Office Action 7: Non-Final Rejection (2021-08-05)
Rejection types: 103 - Obviousness

CLAIMS:
Amendments to the Claims:

Please amend the claims as follows.

1\. (Currently Amended) A method for droplet formation for sample preparation for sequencing, the method comprising:
providing a first aqueous fluid in a first inlet channel on a fluidic chip, the first aqueous fluid comprising primers;
providing, via a second inlet channel on the fluidic chip by positive displacement pressure driven flow in the second inlet channel, a second aqueous fluid comprising nucleic acid molecules;
providing an immiscible carrier fluid in a main channel in the fluidic chip, wherein each of the inlet channels is in fluid communication with the main channel;
coalescing the first aqueous fluid with the second aqueous fluid in the main channel in the fluidic chip, thereby forming a plurality of droplets surrounded by the immiscible carrier fluid in the main channel;
applying a pressure to flow the immiscible carrier fluid in the main channel by positive displacement pressure driven flow in the main channel; and
annealing one of the primers to one of the nucleic acid molecules within one of the plurality of droplets in the main channel.

2\. (Original) The method of claim 1, wherein the pressure in the main channel is derived from pressure in a reservoir in fluid communication with the main channel.

3\. (Original) The method of claim 1, wherein the pressure in the main channel is applied by a single pressure source to the main channel.

4\. (Previously Presented) The method of claim 1, wherein each of the primers is linked to a binding moiety.

5\. (Previously Presented) The method of claim 4, wherein the binding moiety is biotin and the method includes capturing the annealed one of the nucleic acid molecules to a streptavidin- coated surface.

6-11. (Cancelled) 710

12\. (Previously Presented) The method of claim 1, wherein the aqueous fluids further comprise reagents for conducting an amplification reaction.

13\. (Currently Amended) The method of claim 12, wherein the amplification reaction is a PCR reaction and the method comprises performing the PCR reaction in the one of the droplets to yield PCR products and breaking the droplets to release the PCR products from the droplets.

14-18. (Cancelled) 1517

19\. (Previously Presented) The method of claim 1, wherein the first and second inlet channels are in fluid communication with the main channel at a junction on the fluidic chip where the plurality of droplets are formed in the main channel.

20\. (Previously Presented) The method of claim 19, wherein the junction comprises a fluidic interconnect apparatus that forms a seal between the first inlet channel and the main channel.

21\. (Previously Presented) The method of claim 19, wherein the junction defines a nozzle and the nozzle includes at least one oil line that provides a constant flow of the carrier fluid into the nozzle while the plurality of droplets are formed.

22\. (Previously Presented) The method of claim 21, wherein the nozzle design inhibits the first aqueous fluid from wetting surfaces of the nozzle or the main channel.

23\. (Previously Presented) The method of claim 21, wherein the flow of carrier fluid into the nozzle provides complete isolation of the nozzle from the aqueous phase of the first aqueous fluid.

24-30. (Cancelled) 2529

31\. (Previously Presented) The method of claim 1, wherein the immiscible carrier fluid comprises an oil.

32\. (Previously Presented) The method of claim 31, wherein the oil comprises a fluorocarbon oil.

33\. (Previously Presented) The method of claim 31, wherein the oil comprises a surfactant.

34\. (Previously Presented) The method of claim 33, wherein the surfactant is a fluorosurfactant.

