim 8.
It would have been obvious for a person having ordinary skill in the art to extend the teachings of Sunny for detecting metabolically active bacteria to food products that contain probiotics because live probiotics confer health benefit on their hosts, and because lactic acid bacteria are widely used as sources of probiotics, and these bacteria are contained in foods such as yogurts and fermented milk products, see He p.1 introduction. One of ordinary skill in the art would recognize that the similar protocol using FCM would ensure a high probability of successfully applying the methods of Sunny to the analytes of He. It would have been obvious for a person having ordinary skill in the art to carry out routing optimization to arrive at an effective percentage of beads to particles. One of ordinary skill in the art would be motivated to do so in order to calculate the concentration of the bacteria.
Regarding claims 15-16, Sunny teaches an equation to calculate the relative number of population extruding the intracellular accumulated dye and teaches dividing the percentage of cells still stained with cF following glucose addition and incubation by the percentage of cells stained with cF glucose addition (para. 9 equation 3). One of ordinary skill in the art would be motivated to calculate the percentage of living bacteria and compare it to that of the whole population of bacteria in the sample as suggested by Sunny.
Claim 14 is rejected under 35 U.S.C. 103 as being unpatentable over Sunny and He as applied to claim 1 above, and further in view of Abe (Cancer Immunology, Immunotherapy 28.1 (1989): 29-33, of record in Office Correspondence mailed on 3/11/2026).
Sunny and He do not teach plotting a graph of cells which have excreted cF versus time and taking area of the plotted curve to quantify live lactic acid bacteria.
However, Abe teaches plotting concentration of a drug versus time and taking the area under the curve to quantify total serum levels on p.31-32 and table 4. It would have been obvious for a person having ordinary skill in the art to use the method of calculating the area under the curve to obtain a measure of total amount of live lactic acid bacteria and a person having ordinary skill in the art would be motivated to do this to know total amounts of live bacteria that would be available for food production or probiotic benefits to consumers.
Response to Arguments
Applicant's arguments filed 05/26/2026 have been fully considered but they are not persuasive.
Applicant argues that although Sunny discloses kinetic measurements being performed by withdrawing samples every 10 min for 40 min to monitor release of cF from glucose energized cells, Sunny does not disclose or suggest the unexpected result that more accurate measurements are obtained within 10 minutes.
In response to the argument, Sunny teaches that that rapid efflux of intracellular dye from bacteria functions as a measure of viability and teaches withdrawing samples every 10 min for 40 min (including at 10 min) to monitor release of cF from glucose energized cells (p. 185 par. 7, Fig 5). Furthermore, Applicant’s argument of an unexpected result is not commensurate in scope with the claim. Instant Fig 5 shows that the biggest difference in percentage of LcS with excretion activity between the two samples is seen at the 10 min point and at least the first two points (less than 5 min) do not show any difference between the two samples.
Applicant argues that Abe plots a drug versus time whereas the claim requires plotting a percentage of lactic acid bacteria on a vertical axis versus an elapsed time on a horizontal axis.
In response to the argument, Sunny teaches a kinetic measurement of the cF excretion rate and teaches plotting the % of cF-stained cells (a concentration) at culture periods of 0, 10, 20, 30, and 40 minutes (time) in a 20 mM glucose environment (see " 14. Kinetic measurement of the rate of cF extrusion" and Fig. 5). Abe is relied upon for teaching calculating the area under a curve and calculating a concentration as a function of time. One of ordinary skill in the art would be motivated to apply the calculation method taught by Abe to further interpret and display the results of the plot taught by Sunny.
Conclusion
Applicant's amendment necessitated the new ground(s) of rejection presented in this Office action. Accordingly, THIS ACTION IS MADE FINAL. See MPEP § 706.07(a). Applicant is reminded of the extension of time policy as set forth in 37 CFR 1.136(a).
A shortened statutory period for reply to this final action is set to expire THREE MONTHS from the mailing date of this action. In the event a first reply is filed within TWO MONTHS of the mailing date of this final action and the advisory action is not mailed until after the end of the THREE-MONTH shortened statutory period, then the shortened statutory period will expire on the date the advisory action is mailed, and any nonprovisional extension fee (37 CFR 1.17(a)) pursuant to 37 CFR 1.136(a) will be calculated from the mailing date of the advisory action. In no event, however, will the statutory period for reply expire later than SIX MONTHS from the mailing date of this final action.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to MARY A CRUM whose telephone number is (571)272-1661. The examiner can normally be reached M-F 8:00-5:00 CT with alternate Fridays off.
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/MARY A CRUM/ Examiner, Art Unit 1657
/THANE UNDERDAHL/ Primary Examiner, Art Unit 1699
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