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 .
Claims 1-4, 8, 10-17, 19-23, 25, 26, 28, 29, 31, 32 and 39-41 are pending in this application and were examined on their merits.
The objection to the Specification because the Abstract is too short to describe
the disclosure sufficiently to assist readers in deciding whether there is a need for
consulting the full patent text for details has been withdrawn due to the Applicant’s amendments to the Specification filed 03/05/2026.
The objection to the Specification because it contains embedded hyperlinks and/or other form of browser-executable code has been withdrawn due to the Applicant’s amendments to the Specification filed 03/05/2026.
The objection to the Specification because it contains a trade name or mark used in commerce has been withdrawn due to the Applicant’s amendments to the Specification filed 03/05/2026.
The rejection of Claims 13, 14, 16 and 17 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applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention, has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claim 7 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inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention, has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claims 31 and 32 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inventor (or for applications subject to pre-AIA 35 U.S.C. 112, the applicant), regards as the invention, has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claims 1, 2, 10, 11, 12, 15, 23-26, 28, 29, 31 and 32 under 35
U.S.C. § 102(a)(1) as being anticipated by Barbee et al. (1994), has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claims 1, 2, 10, 11, 12, 15, 23-26, 28, 29, 31 and 32 under 35
U.S.C. § 102(a)(1) as being anticipated by Sato et al. (2000), has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23, 24, 25, 26, 28, 29, 31, 32 and 41 under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000), has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claims 1, 2, 3, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23, 24, 25, 26, 28, 29, 31, 32, 39 and 41 under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000), and further in view of Ranjan et al. (1996), has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claims 1, 2, 5, 4, 7, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23, 24, 25, 26, 28, 29, 31, 32 and 41 under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000), and further in view of Hsiai et al. (2003), has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claims 1, 2, 4, 8, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23, 24, 25, 26, 28, 29, 31, 32, 40 and 41 under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000), and further in view of Dardik et al. (2005), has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
The rejection of Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 19, 20, 21, 22, 23, 24, 25, 26, 28, 29, 31, 32 and 41 are rejected under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000), and further in view of Kar et al. (02/04/2021), has been withdrawn due to the Applicant’s amendments to the claims filed 03/05/2026.
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.
Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23 and 41 are newly rejected under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000) in view of Hsiai et al. (2003), both of record, as necessitated by Applicant’s amendments to the claims filed 03/05/2026.
Sato et al. teaches exposing immobilized bovine (mammalian) aortic endothelial cells to a laminar flow shear stress (Pg. 128, Column 1, Lines 9-10 and Column 2, Lines 7-19 and Fig. 1);
then nanoindenting the cells using Atomic Force Microscopy/AFM with a
cantilever which is deflected/bent due to force exerted upon the cantilever by the cells it
contacts (Pg. 129, Column 1, Lines 13-14 and Column 2, Lines 1-16 and Pg. 130,
Column 1, Lines 1-16);
and determining cell stiffness of the immobilized cells at different shear stress levels (6 hour or 24 hour) (Pg. 133, Column 1, Lines 9-17 and Pg. 134, Fig. 10), and reading on Claims 1, 2, 10, 11, 12, 15 and 23,
The teachings of Sato et al. were discussed above.
Sato et al. did not teach a method wherein the cells are exposed to shear stress prior to immobilization or wherein the cell are immobilized using cell and tissue adhesive, as now required by Claim 1;
wherein the cells are suspension cells, as required by Claim 4;
wherein the probe is mechanically lowered from a pre-calibrated distance toward
the surface of the cells, as required by Claim 13;
wherein the probe is lowered for a period of two seconds, as required by Claim 14;
wherein the probe is mechanically raised for a period of two seconds, as required
by Claim 16;
wherein the cantilever is in contact with the cell surface for at least one second,
as required by Claim 17;
wherein the nanoindenter subjects the cells to six rounds of nanoindentation, as
required by Claims 21;
wherein each subsequent nanoindentation is placed 2 µm from the preceding
nanoindentation, as required by Claim 22;
or wherein the cantilever is in contact with the cell surface for five seconds, as
required by Claim 41.
