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 .
Status of the Claims
Claims 1-18 are pending in the application and are the subject of this office action.
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and the fee set forth in 37 CFR 1.17(e) has been timely paid, the finality of the previous Office action has been withdrawn pursuant to 37 CFR 1.114. Applicant's submission filed on 15 April 2026 has been entered.
Claim Rejections - 35 USC § 112(d)
The following is a quotation of 35 U.S.C. 112(d):
(d) REFERENCE IN DEPENDENT FORMS.—Subject to subsection (e), a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
The following is a quotation of pre-AIA 35 U.S.C. 112, fourth paragraph:
Subject to the following paragraph [i.e., the fifth paragraph of pre-AIA 35 U.S.C. 112], a claim in dependent form shall contain a reference to a claim previously set forth and then specify a further limitation of the subject matter claimed. A claim in dependent form shall be construed to incorporate by reference all the limitations of the claim to which it refers.
Claims 7-12 are rejected under 35 U.S.C. 112(d) or pre-AIA 35 U.S.C. 112, 4th paragraph, as being of improper dependent form for failing to further limit the subject matter of the claim upon which it depends, or for failing to include all the limitations of the claim upon which it depends.
Claims 7-12 fail to further limit the subject matter of the claim upon which they depend because claims 1-6 already require that the electrochemical change associated with the formation of the complex of the insulin and the insulin binding protein is detected by EIS, as recited in claim 1.
Applicant may cancel the claim(s), amend the claim(s) to place the claim(s) in proper dependent form, rewrite the claim(s) in independent form, or present a sufficient showing that the dependent claim(s) complies with the statutory requirements.
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 1-18 are rejected under 35 U.S.C. 103 as being unpatentable over Davis et al (US 2015/0177180 A1; IDS entered) in view of Funabashi et al (JP 2016-109666 A; IDS entered) and Garrote et al (Perspectives on and precautions for the uses of electric spectroscopic methods in label-free biosensing applications. ACS Sensors; 4(9), 2216-2227 (2019).; previously cited).
Regarding claims 1, 4-13, and 16-18, Davis teaches an insulin detection method (Abstract) comprising:
Adding a sample to an electrode having an insulin binding protein immobilized on a surface of the electrode, the insulin binding protein specifically recognizing insulin (Par. 33: electrodes of the present invention comprise probe molecules disposed on the planar surface of a substrate, wherein the probe molecules are capable of binding selectively to a target species such as insulin; Par. 124, 139: sample applied to electrode); and
Detecting, using the electrode, a change in electrochemical impedance associated with formation of a complex comprising the insulin and the insulin binding protein, wherein the change in electrochemical impedance is detected by EIS (Abstract: an electrode for use in the electrochemical detection of insulin; Par. 33: the functionalized electrodes may be used for the EIS detection of insulin in a sample; Par. 64).
Davis does not teach that the insulin binding protein includes a first region and second region as described in instant claims 1 and 13, and instead teaches that the insulin binding protein is preferably an anti-insulin antibody (Abstract; Par. 37: most preferably, the probe is an antibody that selectively binds to the target analyte). Davis does not explicitly teach providing the disclosed features and inventions as part of a kit.
Regarding claims 1 and 13, Funabashi teaches an insulin detection kit and method (Par. 1).
Funabashi teaches a method wherein insulin is detected by binding of two labeled polypeptides which are brought into proximity when bound to a common insulin molecule such that they produce a detectable LRET or BRET signal to indicate the presence and concentration of insulin in a sample (Par. 7, 19-20)
Funabashi teaches an insulin binding protein, wherein the insulin binding protein includes:
A first region that includes an alpha-CT segment of an insulin receptor and does not include a beta subunit of the insulin receptor, and a second region that includes an L1 domain of the insulin receptor and receptor and does not include the beta subunit of the insulin receptor, wherein one of the first region and the second region is immobilized on a substrate (Par. 19: a first polypeptide comprising an alpha-CT segment of an insulin receptor and not containing a beta subunit of an insulin receptor. A second polypeptide containing the L1 domain of the insulin receptor and not including the beta subunit of the insulin receptor; Par. 58: the first complex (alpha-CT) and the second complex (L1) may be linked to each other by a linker to form an integrated third complex; Par. 59: linking the alpha-CT segment and L1 domain ensures that both polypeptides are held in proximity to one another such that they may both form a complex with a given molecule of insulin to produce a detectable signal even when concentration of insulin in the sample is low, thus increasing detection sensitivity of the assay; Par. 65: the third complex (i.e. linked alpha-CT segment and L1 domain) can be attached to the surface of a substrate such that the first complex and second complex can be maintained in a suitable arrangement relative to one another).
