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 .
Claim Status
Claims 1-20 are pending in the claim set filed 05/17/2024. All pending claims have been examined on the merits.
Withdrawn
The objections to claims 1-2, 5, 7, 10, 12, 17 and 19 are withdrawn in light of the amendments.
The rejections of claims 1-9, 11-12, 14 and 17-20 under 35 U.S.C. 112(b) are withdrawn in light of the amendments.
The rejection of claims 1-3, 6, 8-9, 11 and 14 under 35 U.S.C. 102 is withdrawn in light of the amendments. See Response to Arguments for further discussion.
New Grounds of Rejection Necessitated by Amendment
Claim Rejections - 35 USC § 103
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.
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) 1-6, 8-9, 11 and 13-14 is/are rejected under 35 U.S.C. 103 as being unpatentable over Handique et al. (US 2013/0190212 A1; previously cited), hereafter “Handique”, and further in view of Pérez-Rodríguez, et al. (Microfluidic devices for studying bacterial taxis, drug testing and biofilm formation. Microb Biotechnol. 2022 Feb;15(2):395-414; cited on Form 892), hereafter, “Perez”.
Regarding claim 1, Handique teaches a cell capture system that includes an inlet manifold (main channel) and an array that includes an inlet channel (branch channel) fluidly connected to a plurality of pores (connection channels), each pore including a chamber (observation chamber) (see pg. 1, para. [0029]). Handique teaches that the cell capture system preferably includes multiple arrays aligned in parallel (see pg. 2, para. [0033]), wherein the multiple arrays are fluidly coupled in parallel by an inlet manifold (main channel) (see pg. 2, para. [0034]). Handique teaches the cell capture system is preferably defined on a microfluidic chip (see pg. 2, para. [0029]). Therefore, Handique teaches a fluidic chip comprising a main channel (inlet manifold) fluidly connected to a plurality of branch channels (inlet channels) fluidly connected to a plurality of connection channels (pores) fluidly connected to a plurality of observation chambers (chambers), which includes all of the structures comprising the “top layer” of the claim.
Regarding the functional limitation, wherein the main channel “facilitates the flow of a sample solution”, Handique teaches the inlet manifold (main channel) is fluidly connected to a plurality of inlet channels (branch channels), as discussed above. Handique teaches that the inlet manifold (main channel) receives a sample from an inlet and distributes the sample to an inlet channel (see pg. 4, para. [0041]) which functions to receive a volume of the sample and to distribute the sample to the pores (see pg. 3, para. [0039]). Therefore, it is understood that the main channel (inlet manifold) functions to facilitate the flow of a sample solution, and Handique teaches the corresponding structure of an inlet manifold which provides a fluidic connection between the inlet and the inlet channel.
Regarding the limitation, wherein the “plurality of branch channels that are configured for facilitating the flow of oil”, Handique teaches the cell capture system to further include an isolation mechanism that functions to isolate cells within individual pores, wherein an isolation inlet is connected to the inlet channel (branch channel), wherein an “isolation material” is flowed through the isolation inlet into the inlet channel (branch channel) (see pg. 6, para. [0053]). Handique teaches that in the “the flow state”, the isolation material is a substantially viscous, immiscible liquid, such as oil, such that the isolation material does not flow into the pores (see pg. 6, para. [0054]). Therefore, Handique teaches the branch channel (inlet channel) is configured for facilitating the flow of oil.
Regarding the limitation, wherein the “plurality of connection channels that connect the branch channel and the observation chambers”, Handique teaches the plurality of connection channels (pores) are fluidly connected to the branch channels (inlet channels) and the observation chambers, as discussed above.
Handique does not teach each observation chamber is a closed chamber connected only to a respective connection channel.
Perez teaches that bacteria have coevolved to establish symbiotic or pathogenic relationships with plants, animals or humans. With human association, the bacteria can cause a variety of diseases. Thus, understanding bacterial phenotypes at the single-cell level is essential to develop beneficial applications. Traditional microbiological techniques have provided great knowledge about these organisms; however, they have also shown limitations, such as difficulties in culturing some bacteria, the heterogeneity of bacterial populations or difficulties in recreating some physical or biological conditions. See Summary.
Perez teaches that understanding bacterial phenotypes at the single-cell level is essential to develop beneficial applications, and microfluidics is an emerging technique that complements current biological assays while allowing for fine-tuning control of the test conditions as well as the recruitment of three-dimensional conditions, in which several processes can be integrated and gradients can be generated, thus imitating physiological 3D environments (see Summary). Perez teaches that devices with linear channels are the most basic design and consist of straight channels with openings at each end, and multiple channels can be added in parallel to create more versatile devices (see pg. 396, col. 2, para. 5).
Perez teaches that microfluidics offers high flexibility in terms of design, making it suitable for numerous research approaches. Developing smaller channels and adjusting them to the measurements of the studied bacteria allows the capture of bacteria according to their size and shape for identification by electron microscopy and testing of their resistance to antibiotics. Moreover, a single bacterium could be trapped to examine its generational growth. See pg. 398, col. 2 para. 3. Another approach to trap bacteria is by designing small side chambers perpendicular to the central channel where the bacteria are analyzed. Multiple E. coli were introduced into these lateral chambers to study their adaptative memory mechanisms when carbon nutrients were removed from the central channel. See pg. 398, col. 2, para. 4. Single bacteria can also be caught in these side chambers. For instance, Aldridge et al. trapped a single Mycobacterium smegmatis in each chamber and discovered that these bacteria did not show symmetrical division, instead preferentially elongating their ‘old pole’. On the other hand, Caspi drew a chamber size similar to E. coli body size, managing to catch bacteria oriented in such a way that their flagellum remained in the central channel. By applying hydrodynamic forces through the central channel, bending forces and filament deformation were calculated. See pg. 399, col. 1.
Perez also teaches that droplet-based microfluidics are characterized for their isolation and confinement of a single bacteria or small populations into individual droplets. These devices include an immiscible two-phase system: an organic liquid, usually oil, that fills the channel and an aqueous liquid that breaks intro droplets when it is introduced into the system. Poisson statistics govern the ratio of organism encapsulation, determining the probability of isolating one or more cells per drop as a function of the starting concentration and occupied fraction of the droplets. See pg. 406, col. 1, para. 4 to col. 2, para. 1. Toxicology and antibiotic resistance have been tested in these devices, analyzing the growth rates inside drops (see pg. 407, col. 1, para. 3).