Hsiai et al. teaches a method comprising exposing Bovine aortic endothelial
cells/BAECs to shear force wherein the cells are seeded on glass slides coated with a tissue and cell adhesive (Pg. 1651, Column 1, Lines 22-30 and Pg. 1652, Column 1, Lines 1-12).
It would have been obvious to those of ordinary skill in the art before the
effective filing date of the claimed invention to modify the method of Sato et al. of
exposing immobilized bovine aortic endothelial cells to a laminar flow shear stress and
then nanoindenting the cells using Atomic Force Microscopy/AFM to expose the cells to shear stress then immobilize because the cells may only be exposed to shear stress either before or after immobilization and is no more than the selection from a number of identified, predictable solutions with a reasonable expectation of success. See the MPEP at 2143, I., E., citing KSR. Further, the selection of any order of performing process steps is prima facie obvious in the absence of new or unexpected results.
See the MPEP at 2144.04, II., C. Those of ordinary skill in the art would have been motivated to make this modification in order to expose cells to shear stress and immobilize the cells for nanoindentation measurements. There would have been a reasonable expectation of success in making this modification because Sato et al. teaches both exposing cells to shear stress and cell immobilization prior to nanoindentation.
It would have been further obvious to those of ordinary skill in the art before the
effective filing date of the claimed invention to modify the method of Sato et al. of
exposing immobilized bovine aortic endothelial cells to a laminar flow shear stress and
then nanoindenting the cells using Atomic Force Microscopy/AFM to use a cell and tissue adhesive to immobilize the cells to a substrate as taught by Hsiai et al. because Sato et al. does not indicate how the BAEC are immobilized for shear stress experiments and Hsiai et al. provides a means for immobilizing BAEC to a solid substrate for shear stress experiments. Those of ordinary skill in the art would have been motivated to make this modification in order to determine the effect of shear stress on immobilized BAEC. There would have been a reasonable expectation of success in making this modification because both references are reasonably drawn to the same field of endeavor, that is, the effects of shear stress on BAEC.
With regard to Claim 4, it would have been obvious to those of ordinary skill in
the art before the effective filing date of the claimed invention to modify the method of
Sato et al. and Hsiai et al. of exposing immobilized Bovine aortic endothelial cells to a laminar flow shear stress and then nanoindenting the cells using Atomic Force Microscopy/AFM to use suspended cells in place of immobilized/adherent cells because cells are cultured only under two conditions, either adherent or suspended. Thus, selection of either condition is no more than the selection from a number of identified, predictable solutions with a reasonable expectation of success. See the MPEP at 2143, I., E., citing KSR. Those of ordinary skill in the art would have been motivated to make this modification in order to determine the effect of shear stress on suspended cells as opposed to adherent cells. There would have been a reasonable expectation of success in making this modification because cells are only cultured under two conditions, either adherent or suspended.
While the references listed above do not specifically teach the limitation of
Claims 13, 14, 16, 17, 21, 22 and 41 that; the optical probe is mechanically lowered
from a pre-calibrated distance toward the surface of the cells, the optical probe is
lowered for a period of two seconds, the cantilever is in contact with the cell surface for
at least one second, the nanoindenter subjects the cells to six rounds of
nanoindentation, wherein each subsequent nanoindentation is placed 2 µm from the
preceding nanoindentation or wherein the cantilever is in contact with the cell surface
for five seconds, one of ordinary skill in the art would recognize that these are result-
effective optimizable variables in the process of Atomic Force Microscopy.