35\. (Previously Presented) The method of claim 33, wherein the pressure to flow the immiscible carrier fluid in the main channel by positive displacement pressure driven flow is applied using a pump.
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OFFICE ACTION:
DETAILED 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 July 21, 2021 has been entered.
Notice of Pre-AIA  or AIA  Status
The present application is being examined under the pre-AIA  first to invent provisions.
Preliminary Remark
Claims 6-11, 14-18, and 24-30 are canceled.
Claim Rejections - 35 USC § 103
The rejection of claims 1-3, 5, 12, 13, 19, 31, and 35 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Nobile et al. (US 2005/0227264 A1, published October 13, 2005, priority January 28, 2004), made in the Office Action mailed on April 22, 2021 is withdrawn.
The rejection of claim 4 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Nobile et al. (US 2005/0227264 A1, published October 13, 2005, priority January 28, 2004) as applied to claims 1-3, 12, 13, and 19 above, and further in view of Vogelstein et al. (US 2011/0059435, published March 2011, priority February 7, 2005), made in the Office Action mailed on April 22, 2021 is withdrawn.
The rejection of claims 20-23 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Nobile et al. (US 2005/0227264 A1, published October 13, 2005, priority January 28, 2004), made in the Office Action mailed on April 22, 2021 is withdrawn.
The rejection of claims 32-34 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Nobile et al. (US 2005/0227264 A1, published October 13, 2005, priority January 28, 2004) as applied to claims 1-3, 12, 13, and 19 above, and further in view of Dorfman et al. (Analytical Chemistry, 2005, vol. 77, pages 3700-3704), made in the Office Action mailed on April 22, 2021 is withdrawn.
Rejection – New Grounds, Necessitated by Amendment
The following is a quotation of pre-AIA  35 U.S.C. 103(a) which forms the basis for all obviousness rejections set forth in this Office action:
(a) A patent may not be obtained though the invention is not identically disclosed or described as set forth in section 102, if the differences between the subject matter sought to be patented and the prior art are such that the subject matter as a whole would have been obvious at the time the invention was made to a person having ordinary skill in the art to which said subject matter pertains. Patentability shall not be negatived by the manner in which the invention was made.
Claims 1-3, 5, 12, 13, 19, 31, and 35 are rejected under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Quake et al. (US 2002/0058332 A1, published May 16, 2002).
With regard to claims 1, 12, 13, and 19 Takahashi discloses a method of forming droplets for sample preparation for sequencing (“present invention provides methods of using inline automated platform of the invention … to prepare, amplify, detect … and/or optionally select for further analysis (e.g., sequence) genomic material isolated…”, section [0038]), the method comprising:
providing a first aqueous fluid in a first inlet channel, the first aqueous fluid comprising primers (see Figure 1A, the first inlet is channel carrying the primers A/B and Markers A/B);
providing via a second inlet channel, a second aqueous fluid comprising nucleic acid molecules (see Figure 1A, where gene is provided);
providing an immiscible carrier in a main channel in a fluidic chip (see element 6, Figure 1A, where “carrier fluid” is provided; “a sample plug is surrounded by an immiscible nonaqueous fluid (e.g., mineral oil)”, section [0050]), wherein each of the inlet channels is in fluid communication with the main channel (see where the inlet containing the gene and the inlet containing the primer are connected to the channel where the carrier liquid flows (element 5);
coalescing the first aqueous fluid with the second aqueous fluid in the main channel in the fluid chip thereby forming a plurality of droplets surrounded by the immiscible carrier fluid in the main channel (see the gene primer become coalesced into a single emulsion, Figure 1A, area 10; “each sample droplet is further prepared at a junction, e.g., a T-shaped junction (10) to form a sample plug by being mixed with a primer plug comprising amplification reagents (e.g., primer(s), nucleotide, polymerase, etc).”, section [0084]);
applying pressure to flow the immiscible carrier fluid in the main channel (“controlling device … such as pumps, valves … may be used to control the movement and timing of each sample plug and/or DNA sample plug .. such controlling devices are well known to those skilled in the art”, section [0115]); and
annealing one of the primers to one of the nucleic acid molecules within one of the plurality of droplets (“mixed with amplification reagents to form a sample plug, it is drawn along the microfluidic inline reaction channel into an amplification area of a device … only DNA sample plugs will comprise DNA that will be amplified”, section [0085]; “low temperature area (13) may be set to a temperature appropriate to effectuate annealing”, section [0088]).
While Takahashi explicitly disclose that carrier fluid is provided for flowing the plugs in the channels, the artisans do not explicitly disclose that the carrier fluid is pressure driven from a reservoir in communication with the main channel through which the carrier fluid flows (claim 2), or that the pressure is applied by a single pressure source to the main channel (claim 3).
Takahashi et al. do not teach that a positive displacement pressure driven flow is employed for providing a second aqueous fluid through the second inlet channel.
While Takahashi explicitly teach and suggests that the amplification products of the invention can be subject to sequencing analysis (“microfluidic devices and instruments of the invention are intended to work together to provide an automated inline platform for either or both PCR and waveform profiling methods and optionally, e.g., sequencing analysis”, section [0047]), the artisans do not explicitly teach that primers are linked to a binding moiety (claim 4), or that the binding moiety is biotin and the method includes capturing the annealed nucleic acid molecules to a biotinylated surface (claim 5).
Quake et al. teach the use of positive pressure driven flow to flow an immiscible oil in a main channel, and generating droplets via first and second inlet channels attached to said main channel, wherein said inlet channels are also taught as being positive pressure driven:
