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 .
Election/Restrictions
Applicants’ election of claims 33, 102-104, 112, 118, 127-131, 134, and 135 (Group I) in the reply filed on 13 July 2026 is acknowledged. Because applicant did not distinctly and specifically point out the supposed errors in the restriction requirement, the election has been treated as an election without traverse (MPEP § 818.01(a)).
Claim Status
Claims 1, 33, 47, 102-104, 112, 118, 127-131, 134, and 135 are pending.
Claims 33, 102-104, 112, 118, 127-131, 134, and 135 were elected.
Claims 1 and 47 were withdrawn.
Claims 33, 102-104, 112, 118, 127-131, 134, and 135 were considered.
Claims 33, 102-104, 112, 118, 127-131, 134, and 135 are rejected.
Priority
The instant application filed on 15 November 2022 claims priority to PCT/US2021/032750 filed 17 May 2021, Provisional 63/142046 files 27 January 2021, Provisional 63/090772 filed 13 October 2020, Provisional 63/074575 filed 4 September 2020, and Provisional 63/025555 filed 15 May 2020. As such the effective filing date of the application is 15 May 2020.
Information Disclosure Statement
The IDS filed 15 November 2022 was considered by the examiner.
Drawings
The drawings filed 15 November 2022 are accepted.
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 130 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. The claim includes the limitation of stopping the assay process if a good regression fit cannot be identified. Neither the claim nor the specification indicates at what point the assay process is stopped. In order to determine the regression fit as good, it would be necessary to complete the steps of claims 102 and 127 (from which 130 depends). It is unclear as to what point in the process one would have enough information regarding the regression fit in order to decide to stop the assay process. As such, the claim is indefinite.
Claim Interpretation
For the purposes of applying prior art, claim 130 will be interpreted as stopping the assay process at any point.
Claim Rejections - 35 USC § 102
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
The following is a quotation of the appropriate paragraphs of 35 U.S.C. 102 that form the basis for the rejections under this section made in this Office action:
A person shall be entitled to a patent unless –
(a)(1) the claimed invention was patented, described in a printed publication, or in public use, on sale, or otherwise available to the public before the effective filing date of the claimed invention.
(a)(2) the claimed invention was described in a patent issued under section 151, or in an application for patent published or deemed published under section 122(b), in which the patent or application, as the case may be, names another inventor and was effectively filed before the effective filing date of the claimed invention.
Claim 33 is rejected under 35 U.S.C. 102(a)(1) and 102(a)(2) as being anticipated by Shepherd and Steinke. (US 6,262,798 B1, 17 July 2001, pages 1-41) (Herein referred to as Shepherd.) Shepherd teaches a method and apparatus that allows accurate spectrophotometric determination of the concentrations of various hemoglobin species in whole blood, to maximize the true optical absorbance of whole blood, and to minimize the effects of light scattering on the spectrophotometric measurements of the concentrations of various constituent components. [Abstract] Shepherd further teaches correcting the hemoglobin concentration measurements for light scattering. [Abstract] Shepherd additionally teaches a preferred embodiment of the present invention which includes a digital computer that computes the apparent optical densities at each of the selected wavelengths, corrects the concentration measurements for the effects of light scattering by the red blood cells and for nonspecific light-scattering losses, corrects for the effects of the finite spectral bandwidth of the plurality of substantially monochromatic wavelengths, and reports the final results to the operator. [Column 8, lines 25-49] Shepherd teaches an apparatus that assesses the optical transmittance of a sample of unaltered whole blood to attain an accurate measurement of its total hemoglobin concentration[column 4, lines 51-55] and measuring at a variety of wavelengths [Figure 1].
Claims 102, 103, 118, and 134 are rejected under 35 U.S.C. 102(a)(1) as being anticipated by Bhutani et al. (Pediatrics, 1 November 2015, pages e1268-e1275) (Herein referred to as Bhutani.)