Sato et al. teaches nanoindenting cells using Atomic Force Microscopy/AFM with an optical probe/cantilever. This is a starting point for the experimentation to determine the optimal distance to lower the optical probe, time to lower the optical probe, time of
contact between the optical probe and the cells, number of rounds of nanoindentation
performed and distance between subsequent nanoindentations. This is motivation for
someone of ordinary skill in the art to practice or test the nanoindentation parameter
values widely to find those that are functional or optimal to sufficiently provide the
desired nanoindentation data which then would be inclusive or cover the instantly
claimed values. Absent any teaching of criticality by the Applicant concerning the
claimed parameters, it would be prima facie obvious that one of ordinary skill in the art
would recognize these limitations are an optimizable variable which can be met as a
matter of routine optimization (see MPEP § 2144.05 (II)(B). Those of ordinary skill in the art before the effective filing date of the claimed invention would have been motivated to make this modification in order to obtain nanoindentation data of the shear stressed cells. There would have been a reasonable expectation of success in making these modifications because the Sato reference is drawn to the Atomic Force Microscopy/nanoindentation analysis of shear stressed cells.
Claims 1, 2, 3, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23, 39 and 41 are newly rejected under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000) in view of Hsiai et al. (2003), as applied to Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23 and 41 above, and further in view of Ranjan et al. (1996), all of record, as necessitated by Applicant’s amendments to the claims filed 03/05/2026.
The teachings of Sato et al. and Hsiai et al. were discussed above.
Neither reference taught a method wherein the cells are Chinese hamster
ovary/CHO cells, as required by Claims 3 and 39.
Ranjan et al. teaches the treatment of CHO cells and bovine aortic
endothelial/BAEC cells to experimental laminar flow stress (Pg. 383, Abstract).
It would have been obvious to those of ordinary skill in the art before the effective
filing date of the claimed invention to modify the method of Sato et al. and Hsiai et al. of determining the mechanical effect of laminar shear stress on immobilized BAEC to substitute the CHO cells of Ranjan et al. for the BAEC cells of Sato et al. because both cell types are art-recognized as suitable for use in experiments involving the effects of laminar flow shear stress. Those of ordinary skill in the art would have been motivated to make this modification based on artisan preference and the availability of cell types. There would have been a reasonable expectation of success in making this modification because at least both Sato et al. and Ranjan et al. are reasonably drawn to the same field of endeavor, that is, in vivo experiments on cell response to shear stress.
Claims 1, 2, 4, 8, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23, 40 and 41 are newly rejected under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000) in view of Hsiai et al. (2003), as applied to Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23 and 41 above, and further in view of Dardik et al. (2005), all of record, as necessitated by Applicant’s amendments to the claims filed 03/05/2026.
The teachings of Sato et al. and Hsiai et al. were discussed above.
Neither reference taught a method wherein the forces that cause shear stress are generated by bioreactor agitation or shake flask agitation, as required by Claim 8;
or wherein the forces that cause shear stress are generated by bioreactor
agitation, as required by Claim 40.
Dardik et al. teaches a method of applying shear stress to BAEC through an
orbital shaker positioned inside an incubator (e.g. a bioreactor) (Pg. 870, Column 1,
Lines 12-15 and 52-56).
It would have been obvious to those of ordinary skill in the art before the effective
filing date of the claimed invention to modify the method of Sato et al. and Hsiai et al. of exposing immobilized Bovine aortic endothelial cells to a laminar flow shear stress and then nanoindenting the cells using Atomic Force Microscopy/AFM to use the bioreactor
agitation method of Dardik et al. to generate the shear stress because both methods are
recognized by the art as suitable for subjecting BAEC to shear stress.
Those of ordinary skill in the art would have been motivated to make this modification in order to determine the effect of shear stress on immobilized BAEC. There would have been a reasonable expectation of success in making this modification because both references are reasonably drawn to the same field of endeavor, that is, the effects of shear stress on BAEC.
Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 19, 20, 21, 22, 23 and 41 are newly rejected under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000) in view of Hsiai et al. (2003), as applied to Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23 and 41 above, and further in view of Kar et al. (02/04/2021), all of record, as necessitated by Applicant’s amendments to the claims filed 03/05/2026.
The teachings of Sato et al. and Hsiai et al. were discussed above.
Neither reference taught a method wherein upon contact with the cell, the
cantilever generates multiple increasing oscillation frequencies of 1F Hz, 2F Hz, 4F Hz,
and 10F Hz, as required by Claim 19;
or wherein no oscillation frequency is generated for a period of two seconds
between the generation of each increasing oscillation frequency, as required by Claim
20.