Therefore, it would have been obvious at the time of filing for a person of ordinary skill to have arrived at the claimed invention by combining the teachings of Handique and Perez for the following reasons. First, both references are in the same field of endeavor and teach the use of microfluidic chips to capture single microorganisms. Further, Perez teaches that it can be useful in many applications to design small, lateral chambers to trap individual microbial cells, based on their size, to make a variety of observations (e.g., identification by electron microscopy, testing of their resistance to antibiotics, etc.). It would be apparent to a person of ordinary skill that these single cell “traps” can be closed chambers with only a single opening connected to the channel that facilitates the flow of sample solution to each chamber. One would have been particularly motivated to do so, because Perez teaches that understanding bacterial phenotypes at the single-cell level is essential to develop beneficial applications, such as studying pathogenic disease, and microfluidics offers a versatile and fine tunable platform for conducting these experiments. One would have also recognized from Perez’ review that the design of such devices is highly versatile, and there are numerous design elements that have been adapted for a wide range of applications that have been met with success in the art, conferring a considerable amount of knowledge for an ordinary artisan to adapt such a platform. Therefore, the results of combining these teachings would have been reasonably predictable, one would have had a reasonable expectation for success. Hence, the combination would have been readily apparent and deemed to be a mere (A) combining of prior art elements according to known methods to yield predictable results (see MPEP 2143(I): Rationales to support rejections under 35 U.S.C. 103).
Regarding claim 2, Handique teaches that the inlet manifold additionally includes an inlet, wherein the inlet manifold (main channel) receives the sample from the inlet (see pg. 4, para. [0041]). Handique teaches that the outlet manifold of the cell capture system includes an outlet, wherein the outlet manifold egresses the filtered sample from the outlet (see pg. 5, para. [0047]).
Regarding claim 3, Handique identifies the inlet channel (branch channel) as part number “240” (see pg. 1, para. [0029]), and discloses a cell capture system comprising 3 inlet channels, as shown in FIG. 2 below:
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Regarding claim 4, Handique teaches a fluidic chip comprising a main channel (inlet manifold) fluidly connected to a plurality of branch channels (inlet channels) fluidly connected to a plurality of connection channels (pores) fluidly connected to a plurality of observation chambers (chambers), as discussed above.
Handique does not explicitly teach the width of the main channel (inlet manifold).
However, Handique teaches that the system may include a single inlet manifold (main channel) that feeds all the inlet channels (branch channels) (see pg. 4, para. [0043]), and the system may also include multiple inlet channels, each having a width of up to 200 micrometers, or, in one instance, a width of 50-100 micrometers (see pg. 4, para. [0039]). In one example, Handique teaches the cell capture system having up to ten inlet channels arranged in parallel which can be connected to a single inlet manifold (see pg. 8, para. [0063]). In view of FIG. 2 of Handique, the inlet manifold spans the width of the fluidic chip and is in connection with the inlet channels longitudinally. Note that a cell capture system comprising, for example, ten inlet channels with a width of 100 micrometers each would comprise a total width of at least 1 mm, and a cell capture system comprising, for example, ten inlet channels with a width of 200 micrometers would have a total width of at least 2 mm. Furthermore, a system comprising fewer inlet channels or narrower inlet channels within the ranges taught by Handique could accommodate additional spacing (e.g., the length of the pores) between the inlet channels and still have an inlet manifold with a width that is within the claimed range. Therefore, Handique teaches embodiments which reasonably include inlet manifolds (main channels) having a width of 1–2 mm.
In view of the instant specification, there is no discussion regarding the criticality of the main channel spanning a width of 1–2 mm. On the contrary, Example 5 of Applicant’s disclosure discusses an embodiment wherein “the width of the main channel can vary from 25 µm to 150 µm” (see pg. 20, para. [0089]), which is substantially smaller than the claimed range. However, in view of Handique’s disclosure, a person of ordinary skill would have recognized that fluidic chips may be scaled in size in accordance with their desired application and that the claimed range is within the scope of Handique’s teachings. Therefore, a width within the claimed range would have been obvious in view of Handique’s disclosure and could have been arrived at through no more than routine optimization.
Regarding claim 5, Handique teaches that the inlet channel (branch channel) preferably has a depth and/or width between 5-200 microns, but can alternatively have any suitable depth and/or width (see pg. 4, para. [0039]). Handique teaches that the inlet channel (branch channel) “preferably has a depth and width larger than the diameter of the cell of interest” (see pg. 4, para. [0039]). Examiner notes that Handique appears to use the term “depth” instead of “height” when describing the fluidic channels of the disclosure. Therefore, a person of skill would have recognized from Handique that the width and the height of the branch channel (inlet channel) may each be any value within the range of 5-200 microns (which overlaps with the claimed range), depending on the size of the cell of interest. Therefore, it would have been obvious to have designed the width and height of the branch channel to be 5-20 micrometers when analyzing a cell that is less than 5-20 micrometers, which is within the claimed ranges. Furthermore, Perez teaches that microfluidics offers high flexibility in terms of design, and developing smaller channels and adjusting them to the measurements of the studied bacteria allows the capture of bacteria according to their size and shape, as previously discussed.
Regarding claim 6, Handique teaches that cells of a predetermined size are preferably trapped within the chamber as the sample flows through the pores (connection channels) (see pg. 2, para. [0031]). Hence, Handique teaches the connection channel (pore) is configured to permit passage of the single sample (cell) into the chamber to be trapped. Furthermore, Perez teaches that microfluidics offers high flexibility in terms of design, and developing smaller channels and adjusting them to the measurements of the studied bacteria allows the capture of bacteria according to their size and shape, as previously discussed.
Regarding claim 8, Handique teaches that the chamber preferably has a length and width configured to retain an isolated cell, wherein the chamber is dimensioned to prevent cell egress from the chamber (see pg. 3, para. [0037]). Hence, Handique teaches the observation chamber is configured for trapping a single sample (isolated cell). Furthermore, Perez teaches that microfluidics offers high flexibility in terms of design and teaches that chambers having similar size to the bacterial cell have been successfully used to capture and trap single microorganisms, as previously discussed.
Regarding claim 9, Handique teaches the chamber “has a rectangular cross-section”, as shown in FIG. 3B (see pg. 3, para. [0037]).
Regarding claim 11, Handique teaches the chamber having a width of 50 micrometers and a length of 50 micrometers (see pg. 3, para. [0037]).