With respect to claim 102, Bhutani teaches whole blood samples were analyzed for G6PD activity. (Page e1269, Middle column, last paragraph – right column, continued paragraph) Bhutani further teaches substrate solution containing D-glucose-6-phosphate, NADP, magnesium chloride, and maleimide in Tris buffer. (Page e1270, left column, last paragraph – middle column, continued paragraph) Bhutani additionally teaches whole blood samples were diluted in lysis solution, hemoglobin concentration was determined by measuring absorbance at 540 nm, the sample was further diluted, the sample was mixed with substrate, fluorescence of NADPH was measured to detect reaction product, and the activity was normalized using hemoglobin concentration. (Page e1270, right column) Bhutani additionally teaches that the measure of G6PD activity was recorded in U/g Hb. (Page e1271, Figure 2, units on graph)
With respect to claim 103, Bhutani teaches using NADP and maleimide. (Page e1270, middle column, lines 2-4)
With respect to claim 118, Bhutani teaches using glucose-6-phosphate and NADP. (Page e1270, middle column, lines 1-2)
With respect to 134, Bhutani teaches that the measure of G6PD activity was recorded in U/g Hb. (Page e1271, Figure 2, units on graph) Bhutani further teaches determining the concentration of hemoglobin in the whole blood sample and that the G6PD reported activity was normalized by using the hemoglobin concentration. (Page e1270, right column, lines 18-19 and 39-40) Bhutani additionally teaches that the samples were initially diluted at a ratio of 1:9 and then later diluted to a final ratio of 1:27. (Page e1270, right column, lines 3-4 and 20-21) For the final measure of G6PD activity to be reported in U/g Hb, calculations to convert units would have been necessary. It was known convention at the time of the instant application to convert units for reporting results from a G6PD assay. An example of this is seen in instructions from Sentinel Diagnostics on how to calculate G6PD activity in mU/g Hb. (Sentinel Diagnostics, 23 June 2015, pages 1-2) (Page 2)
Claim Rejections - 35 USC § 103
In the event the determination of the status of the application as subject to AIA 35 U.S.C. 102 and 103 (or as subject to pre-AIA 35 U.S.C. 102 and 103) is incorrect, any correction of the statutory basis (i.e., changing from AIA to pre-AIA ) for the rejection will not be considered a new ground of rejection if the prior art relied upon, and the rationale supporting the rejection, would be the same under either status.
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 102, 104, 112, 134, and 135 are rejected under 35 U.S.C. 103 as being unpatentable over Bhutani in view of Zhang et al. (US 8192995 B2, 5 June 2012, pages 1-23) (Herein referred to as Zhang.) Bhutani teaches the limitations of claims 102 and 134 as applied above under 35 U.S.C. 102.
Bhutani does not teach correcting absorbance measurements for scatter from non-hemoglobin compounds.
Zhang teaches a method of correction of particle interference to hemoglobin measurement of a blood sample, measuring absorbance of the sample mixture at a predetermined wavelength of a hemoglobin chromogen formed in the sample mixture, obtaining an apparent hemoglobin concentration of the blood sample using obtained absorbance, obtaining a corrected hemoglobin concentration of the blood sample, and reporting the corrected hemoglobin concentration of the blood sample. [Abstract] Zhang further teaches that the hemoglobin chromogen is measured by spectroscopy at a predetermined wavelength, typically between 400 nm and 680 nm [column 7, lines 2-5] and wavelengths of 380 nm-770 nm, inclusive [claim 11], and a wavelength of about 540 nm [claim 12]. Zhang additionally teaches a computer system on the hematology analyzer [column 20, line 38], determining a corrected hemoglobin concentration of said blood sample by removing a contribution of said first and second subpopulations of cellular particles to said apparent hemoglobin concentration [claim 1], and concentrations and said sizes of said subpopulations of cellular particles are measured by a light scatter measurement [claim 10].
It would have been prima facie obvious to one of ordinary skill in the art at the effective
filing date of the invention to have combined the method of Bhutani to assay for G6PD activity in blood samples with the method of Zhang to provide a corrected hemoglobin concentration. Zhang discloses that in a sample mixture used for measuring hemoglobin concentration, red blood cells are lysed to release hemoglobin molecules, but other cellular particles remain in the sample mixture. [Column 1, lines 39-42] Zhang further discloses that it is known that these cellular particles present in the sample mixture interfere with the spectrophotometric measurement of hemoglobin, because the cellular particles absorb and scatter the incident light used in the measurement. [Column 1, lines 43-47] Zhang states that these cellular particles in the sample mixture used for hemoglobin measurement may cause significant overestimation of the hemoglobin concentration. [Column 1, lines 56-58] Since Bhutani’s method relies upon hemoglobin concentration in the samples and determines that concentration based on measuring absorbance, one of ordinary skill in the art would have been motivated to utilize a method to ensure a corrected hemoglobin absorbance measurement. There would have been an expectation of success when incorporating the method of Zhang into the method of Bhutani to provide a corrected hemoglobin absorbance measurement and thereby provide a more accurate measure of G6PD activity. The invention is therefore prima facie obvious.
Claims 102, 127, and 129 are rejected under 35 U.S.C. 103 as being unpatentable over Bhutani in view of Casadevall i Solvas et al (JOVE, 10 December 2011, pages 1-9), F. Zhang et al (Diagnostic Microbiology and Infectious Disease, 15 September 2012, pages 263-266), and SoftMax Pro (Molecular Devices, 4 December 2019, pages, 1-130). (Herein referred to as Solvas, F. Zhang, and SoftMax, respectively.) Bhutani teaches the limitations of claim 102 as applied above under 35 U.S.C. 102.
