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 .
Acknowledgement of Receipt
Applicant’s Response, filed 2/26/2026, in reply to the Office Action mailed 11/26/2025, is acknowledged and has been entered. Claims 1-25 have been amended. Claims 1-28 are pending, of which claims 26 and 27 are withdrawn from consideration at this time as being drawn to a non-elected invention. Claims 1-25 and 28 are examined herein on the merits for patentability.
Response to Arguments
Applicant’s arguments have been fully considered. Any rejection not reiterated herein has been withdrawn. The previous rejection has been modified in view of claim amendment. The Examiner’s response to Applicant’s arguments is incorporated below.
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.
Claim(s) 1-25 and 28 are rejected under 35 U.S.C. 103 as being unpatentable over Gruner et al. (WO 18/189051) in view of Hossain et al. (Appl. Phys. Lett., 2010, 97, 26370).
Gruner teaches a method for X-ray fluorescence measurement, in particular for X-ray fluorescence imaging, in which it is detected whether fluorescent target particles are present in an object to be examined, in particular in the body of an examined subject, and existing target particles are localized in the object. The invention also relates to an X-ray fluorescence measuring device, in particular an X-ray fluorescence imaging device, for carrying out such a method. Applications of the invention are in the X-ray fluorescence imaging, especially for medical purposes and in particular for objects with size scales as in humans.
Aims of medical imaging are the detection, characterization and monitoring of pathological changes in the body being studied and / or pharmacokinetics, whereby the distribution of drugs in the body is measured in vivo. In order to improve these methods, techniques of so-called "molecular imaging" are being developed, for example to allow early tumor diagnosis and better characterization of the tumor tissue or pharmacokinetic examinations.
In particular, the invention should make it possible to detect the X-ray fluorescence of target particles in an object with an increased sensitivity, to render the substrate more efficient. to reduce the radiation intensity and / or to improve the spatial resolution. Furthermore, the invention is intended in particular to permit the hitherto impossible examination of larger objects and to enable clinical X-ray fluorescence imaging, in particular on humans.
The object is irradiated (scanned) with the X-ray beam at a plurality of scanning positions in the first projection plane. The scanning positions (positions where irradiation is performed) are set with a scanning device that moves the source device and the object relative to each other. For each scan position, the X-ray beam cuts the first projection plane at other coordinates. The x-ray beam is preferably aligned perpendicular to the first projection plane. The scan positions are in the first projection plane e.g. distributed as a matrix in rows and columns.
At each scanning position, the X-ray radiation emitted from the object in a plurality of spatial directions is detected with a detector array device fixedly connected to the source device. With the scanning device, the source and detector directions and moves the object relative to each other. Preferably, the source and detector means are moved relative to a stationary held object.
The detector array means comprises an array of spectrally selective detector elements (also referred to as pixels) distributed in a plurality of different spatial directions around the object.
The detector elements are arranged to detect the X-radiation in the plurality of spatial directions. The "detector element" may be a single, spectrally resolving, detecting element, a composite of a plurality (eg, 4 x 4) detecting elements, or a detecting component composed of a plurality of detecting sub-elements. The combination of several detecting elements may in particular have advantages for the statistics of the detected X-ray photons. The component consisting of sub-elements can in particular have advantages for the spectrally resolving detection when using a laser-based Thomson source as the source device, since in this case the photons are detected almost simultaneously and it is ensured by the sub-elements that per sub-element not more than 1 photon per detector time window is detected.
By searching for the significant detector elements, those detector elements are detected whose sum spectrum the X-ray fluorescence is detected with a relatively low background signal in comparison to the other detector elements. As a result, the quantitative knowledge about the anisotropy of the intrinsic subsoil is translated into a reduction of the subsurface with the aid of direction-dependent detection.
According to a preferred application of the invention in X-ray fluorescence imaging, the examined object is a human or animal subject or a body part thereof. The subject the target particles were supplied in advance, z. By oral or other administration or injection. A preparation step with a supply of target particles, in particular by injection into the body of the subject, is not part of the invention. Alternatively, other non-biological objects can be examined. The target particles comprise particles, in particular nanoparticles, which are suitable for the excitation of X-ray fluorescence with incident photons, in particular with an energy of at least 15 keV, and preferably have a mass number in the range of the mass numbers of iodine to gold. These elements have the advantage that the K-shell fluorescence photons have sufficient transmission in the body of the subject so that the target particles are detectable at any depth (as opposed to optical fluorescence). Furthermore, the target particles are functionalized with a marker substance. The marker substance (or biomarker) comprises a substance that is specific to parts of the object being examined, e.g. B. on predetermined cells or cell groups or on predetermined tissue such. As tumor cells or tissue binds. But the target particles can also be functionalized with drugs to allow in vivo pharmacokinetic studies.