Regarding claim 13, Handique teaches that the inlet channel preferably has a length longer than the combined widths of the chambers and may extend to the edge of the substrate (fluidic chip) (see pg. 4, para. [0039]). Handique teaches that each pore includes a chamber (see pg. 1, para. [0029]), each chamber has a width of 5 to 200 microns (see pg. 3, para. [0037]), and each array includes multiple pores, such as 100 or 1000 pores (see pg. 2, para. [0036]). Note that the cell capture system includes one or more arrays, wherein the arrays are arranged in parallel (see pg. 2, para. [0033]). Therefore, the length of the fluidic chip may be estimated from the length of the inlet channel, the combined widths of the chambers, and the number of pores. For example, a fluidic chip comprising 100 pores and chambers 60-200 microns in width would be 6–20 mm in length, and a fluidic chip comprising 1000 pores and chambers 6-20 microns in width would also be 6–20 mm in length. Therefore, Handique teaches embodiments that reasonably include fluidic chips with a length of 6–20 mm.
Furthermore, Perez teaches that microfluidics offers high flexibility in terms of design and teaches that multiple channels can be added in parallel to create more versatile devices, as previously discussed.
In view of the instant specification, there is no discussion regarding the criticality of the fluidic chip spanning 6–20 mm in length. The specification discloses that an 8 mm length of microfluidic chip allowed capturing of 98% viable and culturable bacteria (see pg. 12, para. [0070]); however, the inventors also state that potential chip designs may include microchannels which vary in length in the range of 5 – 15 mm (see pg. 20, para. [0088]) and do not seem to attribute the results of the invention with the length of the chip. Therefore, in view of Handique and Perez, a person of ordinary skill would have recognized that fluidic chips may be scaled in size in accordance with their desired application and that the claimed range is within the scope of Handique’s teachings. Therefore, a length within the claimed range would have been obvious in view of Handique’s disclosure and could have been arrived at through no more than routine optimization.
Regarding claim 14, Handique teaches that polycarbonate can be used for the microfluidic chip’s substrate (see pg. 7, para. [0058]).
Claim(s) 7, 10 and 16 is/are rejected under 35 U.S.C. 103 as being unpatentable over Handique and Perez as applied to claims 1-6, 8-9, 11 and 13-14 above, and further in view of Lawrenz et al. (Geometrical effects in microfluidic-based microarrays for rapid, efficient single-cell capture of mammalian stem cells and plant cells, Biomicrofluidics, 2012 Jun;6(2):24112-2411217; previously cited), hereafter, “Lawrenz”.
Regarding claim 7, Handique teaches a fluidic chip comprising a main channel fluidly (inlet manifold) connected to a plurality of branch channels (inlet channels) fluidly connected to a plurality of connection channels (pores) fluidly connected to a plurality of observation chambers (chambers), as discussed above.
Handique does not explicitly teach the connection channel (pore) having a width ranging from 0.25-50 µm.
However, Perez teaches that microfluidics offers high flexibility in terms of design, and developing smaller channels and adjusting them to the measurements of the studied bacteria allows the capture of bacteria according to their size and shape, as discussed above.
Lawrenz, in the same field of endeavor, discloses a detailed numerical and experimental investigation into the optimization of hydrodynamic micro-trapping arrays for high-throughput capture of single polystyrene (PS) microparticles and three different types of live cells at trapping times of 30 min or less (see Abstract). Lawrenz teaches that microfluidic-based single cell isolation arrays utilize the principal of “paths of least flow resistance” to initially direct a single cell into a trap, which results in this lead cell “blocking” the dominant flow path and thereafter redirecting subsequent cells through the next “path of least flow resistance” containing similar traps (see pg. 3, para. 1). Lawrenz teaches that this particular design concept is well established in the literature and can successfully fill about 10%-80% of the traps with single particles or cells (see pg. 3, para. 1). However, scaling up to a higher density of traps is a significant challenge, as is adapting these devices to a wide variety of cell types (see pg. 3, para. 1). Lawrenz teaches that the devices must be optimized for each cell type and designed principally in accordance with the trap size to cell size ratio (see pg. 17, para. 1). In Lawrenz’ experiments, trapping channels (which are analogous to the pores taught by Handique) were designed for trapping 20 micrometer cells, wherein the channels had a width of 10 to 18 micrometers (see pg. 5, para. 2). However, in order to trap smaller cellular species on the order of 2 to 15 micrometers in diameter, trapping channels were decreased to 1 to 5 micrometers in width (see pg. 5, para. 4).
It would have been obvious at the time of filing for a person of ordinary skill in the art to have arrived at the claimed invention by combining the teachings of Handique, Perez and Lawrenz because each reference teaches microfluidic-based single cell isolation arrays for trapping and analyzing individual cells. One would have been motivated to do so, because Lawrenz teaches optimized strategies for isolating live cells, including smaller cellular species, for higher-throughput operations. One would have recognized from each reference that the dimensions of the branch channels (i.e., trapping channels, pores) must be optimized in accordance with the cell type of interest, and that the teachings of the these references could have been combined using known methods to achieve reasonably predictable results. There would have been a reasonable expectation for success, because Lawrenz teaches this particular design concept is well-established in the art and provides guidance and examples using cells and particles of various sizes and properties. Furthermore, one would have recognized from Perez that the design of microfluidic devices is highly versatile, and there are numerous design elements that have been successfully adapted for a wide range of applications. Hence, the combination would have been readily apparent and deemed to be a mere (A) combining of prior art elements according to known methods to yield predictable results (see MPEP 2143(I): Rationales to support rejections under 35 U.S.C. 103).
Regarding claim 10, Lawrenz teaches that in order to trap smaller cellular species on the order of 2 to 15 micrometers in diameter, trap depths were decreased to 10 micrometers (see pg. 5, para. 4), which is within the claimed range.
Regarding claim 16, Lawrenz’ experiments were conducted using mesenchymal stem cells (MSCs), human embryonic stem cells, and chlorella algae cells (see pg. 6). Therefore, it would have been obvious to have used a sample solution composed of eukaryotes, and to have adapted the size of the chip’s components (i.e., channels, chambers) accordingly.
Claim(s) 12 is/are rejected under 35 U.S.C. 103 as being unpatentable over Handique and Perez as applied to claims 1-6, 8-9, 11 and 13-14 above, and further in view of Chiu et al. (US 2016/0146823 A1; previously cited), hereafter, “Chiu”.
Regarding claim 12, Handique teaches the cross-section of the chamber can be substantially constant, wherein the cross-section is parallel to the broad face of the substrate (pg. 3, para. [0037]). Here, a person of ordinary skill would have recognized that shapes having a substantially constant cross-section parallel to their base include cylinders.