Bhutani does not teach plotting the RFU data vs time.
With respect to claim 127, Solvas teaches an algorithm to differentiate signal bursts from the noise background of the oil phase. (Page 3, Section 4.2, #4) F. Zhang teaches an average of 3 blank tests and that this average intensity of the blank assay was considered as the background noise and was subtracted from the results of the fluorescence detection of the enzyme assays. (Page 265, left column, bottom paragraph) SoftMax teaches blank measurements, averaging group blanks, and subtracting blank values from other non-blank values. (Pages 102, headings 2-5; 116, headings 2-4; and 117, headings 2 and 3) SoftMax further teaches graph functions in general (page 41) and a function to calculate Vmax as the slope of plotted RFU data per time. (Page 53, Vmax heading) SoftMax additionally teaches weight values, identifying outliers, and the ROUT test which removes outliers (page 38, headings 3-5), and a weighting formula (page 51, heading 4). SoftMax teaches calculating slope (page 51, heading 2 and 53, paragraph 2), intercept (page 45, heading 4), R2 (page 50, last heading), and regression parameters (page 81, heading 6). While the SoftMax results would be calculated based on RFU per time, it is standard in the art to convert these units from fluorescence values to total NADPH activity and to convert from NADPH units of activity to a measure of G6PD activity per volume. Cell Biolabs discloses a method of converting from corrected fluorescence to total NADPH. (Cell Biolabs, Inc, 10 September 2018, pages 1-8) (Page 7, bottom) Karakaya and Ozkul disclose a protocol with instructions to convert from NADPH units of activity to G6PD units of activity. (Bioprotocol, 5 October 2016, pages 1-6) (Page 4, Part C: 1-2)
With respect to claim 129, SoftMax teaches calculating the linear regression of subsets of the data using creeping iteration. (Page 56, middle paragraph)
It would have been prima facie obvious to one of ordinary skill in the art at the effective
filing date of the invention to have combined the method of Bhutani with the methods of Solvas and F. Zhang and the software features of SoftMax. Bisswanger discloses that any enzyme assay requires a blank, especially dehydrogenase reactions, and for stopped assays the blank value is obligatory to determine the velocity from the difference between the stopped value and the blank. (Perspectives in Science, 21 March 2014, pages 41-55) (Page 51, right column, first full paragraph) Bisswanger further discloses that continuous assays also need a blank to adjust the instrument to zero, otherwise the reaction may fall outside the observation range of the system. (Page 51, right column, first full paragraph) F. Zhang teaches, as described above, that subtracting the blank value eliminates the background noise. SoftMax discloses that their formulas, accessors, and functions can be used to create powerful data analysis templates for Protocol files in order to completely automate the analysis and results reporting for plate reads. (Page 5, paragraph 1) One of ordinary skill in the art would have been motivated to combine the known and available methods of using a blank, subtracting blank values, plotting data, and regression analysis in order to detect the G6PD-substrate reaction by measuring G6PD activity. It would have been obvious to plot the RFU values against time as a measure of G6PD activity and do a regression analysis to get the best fit data using known software tools. It would also have been obvious to then convert the units to conventional forms frequently used in the art, as was described above in standard protocol documents. The invention is therefore prima facie obvious.
Claim 128 is rejected under 35 U.S.C. 103 as being unpatentable over Bhutani, Solvas, F. Zhang, and SoftMax in view of Rreg (Stata Manual, 6 November 2017, pages 1-7) and Qreg (Stata Manual, 8 March 2018, pages 1-33) Bhutani, Solvas, F. Zhang, and SoftMax teach the limitations of claims of 102 and 127 as described above.
Bhutani, Solvas, F. Zhang, and SoftMax do not teach subjecting positive residuals to a weighting of (1/residual).