The accumulation (concentration) of the target particles in the at least one subregion, e.g. B. to tissue or cell groups. The localization of the target particles comprises the detection of the location of the enrichment, at least with respect to the first projection plane, but preferably with respect to all three spatial dimensions. In particular, the invention has the following advantages for medical applications. The intrinsic background in medical X-ray fluorescence imaging can be reduced so much that at a justifiable radiation dose minimal amounts of functionalized gold nanoparticles can be detected in the body to enable a tumor early diagnosis.
The energy of the photons of the exciting X-ray beam is particularly preferably selected in a narrow energy interval above the K edge of the fluorescent element in the target particles, with gold target particles at approximately 85 keV.
See Figure 13, if a fluorescence signal is present, that is, the two gold fluorescence lines, these are separated from one another in the energy spectrum of the sum signal of the considered detector elements by a pure background region and there are also left and right signal regions Background areas (see Figure 13): in the background area, there can be only background photons and no signal photons. These areas are therefore a reference value. If there is no fluorescence signal, the signal regions would not statistically differ significantly from the background regions.
Figure14shows the X-ray spectrum which would be measured according to the invention and is distinguished by a considerable background reduction. The statistical significance of the signal from both fluorescence lines in Figure 14B is more than 10 times the standard deviation.
Gruner does not teach a second marker substance.
Hossain teaches multiple DNA and protein biomarkers have been detected based on characteristic x-ray fluorescence of a panel of metal and alloy nanoparticles, which are modified with ligands of biomarkers to create a one-to-one correspondence and immobilized on ligand-modified substrates after forming complexes with target biomarkers in three-strand or sandwich configuration. By determining the presence and concentration of nanoparticles using x-ray fluorescence, the nature and amount of biomarkers can be detected with limits of 1 nM for DNA and 1 ng/ml for protein. By combining high penetrating ability of x-rays, this method allows quantitative imaging of multiple biomarkers.
Metallic nanoparticles of metals and alloys such as indium, bismuth, tin, and lead-tin alloy nanoparticles are made by thermal decompositions of organometallic precursors in ethylene glycol in the presence of a surfactant polyvinylpyrrolidone at 200 °C under nitrogen protection. The nanoparticles are surface-oxidized by heating at 110 °C for 10 min in ambient condition, and aminized by incubating in 5% alcoholic solution of aminopropyltriethoxy-silane for 1 h. After removing excess silane by centrifugation, aminized nanoparticles are suspended in dimethyl sulfoxide DMSO containing 5% N-succinimidyl 4-iodoacetyl aminobenzoate SIAB for 20 min. After removing excess SIAB, nanoparticles are incubated with thiolated probe single strand DNA ssDNA in phosphate buffer solution PBS pH 7.0 at room temperature for 3 h. Amine-modified nanoparticles are then incubated with disuccinimidyl suberate DSS in DMSO for 1 h. After removing excess DSS, nanoparticles are incubated with antibody inside PBS for 3 h. Solid substrates i.e., aluminum plate are modified with capture ssDNA or antibody (page 1).
In Figure 3, a color online a XRF spectrum of bismuth nanoparticles after detecting 100 ng/ml of protein biomarker, where the bottom and top lines are background and signal, respectively; b the peak areas of bismuth L1 top and L1 bottom as a function of protein concentration; c XRF spectrum of bismuth nanoparticles immobilized on an aluminum plate with 100 ng/ml of protein biomarker and then covered with 10 mm PMMA; d spatially distributed intensities of two discrete areas on a plastic substrate covered with bismuth and lead-tin nanoparticles.
A similar method has been used to detect ssDNA using indium, tin, or bismuth nanoparticles not shown. Providing other conditions nanoparticle diameter, collection time, x-ray flux, and detector-sample distance are identical, lead-tin alloy and bismuth nanoparticles provide the highest detection sensitivities compared to other nanoparticles due to their higher fluorescence yield. The sensitivity of detection can be enhanced by stacking substrates with nanoparticles, and detecting x-ray emission in transmission mode (page 2).