Kar et al. teaches a method wherein the mechanical response of cells is
measured using dynamic nanoindentation wherein an oscillating dynamic load is
superimposed on a static load over a frequency range of 10-250Hz (overlapping the
claimed range of 10F Hz) and wherein the measurements are used to calculate the
storage modulus (E') and loss modulus (E") (Pg. 12, Lines 41-48).
It would have been obvious to those of ordinary skill in the art before the effective
filing date of the claimed invention to modify the method of Sato et al. and Hsiai et al. of exposing immobilized Bovine aortic endothelial cells to a laminar flow shear stress and then nanoindenting the cells using Atomic Force Microscopy/AFM to use the dynamic
nanoindentation method of Kar et al. because this would allow nanoindentation cell
testing over a broad frequency range over time. Those of ordinary skill in the art would
have been motivated to make this modification in order to dynamically determine the
effect of shear stress on immobilized BAEC. There would have been a reasonable
expectation of success in making this modification because both references are
reasonably drawn to the same field of endeavor, that is, the nanoindentation
measurement of cells.
While the references listed above do not specifically teach the limitation of Claim
20, "wherein no oscillation frequency is generated for a period of two seconds between
the generation of each increasing oscillation frequency, one of ordinary skill in the art
would recognize that this is a result-effective optimizable variable in the process of
dynamic nanoindentation.
Kar et al. teaches the dynamic nanoindenting of cells with an optical probe/nanoindenter. This is a starting point for the experimentation to determine
the optimal time between the generation of each increasing oscillation frequency. This
is motivation for someone of ordinary skill in the art to practice or test the time between
nanoindentation frequency increases widely to find those that are functional or optimal
to sufficiently provide the desired nanoindentation data which then would be inclusive or
cover the instantly claimed values. Absent any teaching of criticality by the Applicant concerning the claimed parameters, it would be prima facie obvious that one of ordinary skill in the art would recognize these limitations are an optimizable variable which can be met as a matter of routine optimization (see MPEP § 2144.05 (II)(B). Those of ordinary skill in the art before the effective filing date of the claimed invention would have been motivated to make this modification in order to obtain nanoindentation data of the shear stressed cells. There would have been a reasonable expectation of success in making these modifications because the Sato reference is drawn to the Atomic Force Microscopy/nanoindentation analysis of shear stressed cells and Kar is drawn to the dynamic nanoindentation analysis of cells.
Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23, 25, 26, 28 and 41 are newly rejected under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000) in view of Hsiai et al. (2003), both of record, as applied to Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23 and 41 above, and further in view of Liao et al. (2018), as necessitated by Applicant’s amendments to the claims filed 03/05/2026.
The teachings of Sato et al. and Hsiai et al. were discussed above.
Neither reference taught a method wherein cell stiffness is determined by calculating Young's modulus (YM) and Effective Young's modulus (EYM), as required by Claim 25;
wherein the YM and EYM of cells after 26 hours and 46 hours of shear stress is less than about 50X Pa, as required by Claim 26;
or wherein the YM and EYM of cells after 72 hours of shear stress is greater than about 500X Pa, as required by Claim 28.
Liao et al. teaches that cellular stiffness can be determined from measuring YM and EYM (Pg. 1, Abstract).
It would have been obvious to those of ordinary skill in the art before the effective
filing date of the claimed invention to modify the method of Sato et al. and Hsiai et al. of exposing immobilized Bovine aortic endothelial cells to a laminar flow shear stress and then nanoindenting the cells using Atomic Force Microscopy/AFM to determine cell stiffness to determine cell stiffness by measuring EM and EYM as taught by Liao et al. because this would provide another means for quantifying measured cell stiffness. Those of ordinary skill in the art would have been motivated to make this modification in order to determine and quantify cell stiffness as measured by AFM.
There would have been a reasonable expectation of success in making this modification because at least the Sato and Liao references are reasonably drawn to the same field of endeavor, that is, the nanoindentation/AFM measurement of cell stiffness.