Chiu, in the same field of endeavor, teaches methods for isolating cells from a sample comprising introducing the sample into a microfluidic chip via a set of tubing, wherein the fluidic chip comprises at least one channel fluidly connected to the set of tubing, and at least one chamber fluidly connected to the at least one channel (see pg. 1, para. [0007]). Chiu further provides a system, method and apparatus for using single-analyte arrays with a fluidic sample, wherein the single-analyte array can comprise a plurality of wells or micro-wells configured so that no more than one analyte in a fluid sample will occupy a particular well (see pg. 4, para. [0054]). Chiu teaches that the fluidic sample comprising analytes can be introduced into the array and the analytes ( e.g., cells) can be partitioned so that at least 80% of the micro-wells of the device contain no more than one analyte (see pg. 4, para. [0054]). Chiu teaches the methods, systems, apparatuses and devices of the disclosure can be used in a wide variety of applications in biology and pathology for the separation and/or isolation of analytes (e.g., cancer cells, cancer stem cells, malaria infected erythrocytes, stem cells, fetal cells, immune cells, infected cells) from fluids (e.g., body fluids) for diagnosis and prognosis of disease, the isolation of single-celled parasites ( e.g., giardia, cryptosporidium) for water quality monitoring, and others (see pg. 4, para. [0056]). Chiu teaches that the chambers of the device can be wells which can trap single cells from a solution, and multiple wells can be arranged in parallel to form the device (see pg. 22, para. [0240]). Chiu teaches the device can comprise chambers with a variety of potential dimensions and shapes, wherein the dimensions and shapes can affect the parallel flow resistance trap design, and the well can be in the shape of a cylinder (see pg. 22, para. [0240]).
Chiu does not explicitly teach the diameter of the cylinder shaped chamber.
However, Handique teaches the chamber has a length and width of 5-200 microns, but can alternatively have any suitable dimension (see pg. 3, para. [0037]). Therefore, when combining the teachings of Handique and Chiu, a person of skill would have recognized that the cylindrical shaped chamber may have a diameter of 5-200 microns, which overlaps with the claimed range from 100-200 µm.
In view of the instant specification, there is no discussion regarding the criticality of the observation chamber having a diameter ranging from 100-300 micrometers or a having a cylindrical shaped chamber. Furthermore, there is no clear disclosure of what shape was used for the observation chambers in Applicant’s Examples. Example 1 states that the observation chambers “are dimensioned at 50 x 100 x 3.7 µm (WxLxH)” and “can be rectangular prism-shaped, pentagonal prism-shaped, heptagonal prism-shaped, octagonal prism-shaped, triangular prism-shaped, square prism-shaped, cylindrical prism-shaped and trapezoidal prism-shaped” (see pgs. 9-10, para. [0056]; Emphasis added). Hence, there is no indication that any of the results disclosed by the inventors were achieved using cylindrical shaped chambers, and there is no evidence to show that the claimed range or chamber shape were critical to the invention.
Therefore, it would have been obvious at the time of filing for a person of ordinary skill in the art to have arrived at the claimed invention by combining the teachings of Handique, Perez and Chiu, because each reference teaches methods and systems for isolating cells from a sample solution using a microfluidic chip comprising chambers configured to isolate individual cells. One would have been motivated to combine these teachings because both references teach such methods are useful for developing systems for cell capture and analysis which can be used in a wide variety of applications. A person of skill would have recognized from Chiu that the cell capture chambers of the system could be configured to have a cylindrical shape which would meet the design need of having a substantially constant cross-section as described by Handique. One would have also recognized that the dimensions of the chambers could be routinely optimized for the cell type of interest and the desired application and, in view of Handique, may have a diameter that is within the claimed range. Furthermore, one would have recognized from Perez that the design of microfluidic devices is highly versatile, and there are numerous design elements that have been successfully adapted for a wide range of applications. Hence, the combination would have been readily apparent and deemed to be a mere (A) combining of prior art elements according to known methods to yield predictable results (see MPEP 2143(I): Rationales to support rejections under 35 U.S.C. 103).
Claim(s) 15 is/are rejected under 35 U.S.C. 103 as being unpatentable over Handique and Perez as applied to claims 1-6, 8-9, 11, and 13-14 above, and further in view of Hsu et al. (US 2018/0243742 A1; previously cited), hereafter, “Hsu”.
Regarding claim 15, Handique teaches the cell capture system is preferably defined on a chip, more preferably a microfluidic chip (see pg. 2, para. [0029]). Handique teaches the substrate is preferably optically transparent (see pg. 7, para. [0058]). Handique teaches that the cell capture system can additionally include optical elements that function to facilitate imaging, such as the ability to adjust incoming light which can be defined by an imaging stage (see pg. 7, para. [0055]) that interfaces with the cell capture system, such that real-time flow through the cell capture system can be visualized (see pg. 15, para. [0103]). Handique teaches the stage preferably includes a retention mechanism that retains the cell capture system and is capable of retaining other imaging equipment, such as glass slides or cell culture plates (see pg. 14, para. [0101]).
Handique does not explicitly teach the fluidic chip comprising a glass bottom layer.
However, Perez teaches that while materials previously used in the pioneering of microfluidic devices were silicon and glass, silicon made them incompatible with microscopy studies. Moreover, glass and silicon are characterized by their fragility, difficult bonding protocols and high cost. Since the 1990’s, the most utilized material has been polydimethylsiloxane (PDMS) which has numerous beneficial properties, including optical transparency, gas permeability, flexibility, chemical inertness, biocompatibility, the ease of bonding and unbonding to other materials, and low cost. See pg. 396, col. 2, para. 2.
Hsu, in the same field of endeavor, teaches a microfluidic chip designed to function as a hydrodynamic shuttling chip device comprising an array of single-cell trapping units (see Abstract; pg. 1, para. [0001]). Hsu teaches that microfabricated devices for single cell trapping, based on their ability to accurately manipulate single cells, are advantageous, because they are high-throughput, low-cost and can reduce reagent consumption (see pg. 1, para. [0002]). Similar to Handique, Hsu teaches the device comprises an array of single-cell trapping units comprising incoming channels, trapping channels, and chambers in fluidic connection with each other (see pg. 1, paras. [0003]-[0007]). Hsu teaches that the microchannel device is made out of PDMS and is bounded to a glass substrate (see pg. 4, para. [0078]) to provide a finished “PDMS-on glass HSC device” (see pg. 4, para. [0079]). Hsu also discloses the capturing of images showing hydrodynamic structures and cells contained in the PDMS-on glass device using a microscope (see pg. 2, para. [0049]; pg. 4, para. [0077]). Hsu provides a detailed method for fabricating the device and achieving permanent bonding of the PDMS (top layer) and a glass slide (bottom layer) (see pgs. 3-4, para. [0073]).