Regarding claims 4-12 and 16-18 Funabashi further teaches that the second polypeptide (i.e. second region) may comprise additional components in addition to the L1 domain (Par. 51: the second polypeptide may consist of the L1 domain of the insulin receptor or may contain a constitution other than the L1 domain of the insulin receptor), and further indicates that an exemplary second polypeptide may have an amino acid sequence which comprises both an L1 domain and a CR domain (Par. 95: L1 encoded by the DNA sequence shown in SEQ ID NO: 3 and having the amino acid sequence shown in SEQ ID NO: 4 was used as L1 in this example; wherein SEQ ID NO: 4 of Funabashi comprises both an L1 domain and a CR domain, as shown in the sequence alignment below which indicates that SEQ ID NO: 4 of Funabashi is a 100% match to SEQ ID NO: 9 of the instant application, which is defined by the instant specification at Pg. 28, Par. 51 to represent an insulin receptor L1 domain and CR domain).
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Garrote discloses limitations and optimizations for electrochemical biosensing (Abstract).
Garrote teaches that one of the limitations of electrochemical biosensing for small molecules is the target to receptor size ratio, wherein a low target to receptor size ratio (i.e. a small target and larger receptor) negatively affects the sensitivity of detection (Abstract; Figs. 5-6; Pg. 2223, Col. 2: low target to receptor size ratio negatively affects sensitivity. It is common for assays with higher target to receptor size ratios to show higher sensitivities than those with lower values; Pg. 2224: the target to receptor size ratio appears to be important in different types of faradaic EIS platforms. This is because the process is based on how the charge transfer resistance between the solution and the metallic electrode surface is impeded or based on changes in the electrochemical occupancy of the redox moieties within the SAM, wherein both receptor and target sizes will proportionally affect the signal. To overcome the lower sensitivity observed in low target to receptor size ratio assay, an ideal solution is to increase the target to receptor size ratio by using a smaller molecular weight receptor).
It would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to modify the insulin detection taught by Davis to use the insulin binding protein comprising a linked alpha-CT segment and L1 domain (such as the one taught by Funabashi) in place of the anti-insulin antibody taught by Davis (wherein the insulin binding protein in the modified invention would not necessarily comprise the detectable labels taught by Funabashi, given that these would be superfluous in the electrochemical detection method taught by Davis, which does not require the use of labels). One would be motivated to use the linked alpha-CT segment and L1 domain in particular (as opposed to non-linked versions of the polypeptides which are also described in Funabashi) because the incorporation of two different binding components in each insulin binding protein would increase the efficiency and completeness of binding between the insulin binding protein and the insulin in the sample by providing two binding sites per probe instead of one. One would be motivated to use an insulin binding receptor wherein the second region comprises both an L1 domain and a CR domain because Funabashi specifically teaches this as an exemplary and functional example of a polypeptide used to bind and detect insulin.
One would be motivated to make this modification because the insulin binding protein taught by Funabashi is significantly smaller than the anti-insulin antibody taught by Davis, such that substitution of the smaller insulin binding protein of Funabashi for the larger antibody taught by Davis would create a larger target to receptor size ratio, and could enable more sensitive detection of lower concentrations of insulin, as taught by Garrote. Wherein both the prior art and the instant specification indicate that an alpha-CT segment of an insulin receptor consists of 16 amino acid residues, an L1 domain consists of about 155 amino acid residues, and the combined L1 and CR domain of Funabashi SEQ ID NO: 4 (equivalent to instant SEQ ID NO: 9) consists of 310 amino acid residues, while the linker used to link the alpha-CT segment and the L1 domain in Funabashi is taught to be 20 or more amino acid residues (Funabashi, Par. 62). Such that the insulin binding protein as a whole may be as small as about 346 amino acid residues in total (when the CR domain is included) which is significantly smaller than an antibody such as the IgG1 anti-insulin antibody taught by Davis (Davis, Par. 119), wherein an IgG antibody has a molecular weight of about 150 kDa. One of ordinary skill in the art would have a reasonable expectation of success in making this modification because Davis teaches that the probe immobilized to the electrode should be capable of selectively binding to insulin and may be a protein (Par. 8, 37), while Funabashi teaches a protein that selectively binds to insulin.