“the device of the invention comprises at least two inlet regions, each connecting to the main channel at a droplet extrusion region.  In particular, the device may comprise a first inlet region in communication with the main channel at a first droplet extrusion region, and a second inlet region in communication with the main channel at second droplet extrusion region.  A fluid containing a first biological material may pass through the first inlet region so that droplets of the fluid containing the first biological materials are sheared into the main channel.  A fluid containing a second biological material may pass through the second inlet region so that droplets of the fluid containing the second biological material are sheared into the main channel.  In various aspects, the droplets of the first material may mix or combine with the droplets of the second biological material, and the first and second biological material may interact with each other upon mixing.” (section [0018])
“An inlet region may have an opening such as in the floor of the microfabricated chip, to permit entry of the sample into a device.  The inlet region may also contain a connector adapted to receive a suitable piece of tubing ….through which a sample may be supplied … Such an arrangement facilitates introducing the sample solution under positive pressure in order to achieve a desired pressure at the droplet extrusion region” (section [0081])
“device of the invention may have an additional inlet region, in direct communication with the main channel … through which a pressurized stream or ‘flow’ of a fluid is introduced into the main channel.  Preferably, this fluid is one which is not miscible with the solvent or fluid of the sample …” (section [0082])
“the invention use pressure drive flow control, e.g., utilizing valves and pumps, to manipulate flow of cells virions, particles, molecules, enzymes or reagents in one or more directions and/or into one or more channels of a microfluidic device” (section [0099])
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was made to combine the teachings of Takahashi et al. and Quake et al., thereby arriving at the invention as claimed for the following reasons.
As discussed above, Takahashi et al. already disclose a method and a microfluidic device comprising a plurality of sample and reagent inlets and an inlet for an immiscible carrier fluid into a main channel for performing continuous generation of emulsion products and PCR.
While Takahashi et al. do not explicitly disclose that the samples and PCR reactants or the oil are provided via a pump that generates a positive pressure, said one of ordinary skill in the art would have recognized that other prior art known pressure driven system utilized for generating droplets in a channel would have worked equally well.  Such a capability was demonstrated by Quake et al. who also teach generation of emulsion droplets comprising PCR reaction mixture in a flowing oil, wherein the artisans explicitly teach the use of pumps which provide a positive pressure flow to oil and the respective reagents from inlets in order to generate emulsion droplets in the main channel (see above).
Therefore, one of ordinary skill in the art would have been motivated to combine the teachings of Takahashi et al. and Quake et al., thereby arriving at a method of performing the method of Takahashi et al. on a system in which the oil and the reagents are provided into the main channel via a pump that produces positive pressure so as to generate emulsion droplets.
In KSR International Co v. Teleflex Inc, the supreme court stated that, “A person of ordinary skill in the art is also a person of ordinary creativity, not an automation” (82 USPQ2d at 1397) and that “in many cases a person of ordinary skill will be able to fit the teachings of multiple patents together like pieces of a puzzle” and take into account, “the inference and creative steps that a person of ordinary skill in the art would employ” (82 USPQ2d at 1396).
Lastly, it is respectfully submitted that such “fitting” of the teachings would not have been “beyond [the] ... skill [level]” of one of ordinary skill in the art
In KSR, the Supreme Court stated:
“When a work is available in one field of endeavor, design incentives and other market forces can prompt variations of it, either in the same field or a different one.  If a person of ordinary skill can implement a predictable variation, 103 likely bars its patentability.  For the same reason, if a technique has been used to improve one device, and a person of ordinary skill in the art would recognize that it would improve similar devices in the same way, using the technique is obvious unless its actual application is beyond his or her skill.” (page 13, emphasis added).
Therefore, the invention as claimed is deemed prima facie obvious over the cited references.Response to Arguments:
While the instant rejection is based on a newly applied reference, Applicants’ arguments presented in the Amendment received on July 21, 2021 are addressed herein to the extent applicable in desire of compact prosecution.
Applicants traverse the rejection (page 5, Response).
Applicants contend that the teachings of Takahashi directed toward the use of “sipping” pressure (i.e., negative pressure) cannot teach the presently amended claim limitation of reagents being delivered and driven through the main channel via positive pressure.
Applicants then contend that Nobile does not bring aqueous template nucleic acid and aqueous amplification reagents on-chip via separate inlet channels (page 7, Response).
This argument is no longer applicable because Nobile reference is not relied upon.
However, it is noted herein that Nobile reference was relied upon for teaching the use of a positive pressure driven flow for generating emulsion droplets in a channel.  Therefore, Applicants’ arguments cannot be deemed persuasive as they are arguing teaching which is not relied upon from Nobile in the present rejection.
Regardless of this finding, as discussed above, Quake et al. teach a method of utilizing positive pressure to move a non-aqueous medium (i.e., oil) in a main channel of a microfluidic chip, as well as using positive pressure to introduce at least two different inlets, each of which comprising immiscible medium (i.e., aqueous) into the main channel, and coalescing them together to mix the two contents therein.
Therefore, one of ordinary skill in the art would have recognized that the method disclosed by Takahashi et al. that provide aqueous solution via sipping (i.e., suction) could also be performed via a positive pressure driven system as taught by Quake et al.