It would have been obvious at the time of filing for a person of ordinary skill in the art to have arrived at the claimed invention by combining the teachings of Handique, Perez and Hsu, because each reference teaches the use of optically transparent substrates in microfluidic chips designed for single cell capture and observation. One would have recognized from Handique and Hsu the advantage of using a glass bottom layer to allow for the passage of light through the chip and would have been motivated to apply the methods of Hsu to have bound this element to the top layer of the chip. One would have also recognized from Perez that the PDMS layer would also ensure optical transparency and the ease of bonding with the glass substrate, while lowering the overall cost of the manufacturing process. One would have had a reasonable expectation of success, because Hsu discloses a method for fabricating and binding these materials together and the use of these materials for microfluidics is well known in the art. Hence, the combination would have been readily apparent and deemed to be a mere (A) combining of prior art elements according to known methods to yield predictable results (see MPEP 2143(I): Rationales to support rejections under 35 U.S.C. 103).
Claim(s) 17-18 and 20 is/are rejected under 35 U.S.C. 103 as being unpatentable over Handique, Perez and Hsu, as applied above, and further in view of Yang, et al. (A single-cell encapsulation method based on a microfluidic multi-step droplet splitting system, Chinese Chemical Letters, Volume 26, Issue 12, 2015, Pages 1450-1454; previously cited), hereafter, “Yang”.
Regarding claim 17, Handique teaches that cell sorting and viability maintenance is accomplished by utilizing suitably sized pores and by controlling the sample flow rate through the system (see pg. 2, para. [0030]).
Hsu teaches that for Design 2 of the microfluidic device, the sample solution was loaded into the microfluidic channel at a flow rate of 0.3 µl/min by using a syringe pump; then, 5 µl DMEM medium was loaded into the microfluidic channel at a flow rate of 0.3 µl/min (see pg. 4, para. [0076]). Therefore, Hsu teaches a) filling the fluidic chip with a growth medium and the sample solution at a flow rate of 0.018 ml/h, which falls within the claimed ranges.
Regarding step “b)”, Handique teaches the cell capture system to further include an isolation mechanism that functions to isolate cells within individual pores, wherein an isolation inlet is connected to the inlet channel (branch channel), wherein an “isolation material” is flowed through the isolation inlet into the inlet channel (branch channel) (see pg. 6, para. [0053]). Handique teaches that in the “the flow state”, the isolation material is a substantially viscous, immiscible liquid, such as oil (see pg. 6, para. [0054]). Hence, Handique teaches infusing oil into the fluidic chip to trap individual microorganisms (cells) inside the observation chamber.
Perez also teaches that droplet-based microfluidics are characterized for their isolation and confinement of a single bacteria or small populations into individual droplets. These devices include an immiscible two-phase system: an organic liquid, usually oil, that fills the channel and an aqueous liquid that breaks intro droplets when it is introduced into the system. Poisson statistics govern the ratio of organism encapsulation, determining the probability of isolating one or more cells per drop as a function of the starting concentration and occupied fraction of the droplets. See pg. 406, col. 1, para. 4 to col. 2, para. 1. Droplet-based microfluidics is the most recent and innovative approach among microfluidic devices, and droplet-based devices introduce the opportunity for the application of certain biomolecular techniques (see pg. 406, col. 2, para. 2). The development of devices based on droplets has led to a great advance in this field, giving the possibility of studying bacteria at a single-cell level (see pg. 410, col. 1, para. 2).
The references do not explicitly teach infusing oil by a syringe pump at a 0.001–0.1 ml/hr flow rate within 15–60 seconds.
Yang teaches that microfluidics can provide rapid, accurate, and cost-effective methods for single-cell analysis, and droplet microfluidics not only has the potential to compartmentalize individual cells into nanoliter or smaller droplets for biochemical analysis, but can also characterize the single-cell analysis with minimal sample dilution, higher sensitivity, shorter reaction time, higher throughput and no cross contamination (see pg. 1450, col. 1, para. 1). Yang teaches that single-cell encapsulation methods based on active encapsulation employ external forces which increase the complexity of manipulating the system without a corresponding increase in throughput, whereas passive hydrodynamic encapsulation methods utilize pressure-driven flow in a simple microfluidic configuration for generating aqueous droplets at the same time entrapping single cells into droplets (see pg. 1450, col. 2, para. 1). Yang’s study aims at developing a simple hydrodynamic method to encapsulate single cells in droplets with high efficiency and throughput (see pg. 1451, col. 1, para. 2).
Yang discloses that two types of microfluidic chips were fabricated, both consisting of flow-focusing water/oil phase channels and a multi-step droplet splitting geometry, in which a main channel was sequentially divided into two branch channels 3 times to form 8 parallel branches (see pg. 1451, col. 1, para. 3). Yang discloses syringe pumps were used to introduce the fluids into the microfluidic chip (see pg. 1451, col. 2, para. 2). Yang found that the droplet generation rate could be sharply raised by simply adjusting the flowrate (see pg. 1452, col. 2, para. 3). An improved single-cell encapsulation efficiency was obtained using a human blood cell suspension, with an aqueous phase flowrate of 0.2 µl/min and an oil phase flowrate of 0.5 µl/min (see page 1454, col. 1, para. 1). This is equivalent to an oil infusion flowrate of 0.03 ml/h, which is within the claimed range. Yang teaches this single-cell encapsulation technique has great potential for single cell analysis by integrating with other functional modules on the chip (see pg. 1454, col. 1, para. 2).
Regarding the limitation, wherein the oil-cutting by oil infusion is conducted “for a period of 15 – 60 seconds to trap 0 or 1 microorganism”, each reference above teaches the goal of single-cell encapsulation. Therefore, it would have been obvious to have conducted this step for whatever period was necessary to trap a single microorganism in each trap. A person of ordinary skill would have recognized that if the infusion was not conducted long enough (i.e., did not trap any microorganisms) or was conducted for too long (e.g., resulting in cell egress from the chambers or the trapping of more than one cell in each chamber) that this period could be easily adjusted to achieve the optimal results. Therefore, a person of ordinary skill could have arrived at the claimed time period through no more than routine optimization.