Additionally, since both Davis and Funabashi are directed to detection of insulin in a sample comprising the use of protein probes which bind specifically to insulin and which can be immobilized to a substrate for the detection assay, one of ordinary skill art would recognize that substitution of the probe taught by Funabashi for the antibody taught by Davis amounts to simple substitution of known elements to achieve predictable results with a reasonable expectation of success.
It would have been obvious to one of ordinary skill in the art to further modify Davis in view of Funabashi and Garrote to include the disclosed features in a kit, as taught by Funabashi, because incorporation of the components and reagents required for performing the insulin detection in a single kit is efficient and useful for the end user. One of ordinary skill in the art would have a reasonable expectation of success in making this modification because both Davis and Funabashi are directed to assays for insulin detection comprising insulin binding proteins.
Regarding claims 2-3 and 14-15, Davis in view of Funabashi and Garrote teaches that the first region and second region are connected by a linker, as described in the rejection of claim 1 above.
Davis teaches that the insulin binding protein is immobilized to the surface of an electrode, as described above.
Funabashi also teaches that the linked first region and second region may be immobilized on a substrate, but is generic regarding the specific orientation of immobilization.
As such, it would have been obvious to one of ordinary skill in the art before the effective filing date of the claimed invention to immobilize the insulin binding protein either at the second region (as in claim 2) or at the first region (as in claim 3), because both orientations are obvious to try based on a finite number of potential orientations for immobilizing the insulin binding protein (i.e. the insulin binding protein must be immobilized at one end or the other, such that either orientation is obvious to try from a total of two potential options). One of ordinary skill in the art would have a reasonable expectation of success in making this modification because both Davis and Funabashi disclose the detection of insulin using an insulin binding protein immobilized to a surface.
Response to Arguments
Applicant’s arguments filed 15 April 2026 have been fully considered.
Regarding the 103 rejection, applicant argues that the teachings of the prior art do not provide a reasonable expectation of success in modifying the EIS insulin sensor of Davis to substitute the insulin-receptor fragment probe taught by Funabashi for the anti-insulin antibody taught by Davis.
Applicant argues that the behavior of receptor fragments outside their native quaternary assembly is often unpredictable, and that immobilization on a solid surface introduces further uncertainty due to orientation, tethering, and surface effects. Applicant argues that surface immobilization can restrict the conformational freedom of proteins and change the effective orientation of binding sites, especially for multi-domain constructs. Applicant argues that the behavior of the linker and the relative positions of the insulin receptor regions near a conductive surface are not predictable from the behavior of the same polypeptides in homogeneous solution (i.e. as is demonstrated in Funabashi). Applicant argues that EIS depends on complex interfacial phenomena including charge transfer resistance, double-layer effects, probe packing density, and orientation, which are not addressed by the Funabashi prior art.
Applicant argues that although Funabashi explicitly states that the receptor fragment probe (i.e. third complex) can be attached to the surface of a substrate for detection of insulin, this bare statement does not remedy the deficiencies noted above. Applicant argues that this statement is not supported by particular data or experimental results which demonstrate a working example of the immobilized embodiments.
These arguments are not persuasive. Funabashi explicitly teaches at Par. 65 that the insulin binding probe may be immobilized for detection of insulin, and explicitly indicates that this immobilization may take into account orientation-related effects on how the probe binds to insulin. In fact, the teaching indicates that such immobilization may be specifically used to ensure proper orientation of the receptor fragments for binding to insulin (Par. 65: The third complex (i.e. linked alpha-CT segment and L1 domain) can be attached to the surface of a substrate such that the first complex and second complex can maintain good arrangement for each other. If the support body is used, insulin can be detected with high sensitivity). Though it is acknowledged that Funabashi does not provide specific data demonstrating reduction to practice of this particular embodiment, a reference is not required to provide explicit reduction to practice of every possible embodiment, and the teachings of the reference indicate that one of ordinary skill in the art had an explicitly stated expectation of success in using the probe in an immobilized embodiment for high sensitivity detection of insulin.
The argument that the behavior of receptor fragments outside their native quaternary assembly is often unpredictable is not persuasive in this instance because Funabashi teaches that the linked alpha-CT segment and L1 domain are fragments of an insulin receptor that are specifically shown to still be capable of binding to insulin outside their native quaternary assembly. While applicant indicates that other factors such as sub-optimal orientation, restricted conformational freedom, or poor packing after immobilization can impact the performance of the receptor, as shown in instant Figs. 3-4, this argument does not undercut a general reasonable expectation of success because while it shows differing levels of detection sensitivity with different receptor arrangement, it shows that the receptor is still functional (i.e. successful) in multiple different arrangements.