Given that both system disclosed by Takahashi et al. and Quake et al. successfully generated emulsion droplets within a channel flowing with an immiscible carrier medium, one of ordinary skill in the art would have expected that the use of positive pressure driven system (of Quake et al.) in the method of Takahashi et al. would have yielded the same predictable outcome.
In KSR, the Supreme Court particularly emphasized “the need for caution in granting a patent based on the combination of elements found in the prior art,” Id. at 415, 82 USPQ2d at 1395, and discussed circumstances in which a patent might be determined to be obvious. Importantly, the Supreme Court reaffirmed principles based on its precedent that “[t]he combination of familiar elements according to known methods is likely to be obvious when it does no more than yield predictable results.” Id. at 415-16, 82 USPQ2d at 1395. The Supreme Court stated that there are “[t]hree cases decided after Graham [that] illustrate this doctrine.” Id.  at 416, 82 USPQ2d at 1395. (1) “In United States v. Adams, . . . [t]he Court recognized that when a patent claims a structure already known in the prior art that is altered by the mere substitution of one element for another known in the field, the combination must do more than yield a predictable result.”
Therefore, Applicants’ arguments are not found persuasive and the rejection is maintained.
Claim 4 is rejected under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Quake et al. (US 2002/0058332 A1, published May 16, 2002), as applied to claims 1-3, 12, 13, and 19 above, and further in view of Vogelstein et al. (US 2011/0059435, published March 2011, priority February 7, 2005).
The teachings of Takahashi et al. and Quake et al. have already been discussed above.
Takahashi, while explicitly suggesting that their disclosed method be for amplification, detection, as well as for sequencing reaction, do not explicitly all possible sequencing means known at the time the invention was made.
Consequently, the artisans do not explicitly disclose all possible means of sequencing reactions, such as those involving primers which are linked to a biotin (claim 4).
Vogelstein et al. disclose a method of amplifying nucleic acids in emulsion particles (region of analyte DNA molecules is amplified using a high fidelity DNA polymerase … Microemulsions comprising said first amplicons and reagents beads are formed”, section [0008]), wherein the primers are biotinylated (“common oligonucleotide … was synthesized with a dual biotin group … coupled to streptavidin-coated magnetic beads”, section [0058]).
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was made to combine the teachings of Takahashi et al. and Quake et al. with the teachings of Vogelstein et al., thereby arriving at the invention as claimed for the following reasons.
As stated above, Takahashi et al. explicitly teach that the amplification products in the emulsion particles of their invention can be employed for further analysis such as amplifying, detecting, and sequencing (see section [0030]), and while the artisans did not explicitly list all possible such “further analysis”, said one of ordinary skill in the art would have had a reasonable expectation of success that any of amplification and analysis means known in the art would have been equally useful and beneficial in the art of molecular analysis/diagnostics.
Given that Vogelstein et al. teach a method of producing emulsion particles comprising therein, amplification products bound on a bead via use of bead-bound primers for the purpose of detecting nucleotide sequence variations, said one of ordinary skill in the art would have been reasonably motivated to adopt the teachings of Vogelstein et al. into the method disclosed by the combination of Takahashi et al. and Quake et al., thereby arriving at the claimed method of generating emulsion particles comprising bead-bound primers and reagents, and template nucleic acids produced from the coalescing means taught by Takahashi et al.
In KSR, the Supreme Court particularly emphasized “the need for caution in granting a patent based on the combination of elements found in the prior art,” Id. at 415, 82 USPQ2d at 1395, and discussed circumstances in which a patent might be determined to be obvious. Importantly, the Supreme Court reaffirmed principles based on its precedent that “[t]he combination of familiar elements according to known methods is likely to be obvious when it does no more than yield predictable results.” Id. at 415-16, 82 USPQ2d at 1395. The Supreme Court stated that there are “[t]hree cases decided after Graham [that] illustrate this doctrine.” Id.  at 416, 82 USPQ2d at 1395. (1) “In United States v. Adams, . . . [t]he Court recognized that when a patent claims a structure already known in the prior art that is altered by the mere substitution of one element for another known in the field, the combination must do more than yield a predictable result.”
Therefore, the invention as claimed is deemed prima facie obvious over the cited references.
Claims 20-23 are rejected under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Quake et al. (US 2002/0058332 A1, published May 16, 2002), as applied to claims 1-3, 12, 13, and 19 above, and further in view of Nobile et al. (US 2005/0227264 A1, published October 13, 2005, priority January 28, 2004).
The teachings of Takahashi et al and Quake et al. have already been discussed above.
Takahashi et al. do not explicitly disclose that the junction comprises a fluidic interconnect apparatus that forms a seal between the inlet and the main channel (carrying the oil) (claim 20), or that junction defines a nozzle which includes an oil line that provides a constant oil flow into the nozzle (claim 21), wherein the design prevents the nozzle or the main channel from becoming wet with the aqueous fluid (claim 22) or that the carrier fluid into the nozzle provides complete isolation of the nozzle from the aqueous phase (claim 23).
While Takahashi et al. do not explicitly disclose the above-discussed features of the junction and nozzle, these features are deemed known and prima facie obvious to one of ordinary skill in the art at the time the invention was made based on the combination of the teachings of Takahashi et al., Quake et al., and Nobile et al. for the reasons discussed below.
Nobile et al. provides the below configuration which graphically represents a junction where the aqueous reagents are disposed into the main channel which carries the oil, which is similar to the configuration disclosed by Takahashi et al.:
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1015