It would have been obvious at the time of filing for a person of ordinary skill in the art to have arrived at the claimed invention by combining the teachings of aforementioned references, because each reference teaches hydrodynamic encapsulation methods for isolating individual cells on microfluidic chips. One would have recognized that Handique suggests pumping oil into the microfluidic chip in order to isolate individual cells, while Yang further discloses methods of encapsulation using an oil phase. One would have been motivated to combine these teachings, because Yang teaches that microfluidic encapsulation methods using droplets to encapsulate single cells are simpler than other methods and have high efficiency and throughput. There would have been a reasonable expectation of success, because Handique also teaches the use of oil in a microfluidic chip to isolate cells, and Hsu provides specific flow rates for both cell suspensions and growth media in the aqueous phase. Therefore, one would have recognized that the elements of each disclosure could be combined using known methods, and the results of the combination would have been reasonably predictable.
Regarding claim 18, Hsu teaches that the cells were cultured in DMEM medium at 37°C in a humidified incubator (see pg. 4, para. [0075]). Hence, it would have been obvious to have incubated the sample solution in a temperature-controlled chamber at 37°C.
Regarding claim 20, Perez teaches the use of microfluidic devices to isolate individual bacterial cells, as discussed above. Yang teaches that the human blood cell encapsulation was carried out using a cell suspension with a concentration of 2.5x104 cells/µl. Hence, it would have been obvious to have used a sample solution comprising bacterial cells with a concentration of 2.5x104 cells/µl, which falls within the claimed range.
Claim(s) 19 is/are rejected under 35 U.S.C. 103 as being unpatentable over Handique, Perez, Hsu, and Yang, as applied above, and further in view of Son, et al. (Detecting cell-secreted growth factors in microfluidic devices using bead-based biosensors, Microsyst Nanoeng, 2017;3:17025; previously cited), hereafter, “Son”, and Eyer et al. (A Microfluidic Chip for the Versatile Chemical Analysis of Single Cells, J. Vis. Exp. (80), e50618, 2013; previously cited), hereafter “Eyer”.
Regarding claim 19, Handique teaches that priming the cell capture system may include introducing an initial buffer (e.g., 1% BSA+0.1% triton X in 1x PBS) and removing air bubbles from the system (see pg. 12, para. [0087]).
Similarly, Hsu teaches that preparing the microfluidic device included filling the microfluidic device with deionized water, soaking the device in deionized water to remove air bubbles, injecting 5% BSA in 1x PBS into the microfluidic channel, and subsequently incubating for 30 minutes to prevent immediate cell adhesion to the PDMS surface (see pg. 4, para. [0074]).
Handique and Hsu do not explicitly teach “immersing” the top layer in the PBS “overnight” or adding “500 µl” of the PBS to the bottom layer.
Son discloses the designing of a microfluidic device with parallel microchambers for the cultivation of cells and the detection of secreted growth factors (see pg. 2, col. 1, para. 1). Son discloses that the chambers were separated by a hydrogel barrier that allowed secreted factors to diffuse unimpededly, while preventing cells from crossing over into sensing channels (see pg. 2, col. 1, para. 1). Son teaches this system may be easily adapted to accommodate a variety of cell types and cell-secreted signals (see pg. 2, col. 1, para. 1). Son teaches the fabrication of the microfluidic chip included irreversibly bonding a PDMS layer and a glass layer (see pg. 3, col. 1, para. 2). Son teaches that all chambers of the microfluidic device were filled with PBS and stored for 2–5 days to remove any unpolymerized PEG prepolymer solution and to make the channel surfaces hydrophilic (see pg. 3, col. 1, para. 3).
Eyer, in the same field of endeavor, discloses a microfluidic device that enables the quantitative determination of intracellular biomolecules in multiple single cells in parallel, wherein the cells are passively trapped in the middle of a microchamber (see Abstract). Similar to Hsu and Son, Eyer teaches the microfluidic device is finalized by bonding the PDMS parts onto a glass slide (see pg. 3, Protocol A, step 1). Eyer teaches that the chip is filled with fluids after bonding, and the control layer inlets are filled with an isosmotic solution, such as PBS (see pg. 3, Protocol A, Step 3). Eyer also teaches the chip is washed out with PBS at a flow rate of 10 µl/min using a syringe pump before the device is ready for cell experiments (see pg. 3, Protocol A, Step 4).
Regarding the limitation, “the top layer is immersed in 1-10XPBS overnight”, Son, Eyer and Hsu all disclose the filling of microfluidic channels with PBS after bonding the top and bottom layers of the microfluidic chip. Son further teaches incubating the microfluidic chip for 2-5 days in PBS, which Son teaches is important for making the channel surfaces hydrophilic. Similarly, Hsu teaches incubating the device with PBS for 30 minutes helps prevent cell adhesion. Furthermore, Handique and Hsu both teach introducing PBS buffer in the microfluidic channels and also teach the removal of air bubbles from the chip by adding fluids, including 1x PBS, while Hsu also teaches the chip can be “soaked” to remove said air bubbles. Therefore, a person of skill would have envisaged from this disclosure that immersing the chip, including the PDMS top layer, in the 1x PBS for an extended period of time would achieve the same effect of making the microchannel surfaces hydrophilic and preventing cell adhesion. One would have also recognized that incubation periods ranging from 30 minutes to several days are result-effective, and therefore, one would have recognized that an incubation period of “overnight” would have been a convenient and effective timeframe when preparing the microfluidic chip.
Regarding the limitation “500 µl 1-10XPBS is added to the bottom layer”, Eyer teaches filling the completed chip with fluids, including PBS buffer, and the washing of the channels with said buffer using a syringe pump. While Eyer does not explicitly teach injecting “500 µl” of the solution, there is no evidence to suggest that this limitation was critical to the results achieved by the inventors. In view of the instant specification, the top layer “is immersed” in 1x PBS overnight and 500 µl of 1x PBS “is added” to the bottom layer “after assembling” the microfluidic chip and “prior to operation” (see pg. 11, para. [0064]). First, it should be noted that this passage from Applicant’s Examples is recited in the present tense, and it is therefore unclear whether this method step was reduced to practice. Regardless, there is no evidence of criticality regarding this limitation. Furthermore, it would appear that both the top layer and the bottom layer are immersed in the 1x PBS, because this step is “after assembling” the chip, wherein the these layers were bonded together (see pg. 10, para. [0059]). Hence, a person of skill would have recognized that a 6-20 mm microfluidic chip immersed in 1x PBS would reasonably already be exposed to more than 500 µl of 1x PBS. Therefore, the limitation of “adding” the PBS to the bottom layer is reasonably met by the combination of references, and it would have required no more than routine optimization for a person of ordinary skill to have arrived at this amount.