Applicant's argument that EIS depends on complex interfacial phenomena including charge transfer resistance, double-layer effects, probe packing density, and orientation, which are not addressed by the Funabashi prior art is not persuasive because Funabashi is not relied upon to teach EIS in particular, but rather is relied upon to teach a particular insulin binding probe structure which binds specifically to insulin in a sample. The particulars of EIS detection are taught by Davis which teaches that insulin may be detected via EIS using an insulin-binding probe immobilized on an electrode. The teachings of Funabashi are relied upon in the rejection to teach that the structure of the insulin binding probe was known in the art and that one of ordinary skill in the art would have had a reasonable expectation of success in replacing the insulin-binding probe taught by Davis with the insulin-binding probe taught by Funabashi.
Additionally, it is noted that while Applicant argues that there is some unpredictability in immobilization of the insulin-binding probe, this does not effectively disprove reasonable expectation of success, as reasonable expectation of success does not require absolute predictability of success (MPEP 2143.02: conclusive proof of efficacy is not required to show a reasonable expectation of success, nor is absolute predictability of success required. The expectation of success need only be reasonable, not absolute). The teachings of the prior art provide one of ordinary skill in the art with enough evidence and enough showing of predictability to support a reasonable expectation of success (i.e. Funabashi teaches that the probe (which comprises receptor fragments outside their native quaternary assembly) can be effectively used to bind insulin in solution, and explicitly teaches that the probe can also be immobilized, wherein this teaching explicitly addresses factors highlighted in Applicant's arguments such as ensuring proper orientation of the binding fragments).
Applicant further argues that Davis and Garrote are based on an increase in impedance cause by steric hindrance at the electrode surface, where the bound analyte obstructs access of redox species to the electrode, and that this is different from the instantly claimed invention wherein binding of insulin induces a large conformational change in the artificial insulin receptor itself which significantly increases the access barrier to the electrode surface, and that this conformational change results in a stronger impedance response. Applicant further argues that this mechanism was not predictable and yielded unexpectedly improved results, wherein the instantly disclosed sensor is capable of detecting insulin at low nanomolar concentration substantially better than the 50-800 nM limits observed in the prior solution-phase LRET/BRET system of the Funabashi reference.
These arguments are not persuasive. Regarding the exact mechanism of impedance change, Davis, Garrote, and the instant disclosure all teach EIS detection methods wherein a detectable change in impedance is caused by an increase in the access barrier to the electrode surface caused by binding of the target protein (i.e. insulin) to the probe immobilized on the electrode. While the prior art may not explicitly teach the particular conformational change of the insulin-receptor probe which is discussed in the instant disclosure, this does not render the claimed invention un-obvious over the prior art, since this prior art provides a reasonable expectation of success in modifying the EIS detection system of Davis in view of Funabashi and Garrote with the instantly disclosed probe, as discussed above. One of ordinary skill in the art making such a modification would still expect to see a detectable change in impedance caused by an increase in the access barrier to the electrode surface caused by binding of the target protein to the probe immobilized on the electrode.
Additionally, Applicant's assertion of unexpectedly improved results, which relies on the particular lower limit of detection of the instantly disclosed sensor is not persuasive because the results generally follow from the teachings of the prior art. That is, Davis explicitly teaches that an EIS system can be used to detect insulin at concentrations even lower than the limit of detection demonstrated in the instant disclosure (see Davis Par. 127 which indicates that insulin can be detected by EIS in the low picomolar range, while the instant specification discloses detection in the low nanomolar range). The fact that one detection mechanism (i.e. EIS as taught by Davis) would yield a different (and potentially improved) limit of detection as compared to a second detection mechanism (i.e. LRET/BRET, as disclosed by Funabashi) even when using the same insulin binding probe is not particularly unexpected since different detection mechanisms often yield different levels of sensitivity. This is further bolstered by the explicit teaching of Davis that different insulin detection mechanisms and methods which were previously known in the prior art are not capable of detecting insulin with the same sensitivity as the EIS method disclosed by Davis (Par. 3).
As such, the showing in the instant specification that substitution of the insulin-binding probe taught by Funabashi into an EIS detection system such as the one taught by Davis yields a lower limit of detection than the LRET/BRET assay taught by Funabashi, but a higher limit of detection than the EIS method taught by Davis is not a persuasive showing of unexpectedly improved results which renders the instantly claimed invention non-obvious over the prior art.
Conclusion
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/ELLIS FOLLETT LUSI/Examiner, Art Unit 1677
/CHRISTOPHER L CHIN/Primary Examiner, Art Unit 1677
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