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Merriam-Webster defines “nozzle” a short tube with a taper or constriction used to speed up or direct a flow of fluid (see attached)1.
Therefore, the main channel has a nozzle 406 (the narrow portion) found on the main channel wherein the oil flows through and the upper reservoir (412) (or first inlet) is in communication with the main channel at a junction at which the nozzle is found and the droplets are formed therefrom.
With regard to claim 20, the junction formed between the inlet and the nozzle of the main channel is configured to generate a plurality of droplets and while the artisans do not explicitly state that there is a seal between this inlet channel and the main channel, it would have been an obvious to one of ordinary skill in the art to make sure that there was a tight seal where the inlet and main channel intersected so as to contain the emulsion oil flowing through the main channel without escape and the reagents of the reaction flow into the main channel without any escape into a gap therebetween.
With regard to claim 21, the left-side of the narrow junction (i.e., nozzle) is considered to be an oil line and it feeds the carrier oil to the nozzle.
With regard to claim 22, the nozzle design inhibits the first aqueous fluid from wetting the surface of the nozzle of the main channel.  This is because the artisans explicitly state that at the narrow junction (i.e., nozzle), the aqueous reagents from the inlet is introduced and shears off in to a plurality of droplets and the formed droplets continue their travel in the main channel surrounded by the carrier oil (sections [0079] and [0080]).
With regard to claim 23, the nozzle is surrounded by the oil flowing there-through and therefore, isolated from the aqueous phase entering from the inlet.
Therefore, the invention as claimed is deemed prima facie obvious over the cited references.
Claims 32-34 are rejected under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Quake et al. (US 2002/0058332 A1, published May 16, 2002), as applied to claims 1-3, 12, 13, and 19 above, and further in view of Dorfman et al. (Analytical Chemistry, 2005, vol. 77, pages 3700-3704).
The teachings of Takahashi et al. and Quake et al. have already been discussed above.
Takahashi et al., while explicitly teach performing PCR in aqueous droplets in surrounded by an immiscible oil medium, do not teach every possible such medium or variations known in the art.  Consequently, the artisans do not explicitly teach that the oil is fluorinated (claim 32), or that the oil comprises fluorosurfactant (claims 33 and 34).
Dorfman et al. disclose a method of performing a continuous PCR reaction in an aqueous droplets surrounded by an immiscible oil medium (“[p]erformign continuous flow PCR for high-throughput clinical and quantitative applications requires encapsulating the sample and preventing contamination between them … this can be accomplished in two ways: either the sample can be a droplet entrained in an immiscible solvent …”, page 3700, 2nd column).
Dorfrman et al. disclose the use of fluorinated oil as the immiscible medium when performing the continuous emulsion PCR (“an improvement system for performing continuous flow PCR using droplets in an immiscible fluorinated solvent …”, page 3701, 1st column, 2nd paragraph; “primed with either the bulk fluorinated oil FC-40”, page 3701, 1st column, 3rd paragraph) or oil comprising a fluorosurfactant (“or FC-40 containing various amounts of fluoro alcohol surfactant”, page 3701, 1st column, 3rd paragraph).
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was made to combine the teachings of Takahashi et al. and Quake et al. with the teachings of Dorfman et al., thereby arriving at the invention as claimed for the following reasons.
Dorman et al. discuss potential problems associated with performing emulsion droplet PCR in an immiscible medium:
“wall treatment [of the channel] are necessary to prevent adsorption of DNA to the walls and subsequent transfer between samples … wall contamination problem is a significant drawback … bubbles in the stream may introduce irregularities in the thermal history of the droplets, making the technique less attractive … flowing droplet system does not suffer from either of these drawbacks … However, the initial work on this system suffered from contamination between droplets” (page 3701, 1st column, 1st paragraph).
In solving the drawback so identified, Dorman et al. disclose that immiscible solvent, “prevent[ed] the droplet breakage … thereby eliminating the contamination between droplets” (page 3701, 1st column, 2nd paragraph), producing “adequately reproducible amplification between different drops without any detectable contamination between them” (page 3701, 1st column, 2nd paragraph).
Based on such findings, one of ordinary skill in the art would have been motivated to utilize a fluorinate immiscible oil or oil comprising fluorosurfactants into the teachings of Takahashi et al. and Quake et al., for the same recognized benefit of producing reproducible amplification reaction between emulsion droplets as well as preventing contamination therebetween.
Given that Takahashi et al., Quake et al. and Dorfman et al. performed emulsion amplification reactions in a flowing oil medium, said one of ordinary skill in the art would have also expected that addition of fluorosurfactants or using fluorinated oil in the method of Takahashi et al. would have yielded the same predictable benefit as observed in Dorman et al.
In KSR, the Supreme Court particularly emphasized “the need for caution in granting a patent based on the combination of elements found in the prior art,” Id. at 415, 82 USPQ2d at 1395, and discussed circumstances in which a patent might be determined to be obvious. Importantly, the Supreme Court reaffirmed principles based on its precedent that “[t]he combination of familiar elements according to known methods is likely to be obvious when it does no more than yield predictable results.” Id. at 415-16, 82 USPQ2d at 1395. The Supreme Court stated that there are “[t]hree cases decided after Graham [that] illustrate this doctrine.” Id.  at 416, 82 USPQ2d at 1395. (1) “In United States v. Adams, . . . [t]he Court recognized that when a patent claims a structure already known in the prior art that is altered by the mere substitution of one element for another known in the field, the combination must do more than yield a predictable result.”
Therefore, the invention as claimed is deemed prima facie obvious over the cited references.
Conclusion
No claims are allowed.
Inquiries