Therefore, it would have been obvious at the time of filing for a person of ordinary skill to have arrived at the claimed invention by combining the teachings of the references above, because each reference teaches the preparation and use of microfluidic devices for the isolation of single cells. One would have recognized from Handique, Hsu, Son and Eyer that PBS buffer is commonly used to prime microfluidic cell capture systems, and the preparation of such devices commonly include the injecting and pumping of isosmotic solutions (e.g., PBS) into the microchannels of said devices. One would have also recognized that such devices could be “soaked” in such fluids for the removal of air bubbles and would have immediately envisaged adding 1x PBS to both layers of the device and/or immersing the device in 1x PBS prior to operation. One would have been motivated to do so, because Hsu and Son teach that such steps are necessary to prevent cell adhesion and maintain hydrophilicity of the microchannels. Hence, the combination would have been readily apparent and deemed to be a mere (A) combining of prior art elements according to known methods to yield predictable results (see MPEP 2143(I): Rationales to support rejections under 35 U.S.C. 103).
Response to Arguments
Regarding the rejections under 35 U.S.C. 102, Applicant argues that amended claim 1 defines a different microfluidic architecture in which each observation chamber is an individual closed chamber connected only through a respective connection channel. Therefore, the observation chamber is structurally different from the flow-through chamber disclosed by Handique.
Applicant’s arguments with respect to the rejection(s) of claim(s) 1-3, 6, 8, 9, 11 and 14 under 35 U.S.C. 102 have been fully considered and are persuasive. Therefore, the rejection has been withdrawn. However, upon further consideration, a new ground(s) of rejection under 35 U.S.C. 103 is made in view of Handique and Perez, which is necessitated by Applicant’s amendment.
Regarding the rejections under 35 U.S.C. 103, Applicant argues that Handique is directed to a continuous-flow cell capture device in which each capture chamber is intentionally configured to remain fluidly connected to both an inlet channel and an outlet channel through a pore channel. In contrast, amended independent claim 1 adopts a fundamentally different technical approach, because after microorganisms are introduced into the observation chamber through the connection channel, oil is subsequently introduced to perform oil-liquid cutting at the connection channel, thereby terminating fluid communication between the observation chamber and the remaining fluidic network. Accordingly, a person having ordinary skill in the art would not have been motivated to modify Handique to arrive at the closed observation chamber recited in amended Claim 1, as doing so would fundamentally alter Handique's continuous-flow architecture. Furthermore, regarding claims 4, 5 and 13, the channel dimensions disclosed in Handique are defined for this continuous-flow channel architecture and are selected according to the size of the target cells. Handique neither teaches nor suggests that such dimensions may be freely modified while preserving the disclosed channel architecture and cell-capture configuration.
Applicant’s arguments with respect to claim(s) 1, 4-5 and 13 have been considered but are moot because the new ground of rejection does not rely on any reference applied in the prior rejection of record for any teaching or matter specifically challenged in the argument.
In the instant case, Perez teaches the trapping of bacteria in small side chambers and the use of oil-based droplets to isolate individual cells. Furthermore, Perez teaches the modification of microfluidic systems to accommodate cells according to their size and shape.
Regarding claims 7, 10 and 16 in further view of Lawrenz, Applicant argues that Handique and Lawrenz employ channel architectures fundamentally different from that of the present invention, and neither reference teaches or suggests that the channel dimensions disclosed in Lawrenz are applicable to the closed observation chamber architecture of the present invention.
Applicant’s arguments have been considered but are moot because the new ground of rejection does not rely on any reference applied in the prior rejection of record for any teaching or matter specifically challenged in the argument.
In the instant case, Perez teaches the trapping of bacteria in small side chambers and the use of oil-based droplets to isolate individual cells. Furthermore, Perez teaches the modification of microfluidic systems to accommodate cells according to their size and shape.
Applicant further argues that the Office Action established that a person of ordinary skill in the art would have modified the channel dimensions of Handique in view of Lawrenz while preserving the intended structures and functions of the respective references. The proposed modification can therefore only be derived from Applicant’s disclosure using impermissible hindsight reconstruction.
Applicant’s arguments have been fully considered but they are not persuasive.
First, Applicant’s arguments are, at least in part, moot, because in the new grounds of rejection, Perez is relied upon to show that microfluidic devices are highly flexibility and versatile in terms of design, and developing smaller channels and adjusting them to the measurements of the studied bacteria allows the capture of bacteria according to their size and shape.
Second, in response to applicant's argument that the examiner's conclusion of obviousness is based upon improper hindsight reasoning, it must be recognized that any judgment on obviousness is in a sense necessarily a reconstruction based upon hindsight reasoning. But so long as it takes into account only knowledge which was within the level of ordinary skill at the time the claimed invention was made, and does not include knowledge gleaned only from the applicant's disclosure, such a reconstruction is proper. See In re McLaughlin, 443 F.2d 1392, 170 USPQ 209 (CCPA 1971). In the instant case, Lawrenz teaches optimized strategies for isolating live cells, including smaller cellular species, for higher-throughput operations, and teaches channel dimensions that are in accordance with the size of the cells to be captured. Applicant’s argument fails to specifically point out how these proposed modifications can “only be derived from Applicant’s disclosure”.
Applicant argues that Chiu likewise does not teach or suggest a closed observation chamber connected only to a respective connection channel. Therefore, the Office Action has not established that a person of ordinary skill in the art would have modified Handique to arrive at the claimed subject matter.
Applicant’s arguments have been considered but are moot because the new ground of rejection does not rely on any reference applied in the prior rejection of record for any teaching or matter specifically challenged in the argument.
In the instant case, Perez teaches the trapping of bacteria in small side chambers and the use of oil-based droplets to isolate individual cells. Furthermore, Perez teaches the modification of microfluidic systems to accommodate cells according to their size and shape.
Applicant further argues that Hsu merely discloses bonding a PDMS microfluidic device to a glass substrate to facilitate fabrication and optical observation. However, Hsu employs a fundamentally different microfluidic architecture in which trapping channels, chambers, and bypass channels remain fluidically interconnected. Hsu's disclosure of a glass substrate is inseparable from its fundamentally different microfluidic architecture, a person of ordinary skill in the art would not have been motivated to modify the continuous-flow architecture of Handique to arrive at Applicant's claimed fluidic chip.
Applicant’s arguments have been fully considered but they are not persuasive.
First, Applicant’s arguments are, at least in part, moot, because in the new grounds of rejection, Perez is relied upon to show that microfluidic devices are highly flexibility and versatile in terms of design, and developing smaller channels and adjusting them to the measurements of the studied bacteria allows the capture of bacteria according to their size and shape into small side chambers to isolate individual cells.