The feasibility of in vivo protein biomarker detection/ imaging using XRF has been studied after immobilizing prostate specific membrane antigens PSMAs onto substrates. This method has the potential to detect multiple cancer biomarkers inside the human body. Nanoparticles with suitable surface modification can be taken into body, and conjugated onto tumors through antigen-antibody interaction. The penetrating power of incoming x-rays can be tuned by using beams of higher energy. The dose absorbed by the body can be further reduced by using monochromatic beams or beams with less background by appropriate filtration. By quantitatively imaging multiple surface biomarkers of cancer cells, the reliability of in vivo cancer detection can be hopefully enhanced without much harmful effect to human (page 3).
It would have been obvious to one of ordinary skill in the art at the time of the invention to provide a second marker substance in the methods of x-ray fluorescence imaging of a sample including a body part and a substance comprising metal (gold) nanoparticles coupled to a targeting ligand as a marker substance, including detection of spatial resolution and distribution thereof, when the teaching of Gruner is taken in view of Hossain. One would have been motivated to do so, with a reasonable expectation of success, because Hossain teaches that provision of first and second metallic nanoparticles coupled to biomarkers allows for imaging of multiple biomarkers, such as potential to detect multiple cancer biomarkers inside the human body.
With regard to the amended limitation directed to irradiation of the sample with monochromatic or narrow-band X-ray radiation, it is noted that Gruner teaches photons of the exciting X-ray beam is particularly preferably selected in a narrow energy interval above the K edge of the fluorescent element in the target particle (page 7).
With regard to the amended limitation wherein the irradiation photon energy of the X-ray radiation has a distance from a highest absorption edge of all marker substances in the sample at which simultaneously the background noise levels of the marker substances are minimal and the fluorescence probabilities are similar, it is noted that Gruner teaches greatly reduced background signal is achieved in order to achieve a significantly higher efficiency of the method with regard to dose and irradiation time (page 8), and further both Gunther and Hossain teach marker substances according to the dependent claims including metal nanoparticles, etc; it is noted the recitation of similar fluorescence probability is a term of degree, and further that the fluorescence probabilities are interpreted to be capable of being similar, as the same materials are meet the structural requirements of the instant claims. A composition and its properties are inseparable.
Response to arguments
Applicant argues that WO 2018/189051 A1 represents technological background with regard to in vivo XRF imaging with a single marker substance and that Hossain discloses multiplexed sensing of multiple elements with XRF measurements (without practical imaging), demonstrated with plate shaped model samples. Applicant asserts that the subject matter of claim 1 cannot be obvious in view of WO 2018/189051 Al in combination with Hossain because at least the following features of claim 1 are not suggested by the combination of conventional techniques: (i) the first and the at least one further marker substance exhibit fluorescence probabilities, attenuations of the X-ray fluorescence in the sample, and background noise levels on account of scattering in the sample which are equal or are similar to such an extent that the detection of the X-ray fluorescence at the same concentration of the marker substances results in comparable statistical significance levels, and (ii) the irradiation photon energy of the X-ray radiation is above the absorption edges of all the marker substances, wherein the irradiation photon energy of the X-ray radiation has a distance from a highest absorption edge of all marker substances in the sample at which simultaneously the background noise levels of the marker substances are minimal and the fluorescence probabilities are similar. Applicant argues that defining a combination of marker substances according to (i) cannot be disclosed in WO 2018/189051 A1 because both documents are silent with regard to multiplex imaging of different markers. Applicant further argues that Hossain is also silent with regard to (i) because background noise does not play a role in Hossain.
Applicant’s arguments have been fully considered but are not found to be persuasive. It is respectfully submitted that in response to applicant's arguments against the references individually, one cannot show nonobviousness by attacking references individually where the rejections are based on combinations of references. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981); In re Merck & Co., 800 F.2d 1091, 231 USPQ 375 (Fed. Cir. 1986). In the instant case, while Gunther does not specifically teach a second marker substance, it is known to provide first and second markers, as first and second metallic nanoparticles coupled to biomarkers allows for imaging of multiple biomarkers, such as potential to detect multiple cancer biomarkers inside the human body. With regard to the amended limitation directed to irradiation of the sample with monochromatic or narrow-band X-ray radiation, it is noted that Gruner teaches photons of the exciting X-ray beam is particularly preferably selected in a narrow energy interval above the K edge of the fluorescent element in the target particle (page 7).