APPLICANT REBUTTAL: Remarks
With entry of this amendment, claims 1-4, 12, 13, 21-23, and 3 1-35 are pending, with claim 1 being the only independent claim. Claims 1-4 and 21 are amended.
Support for the amendments may be found throughout parent U. S. application No.
11/803,104 as-filed, for example, at least at lines 32-33 on page 19, line 18 on page 22 through line 14 on page 23, and in FIG. 4.
No new matter is added.
Rej
ections under 3 5 U.
S.
C. 103
The Office Action on page 4 states that claims 1-3, 5, 12, 13, 19, 31, and 35 are rejected under §103 over U. S. Pub. 2006/0257893 to Takahashi in view of U. S. Pub. 2002/0058332 to Quake. On page 12, claim 4 is so rejected over Takahashi, Quake, and U.S. Pub. 2011/0059435 to Vogelstein. On page 14, claims 20-23 are so rejected over Takahashi, Quake, and U.S. Pub.
2005/0227264 to Nobile. On page 17, claims 32-3 4 are so rejected over Takahashi, Quake, and Dorfman, 2005, Anal Chem 77:3700-3704.
Applicant submits that the pending, amended claims are not obvious.
The Patent Office must establish three basic criteria in order to establish a prima facie case of obviousness. There must be a suggestion to modify a reference or to combine references to lead one of ordinary skill in the art to the claimed invention; the cited references must convey a reasonable expectation of success in arriving at the claimed invention; and the suggestion to modify the references must be found in the prior art, not in the applicant's disclosure. In re Vaeck, 947 F.2d 488 (Fed Cir. 199]). In addition, an obviousness rejection cannot be established in the first place unless each claim element is disclosed in the cited art. In re Royka, 490 F. 2d 98] (CCPA 1974).
Claim 1 is amended to the embodiments represented by FIG. 4. As amended, claim 1 recites:
1.    A method for droplet formation for sample preparation for sequencing, the method comprising:
providing a first aqueous fluid in a first inlet channel, the first aqueous fluid comprising biological material;
providing, via a second inlet channel, a second aqueous fluid comprising primers; coalescing the first aqueous fluid with the second aqueous fluid to form an aqueous sample fluid;
providing an immiscible carrier fluid via at least one oil supply line; applying a pressure to flow the immiscible carrier fluid into a nozzle disposed within the oil and into a main channel within the nozzle; and
adjusting pressure in one or more of the inlet channels, thereby to produce a plurality of droplets comprising the aqueous sample fluid in the main channel.
Features of claim 1 are shown in FIG. 3 and FIG. 4, reproduced in-part here:
Notably, as shown, the claimed method includes applying a pressure to flow the immiscible
carrier fluid into the nozzle
. The nozzle is disposed within oil. The oil flows into a main channel within the nozzle. Pressure adjustments at the inlet channels produces droplets of the aqueous sample fluid in the main channel as shown in the figures above.
The depicted apparatus and droplet formation process cannot be found among the asserted prior art references. Claim 1 cannot be said to be obvious over any combination of Takahashi, Quake, Vogelstein, Nobile, and Dorfman because none of those references show an apparatus in which the ferrule of a nozzle is immersed in, and surrounded by, oil such that droplets are formed by flowing into the nozzle without ever letting any of the aqueous phase touch any surface (interior or exterior) of the nozzle through which the main channel extends.
Takahashi reports a device for preparing and detecting genomic material. Takahashi, [0030]. The device includes a sipper to pick up sample and a microfluidic inline reaction channel that runs through a reagent assembly area, temperature-controlled amplification, and a detection area. Id., [0040]. Detecting includes aspirating a sample droplet into the reaction channel, mixing the sample droplet with a primer plug to form a sample plug, and then heating and cooling for amplification and subsequent detection. Id, [0052]. Takahashi says nothing of a nozzle disposed in oil, where oil is flowed into the nozzle to form droplets in a manner such that the nozzle is NEVER wetted with aqueous liquid.
Quake and Nobile actually both operate by a principle quite opposite of what is claimed.
FIG. 22 in Quake
FIG. 4 in Nobile
In Quake and Nobile it is quite clear that aqueous fluid emerges from an inlet channel into an oil-filled main channel. Neither Quake nor Nobile shows a channel within a nozzle, with the nozzle sitting in oil, with oil being pumped in such that oil flows into the nozzle forming aqueous droplets within a channel in the nozzle. The oil channels in Quake and Nobile do not have any feature or terminus that matches the claimed nozzle or ferrule.
Unlike Quake and Nobile, the invention uses a nozzle that juts out, into the oil. Pressure- driven flow (look again at FIGs. 3 & 4) causes: the oil to flow into the nozzle; droplets to form at the ferrule and flow into the nozzle; and isolation of the nozzle and ferrule completely from the aqueous phase. The as-filed application describes benefits of the invention that are not provided by Quake or Noble. For example, the as-filed application reports that
This nozzle design eliminates the surface wetting issues surrounding the quasi.-21) flow associated with typical nmicro fluidic nozzles trade using soft lithography or oilher standard microfluidic chip manufacturing techniques. This is because the nozzle design is fuilly 3-dimensional, resulting is a complete isolation of the noz'zle section from the continuous aqueous phase. This same Application as-filed, lines 19-23 on page 29; Parent application] 11803,104, lines 8-]] on page 23. Amended claim 1 recites a nozzle design that eliminates surface wetting problems found in prior art designs such as Takahashi, Nobile, and Quake. Nothing in those references teaches or suggests the "complete isolation of the nozzle section from the continuous aqueous phase" benefits provided by the invention.
Vogelstein and Dorfman also do not teach or suggest the claimed nozzle design.
Vogelstein is a U.S. publication of a patent application for BEAMing, which refers to Bead Emulsion Amplification with Magnets. Vogelstein, [0006]. Vogelstein does not mention microfluidics whatsoever, much less using positive pressure. In fact, Vogelstein does not even use any of the terms "microfluidics" or "channel" or "chip" or any synonyms thereof.
Dorfman, like Takahashi teaches using aspiration in a microfluidics system. Dorfinan, 370]. There simply isn't any nozzle, much less one that juts out into the oil phase.
For clarity, applicant notes that, among other things, amended claim 1 recites that the immiscible carrier fluid flows INTO the nozzle. Also, the nozzle is never wetted by aqueous phase liquids. The closest thing to a nozzle in the prior art (FIG. 4 in Nobile) shows PCR solution flowing OUT OF the nozzle. Also, the bore of Nobile's nozzle is fully wetted by aqueous PCR solution.
Claim 1 is amended to recite an invention found nowhere among the asserted references. Accordingly, amended claim 1 cannot reasonably be said to be obvious over those references.
Because the only independent claim is not obvious, none of the claims are obvious.
Applicant believes that the rej ections under § 103 are overcome.
Summary
On the basis of the foregoing amendments and reasons, Applicant submits that the pending claims are in condition for allowance, which is respectfully requested.
Dated: October 29, 2021
Respectfully submitted,BROWN RUnNICK LLP