Second, regarding Applicant’s statement that “Hsu’s disclosure of a glass substrate is inseparable from its fundamentally different microfluidic architecture”, Applicant is reminded that “The use of patents as references is not limited to what the patentees describe as their own inventions or to the problems with which they are concerned. They are part of the literature of the art, relevant for all they contain." In re Heck, 699 F.2d 1331, 1332-33, 216 USPQ 1038, 1039 (Fed. Cir. 1983) (quoting In re Lemelson, 397 F.2d 1006, 1009, 158 USPQ 275, 277 (CCPA 1968)). Further, “[a] reference may be relied upon for all that it would have reasonably suggested to one having ordinary skill in the art, including nonpreferred embodiments. Merck & Co. v. Biocraft Labs., Inc. 874 F.2d 804, 10 USPQ2d 1843 (Fed. Cir. 1989), cert. denied, 493 U.S. 975 (1989). See MPEP 2123. In the instant case, it is unclear what makes Hsu’s disclosure of a glass substrate “inseparable” from the rest of the disclosure. It should be noted that Perez, Handique, Tsu, Son and Eyer all teach glass substrates and/or the bonding of PDMS to a glass slide, and this appears to be a common technique in microfluidics. Therefore, Applicant’s argument fails to specifically point how Hsu’s architecture makes it inseparable.
Applicant further argues that Yang generates discrete droplets in a droplet-generation microfluidic device in which aqueous droplets are formed and transported by an oil phase. Handique employs a continuous flow-through architecture in which each chamber remains fluidly connected to a downstream outlet channel via a pore channel. Accordingly, simply introducing oil into Handique-as proposed in the Office Action would not produce the claimed oil-liquid cutting and trapping mechanism. Because Handique's chambers remain open to the downstream outlet, the aqueous phase would maintain fluid communication with the outlet channel through the pore channel. As a result, introducing oil into Handique's open, flow-through system would fail to create the isolated pico-chamber structure recited in the present application.
Applicant’s arguments have been considered but are, at least in part, moot because the new ground of rejection does not rely on any reference applied in the prior rejection of record for the specific “chamber structure” now recited in the amended claims.
In the instant case, Perez teaches the trapping of bacteria in small side chambers based on the size and shape of the cells and the use of oil-based droplets to isolate individual cells. Furthermore, Perez teaches the design of microfluidic devices to be highly flexible. Therefore, a person of skill could have arrived at the same chamber structure as now recited in the amended claims by modifying Handique, as discussed in the present rejections.
Applicant is also reminded that the test for obviousness is not whether the features of a secondary reference may be bodily incorporated into the structure of the primary reference; nor is it that the claimed invention must be expressly suggested in any one or all of the references. Rather, the test is what the combined teachings of the references would have suggested to those of ordinary skill in the art. See In re Keller, 642 F.2d 413, 208 USPQ 871 (CCPA 1981). In the instant case, a person of skill could have modified the design of the microfluidic system taught by Handique in order to practice the droplet-based isolation technique taught by Perez and Yang.
Regarding claim 19, Applicant argues that while the cited references (Son, Eyer, and Hsu) teach filling or washing internal microchannels with PBS to prime channels or prevent cell adhesion, none of these references teaches or suggests immersing the top layer in 1-l0x PBS overnight prior to operation as recited in Claim 19. In the context of the present invention, this specific overnight immersion of the top layer matrix is designed to pre-saturate the bulk polymer, thereby preventing localized medium evaporation inside the closed observation chambers during subsequent monitoring. Therefore, a person of skill would have lacked any motivation to employ PBS for the same purpose.
Applicant’s arguments have been fully considered but they are not persuasive.
In response to applicant's argument that the cited references employ PBS for purposes different from the present invention, the fact that the inventor has recognized another advantage which would flow naturally from following the suggestion of the prior art cannot be the basis for patentability when the differences would otherwise be obvious. See Ex parte Obiaya, 227 USPQ 58, 60 (Bd. Pat. App. & Inter. 1985). In the instant case, it would have been obvious to have immersed the microfluidic chip in PBS for a duration that is equivalent to “overnight” for the reasons discussed in the present rejection.
Regarding claim 20, Applicant argues that Yang teaches cell concentration parameters specifically tailored for human blood cell encapsulation in dynamic droplets, while Chiu merely genericizes microfluidic isolation to include bacteria without providing operational concentration parameters. Neither reference teaches, suggests, or provides any motivation to apply Yang's human blood cell concentration to Chiu's bacterial isolation, let alone to arrive at Applicant's claimed range of 5xl03- 5x104 CFU/ml. A person of ordinary skill in the art would recognize that human blood cells and bacteria differ fundamentally in size and volume by orders of magnitude, and Yang's concentration parameters for blood cells cannot be directly applied to bacteria. In the present invention, the claimed bacterial concentration range is specifically calibrated to operate synergistically with the blind-end geometry of claim 1 and the timed oil-liquid cutting mechanism of claim 17. Executing oil-liquid cutting within this precise concentration window ensures that single bacterial cells are isolated inside the closed observation chambers with high efficiency-consistently achieving 0 or 1 microorganism per chamber rather than causing overcrowding or leaving chambers empty.
Applicant’s arguments have been fully considered but they are not persuasive.
First, Applicant’s arguments are, at least in part, moot, because in the new grounds of rejection, the prior art of Chiu is not relied upon. Instead, Perez is relied upon to show that microfluidic devices are highly flexibility and versatile in terms of design, can have channels and side traps based on the size and shape of the bacteria to be isolated, and may incorporate droplet-based isolation techniques.
Applicant’s arguments fail to provide objective evidence as to why a person of skill would not be motivated to use the same concentration taught by Yang. Yang provides a result-effective variable for a concentration of cells for use in a droplet-based method for isolating individual cells. Therefore, it would have been obvious to have started with this concentration when testing a microfluidic chip designed for bacteria. Applicant seems to suggest that the “blind-end geometry of claim 1” is only compatible with bacteria at the recited concentrations. However, it should be noted that the sample solution of claim 1 includes bacteria, fungi, archaea, protists and eukaryotes (see claim 16). These structures also differ in size and volume, but are all within the scope of the method of using the same fluidic chip of claim 1, as recited in claim 17. It should be noted that the fluidic chip of claims 1 and 17 is not limited to any particular size or volume, nor are any of its internal structures. Therefore, a person of skill, particularly in view of Perez, would appreciate that these parameters may be modified based on the size and shape of the microorganism selected for isolation. See the present rejection for further discussion.
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.
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/DENNIS IGNATIUS ARMATO JR/Examiner, Art Unit 1651
/MELENIE L GORDON/Supervisory Patent Examiner, Art Unit 1651