With regard to the amended limitation wherein the irradiation photon energy of the X-ray radiation has a distance from a highest absorption edge of all marker substances in the sample at which simultaneously the background noise levels of the marker substances are minimal and the fluorescence probabilities are similar, it is noted that Gruner teaches greatly reduced background signal is achieved in order to achieve a significantly higher efficiency of the method with regard to dose and irradiation time (page 8), and further both Gunther and Hossain teach marker substances according to the dependent claims including metal nanoparticles, etc; it is noted the recitation of similar fluorescence probability is a term of degree, and further that the fluorescence probabilities are interpreted to be capable of being similar, as the same materials are meet the structural requirements of the instant claims. A composition and its properties are inseparable. With regard to the assertion that background noise does not play a role in Hossain, it is noted that reduction of background noise is readily stated in Gunther, e.g. see page 7; and that Hussain teaches using monochromatic beams or beams with less background by appropriate filtration (page 3).
Applicant further argues that with respect to feature (ii), it is noted that Hossain does not and cannot give any indication of the selection of an irradiation photon energy according to claim 1. Applicant asserts that the X-ray tube used in WO 2018/189051 A1 does not allow the selection of an irradiation photon energy. Applicant further notes that secondly, no reference is made to any selection of an irradiation photon energy. Assuming arguendo that Hossain is the closest prior art, features (i) and (ii) have the technical effect that multiparametric X-ray fluorescence imaging of biological organisms is realized, whereby several marker substances can be detected separately in X-ray spectra. Accordingly, the technical problem underlying the invention can be seen with respect to Hossain in how multiparametric X-ray fluorescence imaging is realized on real objects in the form of biological organisms. The solution of the said technical problem by features (i) and (ii) is not suggested by Hossain insofar as it is considered on its own. Hossain makes no reference to feature (ii) in particular. To transfer the measurement on a tabular model sample described in Hossain to measurements on the human body, it is only suggested to use X-rays with a higher energy (to increase the penetration depth) and to use monochromatic X-rays or radiation with a filtered background (to reduce the dose), see Hossain, page 263704-3, right column, para. 3. However, this is not a reference to feature (ii), but a suggestion in a different direction, deviating from feature (ii). Applicant argues that Hossain had no knowledge of the effect of background noise on the measurement of biological organisms. In order to measure with the highest possible sensitivity, the skilled person would therefore choose the single-beam photon energy at or as close as possible to the absorption edge of the marker substance, based on Hossain. Only the investigations of the background noise and the realization of a minimum in the background region by the inventors have shown that it is necessary to choose the irradiation photon energy at a distance from the absorption edge in order to excite the marker substances in such a way that their emissions are in the range of the minimum of the background noise, while at the same time the fluorescence probabilities are similar.
Applicant’s arguments have been fully considered but are not found to be persuasive. With regard to the assertion that Hossain does not and cannot give any indication of the selection of an irradiation photon energy and that the X-ray tube used in WO 2018/189051 A1 does not allow the selection of an irradiation photon energy, it is respectfully submitted that Gruner readily teaches photons of the exciting X-ray beam is particularly preferably selected in a narrow energy interval above the K edge of the fluorescent element in the target particle (page 7). With regard to the assertion that
Hossain had no knowledge of the effect of background noise on the measurement of biological organisms and that the skilled person would therefore choose the single-beam photon energy at or as close as possible to the absorption edge of the marker substance, based on Hossain, wherein only the investigations of the background noise and the realization of a minimum in the background region by the inventors have shown that it is necessary to choose the irradiation photon energy at a distance from the absorption edge in order to excite the marker substances in such a way that their emissions are in the range of the minimum of the background noise, while at the same time the fluorescence probabilities are similar, see Gunther, which teaches methods reduction of background noise at page 7, as well as background reduction based on a pixel selection.
With regard to the amended limitation wherein the irradiation photon energy of the X-ray radiation has a distance from a highest absorption edge of all marker substances in the sample at which simultaneously the background noise levels of the marker substances are minimal and the fluorescence probabilities are similar, it is noted that Gruner teaches greatly reduced background signal is achieved in order to achieve a significantly higher efficiency of the method with regard to dose and irradiation time (page 8), and further both Gunther and Hossain teach marker substances according to the dependent claims including metal nanoparticles, etc; it is noted the recitation of similar fluorescence probability is a term of degree, and further that the fluorescence probabilities are interpreted to be capable of being similar, as the same materials are meet the structural requirements of the instant claims. A composition and its properties are inseparable.
Conclusion
No claims are allowed at this time.
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.
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/LHS/
/Michael G. Hartley/ Supervisory Patent Examiner, Art Unit 1618
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