NEXT OFFICE ACTION: DETAILED ACTION
The present Office Action is responsive to the Amendment received on October 29, 2021.
Notice of Pre-AIA  or AIA  Status
The present application is being examined under the pre-AIA  first to invent provisions.
Preliminary Remark
Claims 5-11, 14-20, and 24-30 are canceled.
Information Disclosure Statement
The IDS received on August 17, 2021, October 27, 2021, October 29, 2021, and December 17, 2021 are proper and are being considered by the Examiner.  All IDSs were received with a fee under 37 CFR 1.17(p).
Claim Rejections - 35 USC § 112
The following is a quotation of 35 U.S.C. 112(b):
(b)  CONCLUSION.—The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the inventor or a joint inventor regards as the invention.
The following is a quotation of 35 U.S.C. 112 (pre-AIA ), second paragraph:
The specification shall conclude with one or more claims particularly pointing out and distinctly claiming the subject matter which the applicant regards as his invention.
Claim 35 is rejected under 35 U.S.C. 112(b) or 35 U.S.C. 112 (pre-AIA ), second paragraph, as being indefinite for failing to particularly point out and distinctly claim the subject matter which the inventor or a joint inventor (or for applications subject to pre-AIA  35 U.S.C. 112, the applicant), regards as the invention.
Claim 35 recites the phrase, “the main channel by positive displacement pressure driven flow is applied using a pump”.  There is no longer a proper antecedent basis for the limitation, “positive displacement pressure driven flow” in the main channel from the parent claim 1 as the limitation has been deleted therefrom.
Claim Rejections - 35 USC § 103
The rejection of claims 1-3, 5, 12, 13, 19, 31, and 35 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Quake et al. (US 2002/0058332 A1, published May 16, 2002), made in the Office Action mailed on August 5, 2021 is withdrawn in view of the Amendment received on October 29, 2021.
Specifically, the nozzle disclosed by Takahashi et al. and Quake et al. is not disposed within the immiscible fluid.
The rejection of claim 4 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Quake et al. (US 2002/0058332 A1, published May 16, 2002), as applied to claims 1-3, 12, 13, and 19 above, and further in view of Vogelstein et al. (US 2011/0059435, published March 2011, priority February 7, 2005), made in the Office Action mailed on August 5, 2021 is withdrawn in view of the Amendment received on October 29, 2021.
The rejection of claims 20-23 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Quake et al. (US 2002/0058332 A1, published May 16, 2002), as applied to claims 1-3, 12, 13, and 19 above, and further in view of Nobile et al. (US 2005/0227264 A1, published October 13, 2005, priority January 28, 2004), made in the Office Action mailed on August 5, 2021 is withdrawn in view of the Amendment received on October 29, 2021.
The rejection of claims 32-34 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) in view of Quake et al. (US 2002/0058332 A1, published May 16, 2002), as applied to claims 1-3, 12, 13, and 19 above, and further in view of Dorfman et al. (Analytical Chemistry, 2005, vol. 77, pages 3700-3704), made in the Office Action mailed on August 5, 2021 is withdrawn in view of the Amendment received on October 29, 2021.
Rejection – New Grounds, Necessitated by Amendment
The following is a quotation of pre-AIA  35 U.S.C. 103(a) which forms the basis for all obviousness rejections set forth in this Office action:
(a) A patent may not be obtained though the invention is not identically disclosed or described as set forth in section 102, if the differences between the subject matter sought to be patented and the prior art are such that the subject matter as a whole would have been obvious at the time the invention was made to a person having ordinary skill in the art to which said subject matter pertains. Patentability shall not be negatived by the manner in which the invention was made.
Claims 1-4, 12, 13, 21-23, 31, and 35 under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Link et al. (WO 2004/091763 A2, published October 2004) in view of Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) and Quake et al. (US 2002/0058332 A1, published May 16, 2002).
Link et al. teach a method of generating a plurality of emulsion droplets in a carrier oil, wherein the artisans employ the below configuration (from Figure 11B):
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As seen, the nozzle (the narrow opening) is surrounded by oil which shears off the aqueous reagent flowing from the above.  Therefore, Link et al. teach: a) providing a first aqueous fluid in a first inlet channel; b) providing an immiscible carrier fluid via at least one oil supply line, wherein the flow of the immiscible carrier fluid results in the nozzle being disposed within the oil into a main channel (see the lower larger channel where emulsion droplets are formed).
Link et al. also teach additional channels from which to generate additional droplets which could be coalesced to mix the contents therein:
“method includes a step of producing, in a microfluidic channel, a first droplet comprising a first liquid, where the first droplet is surrounded by a carrying liquid. The method may also include steps of producing a second droplet comprising a second liquid, and allowing the first droplet and the second droplet to coalesce into one combined droplet” (page 3, lines 13-17)