With regard to Claims 26 and 28, the YM and EYM would be expected to vary dependent upon the type of cell used, the nature and force of the applied shear stress, the time that shear stress is applied and general cell culture conditions, such that the resulting effects on YM and EYM could be determined by routine experimentation and optimization of the above result-effective variables.
Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23, 29, 31, 32 and 41 are newly rejected under 35 U.S.C. § 103 as being unpatentable over Sato et al. (2000) in view of Hsiai et al. (2003), both of record, as applied to Claims 1, 2, 4, 10, 11, 12, 13, 14, 15, 16, 17, 21, 22, 23 and 41 above, and further in view of Chim et al. (2018), as necessitated by Applicant’s amendments to the claims filed 03/05/2026.
The teachings of Sato et al. and Hsiai et al. were discussed above.
Neither reference taught a method wherein cell stiffness is determined by calculating a storage modulus (an E' value) or a loss modulus (an E" value), as required by Claim 29;
wherein an E' value calculated at frequencies of 1F, 2F, and 10F Hz is higher than an E" value calculated at frequencies of 1F, 2F, and 10F Hz after at least two days of agitation, thereby indicating elasticity of the immobilized cells, as required by Claim 31;
or wherein an E" value calculated at a frequency of 4F Hz is higher than [[the]]an E' value calculated at a frequency of 4F Hz after at least two days of agitation, thereby indicating viscosity of the immobilized cells, as required by Claim 32.
Chim et al. teaches a method wherein AFM is used determine cell stiffness based on calculating the storage modulus (G’(ω)) and loss modulus (G”(ω)) (Pg. 3, Lines 8-16 and Pg. 6, Lines 6-11).
It would have been obvious to those of ordinary skill in the art before the effective
filing date of the claimed invention to modify the method of Sato et al. and Hsiai et al. of exposing immobilized Bovine aortic endothelial cells to a laminar flow shear stress and then nanoindenting the cells using Atomic Force Microscopy/AFM to determine cell stiffness to determine cell stiffness by measuring storage and loss modulus as taught by Chim et al. because this would provide another means for quantifying measured cell stiffness. Those of ordinary skill in the art would have been motivated to make this modification in order to determine and quantify cell stiffness as measured by AFM.
There would have been a reasonable expectation of success in making this modification because at least the Sato and Chim references are reasonably drawn to the same field of endeavor, that is, the nanoindentation/AFM measurement of cell stiffness.
With regard to Claims 31 and 32, the storage modulus and loss modulus values would be expected to vary dependent upon the type of cell used, the nature and force of the applied shear stress, the time that shear stress is applied, the frequency used in calculation and general cell culture conditions, such that the resulting effects on the storage and loss moduli could be determined by routine experimentation and optimization of the above result-effective variables.
Response to Arguments
Applicant’s arguments, see Remarks, filed 03/05/2026, with respect to the rejection(s) of claim(s) under 35 U.S.C. § 112(b) or 35 U.S.C. § 112 (pre-AIA ), second paragraph, 35 U.S.C. § 102(a)(1) and 35 U.S.C. § 103 have been fully considered and are persuasive. Therefore, the rejections have been withdrawn. However, upon further consideration, a new ground(s) of rejection is made in view of Sato et al. (2000) in view of Hsiai et al. (2003) as set forth in the new rejections above.
Conclusion
No claims are allowed.
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 PAUL C MARTIN whose telephone number is (571)272-3348. The Examiner can normally be reached Monday-Friday 12pm-8pm EST.
Examiner interviews are available via telephone, in-person, and video conferencing using a USPTO supplied web-based collaboration tool. To schedule an interview, Applicant is encouraged to use the USPTO Automated Interview Request (AIR) at http://www.uspto.gov/interviewpractice.
If attempts to reach the Examiner by telephone are unsuccessful, the Examiner’s supervisor, Sharmila G Landau can be reached at (571) 272-0614. The fax phone number for the organization where this application or proceeding is assigned is 571-273-8300.
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/PAUL C MARTIN/Examiner, Art Unit 1653 03/25/2026
/SHARMILA G LANDAU/Supervisory Patent Examiner, Art Unit 1653
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