Rreg teaches a Robust Regression by performing a regression, calculating case weights from absolute residuals, and regressing again using those weights. (Page 6, lines 7-9) Rreg further teaches that iterations stop when the maximum change in weights drops below tolerance. (Page 6, lines 9-10) Rreg teaches using Huber weights which are based on the inverse of the residual. (Page 6, paragraph 4) Qreg teaches Quantile Regression and weights positive and negative outliers differently. (Page 31, last paragraph)
It would have been prima facie obvious to one of ordinary skill in the art at the effective
filing date of the invention to have combined the methods of Bhutani, Solvas, F. Zhang, and SoftMax with the Rreg and Qreg methods of Stata. MATLAB Help Center discloses that if the distribution of errors is asymmetric or prone to outliers, model assumptions are invalidated, and parameter estimates, confidence intervals, and other computed statistics become unreliable. (MATLAB Help Center, Reduce Outlier Effects Using Robust Regression, downloaded 2 October 2026, pages 1-5) (Page 1, paragraph 2) MATLAB Help Center further discloses that robust regression uses a method called iteratively reweighted least squares to assign a weight to each data point and this method is less sensitive to large changes in small parts of the data. (Page 1, paragraph 3) In the instant application, the specification states that the applicants observed the G6PD assay shows that interferences in the assay only produce a positive fluorescence signal and do not produce a lower fluorescing signal, so they weigh the positive and negative outliers differently. [Page 29, middle paragraph] Given this disclosed observation, one of ordinary skill in the art would expect to see success in using a robust regression analysis while applying weights asymmetrically when doing a regression analysis. As a result, one of ordinary skill in the art would be motivated to combine the methods of Bhutani, Solvas, F. Zhang, and SoftMax with the known Stata tools or Rreg and Qreg. The invention is therefore prima facie obvious.
Claim 130, and dependent claim 131, are rejected under 35 U.S.C. 103 as being unpatentable over Bhutani, Solvas, F. Zhang, and SoftMax in view of Pamula et al. (US 2010/0041086 A1, 18 February 2010, pages 1-21) (Herein referred to as Pamula.) Bhutani, Solvas, F. Zhang, and SoftMax teach the limitations of claims 102 and 127 as described above.
Bhutani, Solvas, F. Zhang, and SoftMax do not teach stopping the assay process.
Pamula teaches combining the reaction droplet with a quench droplet comprising one or more quenching reagents selected to substantially stop the enzyme reaction. [Claim 131]
It would have been prima facie obvious to one of ordinary skill in the art at the effective
filing date of the invention to have combined the methods of Bhutani, Solvas, F. Zhang, and SoftMax with the method of Pamula. Millington et al discloses that enzymatic assays are the most straightforward type of assay to perform on the digital microfluidics platform because all that is required is to mix sample with reagent, stop the reaction after a defined time and then detect the product. (Expert Review of Molecular Diagnostics, 13 Jul 2018, 701-712) (Page 704, left column, middle paragraph) Bisswanger discloses that to determine the enzyme reaction velocity, the reaction must be stopped after a defined time and the amount of product formed or substrate converted must be analyzed thereafter by a subsequent chemical indicator reaction or a separation method. (Page 44, right column, middle paragraph) One of ordinary skill in the art would have been motivated to incorporate the known method of incorporating a stopping reagent into an enzymatic assay in order to be able to stop the reaction at a defined point. The invention is therefore prima facie obvious.
Claim 131 is rejected under 35 U.S.C. 103 as being unpatentable over Bhutani, Solvas, F. Zhang, SoftMax, and Pamula in view of Westgard. (WestgardQC, February 2004, pages 1-6) Bhutani, Solvas, F. Zhang, SoftMax, and Pamula teach the limitations of claims 102, 127, and 130 as described above.
Bhutani, Solvas, F. Zhang, SoftMax, and Pamula do not teach flat allowable error for fitting residuals. SoftMax teaches calculating residuals (page 50, heading 4) and flagging results (page 78, heading 4).
Westgard teaches that for analytical performance, a quality requirement can be defined in terms of an “allowable total error,” such as specified in the CLIA proficiency testing criteria for acceptable performance. (Page 3, paragraph 4) Total allowable errors can include both set, “flat”, error thresholds and percent-based thresholds. This can be seen in Westgard’s exemplary listing of CLIA-defined allowable total errors. (Page 3, paragraph 5)
It would have been prima facie obvious to one of ordinary skill in the art at the effective
filing date of the invention to have combined the methods of Bhutani, Solvas, F. Zhang, SoftMax, and Pamula with the teachings of Westgard. Westgard discloses examples of total allowable error based on Clinical Laboratory Improvement Amendments (CLIA) guideline. These guidelines are established and well known in the art for equivalent quality testing. Therefore, one of skill in the art would have been motivated to incorporate the CLIA guidelines described by Westgard into the G6PD assay analysis. The invention is therefore prima facie obvious.
Conclusion
Claims 1, 33, 47, 102-104, 112, 118, 127-131, 134, and 135 are eligible under 35 USC 101 (eligibility). The claims are drawn to methods that do not constitute abstract ideas.
Any inquiry concerning this communication or earlier communications from the examiner should be directed to SHARON LEVINE GRAFF whose telephone number is (571)317-0219. The examiner can normally be reached Mon - Fri 7:30 AM - 4 PM.
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/S.L.G./Examiner, Art Unit 1687
/Karlheinz R. Skowronek/Supervisory Patent Examiner, Art Unit 1687