Link et al., while explicitly teaching that the droplets can comprise nucleic acids, such as DNA, do not explicitly teach that the droplets are coalesced for the purpose of performing a PCR reaction.
With regard to claims 2 and 3, the electrode (element 111, see page 34, lines 8-9) that surrounds the nozzle is considered to be a ferrule.
With regard to claims 4 and 23, the nozzle is surrounded by oil and therefore, isolates the nozzle from the aqueous reagent flowing thereto (see above figure).
With regard to claims 21 and 22, the oil supply line provides the constant oil flow and the droplets are formed, wherein because the nozzle is immersed in oil, the aqueous reagent does not wet the nozzle (see above figure).
Link et al. do not teach that a positive displacement pressure driven flow is employed for providing a second aqueous fluid through the second inlet channel.
Takahashi discloses a method of forming droplets for sample preparation for sequencing (“present invention provides methods of using inline automated platform of the invention … to prepare, amplify, detect … and/or optionally select for further analysis (e.g., sequence) genomic material isolated…”, section [0038]), the method comprising:
providing a first aqueous fluid in a first inlet channel, the first aqueous fluid comprising primers (see Figure 1A, the first inlet is channel carrying the primers A/B and Markers A/B);
providing via a second inlet channel, a second aqueous fluid comprising nucleic acid molecules (see Figure 1A, where gene is provided);
providing an immiscible carrier in a main channel in a fluidic chip (see element 6, Figure 1A, where “carrier fluid” is provided; “a sample plug is surrounded by an immiscible nonaqueous fluid (e.g., mineral oil)”, section [0050]), wherein each of the inlet channels is in fluid communication with the main channel (see where the inlet containing the gene and the inlet containing the primer are connected to the channel where the carrier liquid flows (element 5);
coalescing the first aqueous fluid with the second aqueous fluid in the main channel in the fluid chip thereby forming a plurality of droplets surrounded by the immiscible carrier fluid in the main channel (see the gene primer become coalesced into a single emulsion, Figure 1A, area 10; “each sample droplet is further prepared at a junction, e.g., a T-shaped junction (10) to form a sample plug by being mixed with a primer plug comprising amplification reagents (e.g., primer(s), nucleotide, polymerase, etc).”, section [0084]);
applying pressure to flow the immiscible carrier fluid in the main channel (“controlling device … such as pumps, valves … may be used to control the movement and timing of each sample plug and/or DNA sample plug .. such controlling devices are well known to those skilled in the art”, section [0115]); and
annealing one of the primers to one of the nucleic acid molecules within one of the plurality of droplets (“mixed with amplification reagents to form a sample plug, it is drawn along the microfluidic inline reaction channel into an amplification area of a device … only DNA sample plugs will comprise DNA that will be amplified”, section [0085]; “low temperature area (13) may be set to a temperature appropriate to effectuate annealing”, section [0088]) (addressing the limitations of claims 1, 12, and 13).
Quake et al. teach the use of positive pressure driven flow to flow an immiscible oil in a main channel, and generating droplets via first and second inlet channels attached to said main channel, wherein said inlet channels are also taught as being positive pressure driven:
“the device of the invention comprises at least two inlet regions, each connecting to the main channel at a droplet extrusion region.  In particular, the device may comprise a first inlet region in communication with the main channel at a first droplet extrusion region, and a second inlet region in communication with the main channel at second droplet extrusion region.  A fluid containing a first biological material may pass through the first inlet region so that droplets of the fluid containing the first biological materials are sheared into the main channel.  A fluid containing a second biological material may pass through the second inlet region so that droplets of the fluid containing the second biological material are sheared into the main channel.  In various aspects, the droplets of the first material may mix or combine with the droplets of the second biological material, and the first and second biological material may interact with each other upon mixing.” (section [0018])
“An inlet region may have an opening such as in the floor of the microfabricated chip, to permit entry of the sample into a device.  The inlet region may also contain a connector adapted to receive a suitable piece of tubing ….through which a sample may be supplied … Such an arrangement facilitates introducing the sample solution under positive pressure in order to achieve a desired pressure at the droplet extrusion region” (section [0081])
“device of the invention may have an additional inlet region, in direct communication with the main channel … through which a pressurized stream or ‘flow’ of a fluid is introduced into the main channel.  Preferably, this fluid is one which is not miscible with the solvent or fluid of the sample …” (section [0082])
“the invention use pressure drive flow control, e.g., utilizing valves and pumps, to manipulate flow of cells virions, particles, molecules, enzymes or reagents in one or more directions and/or into one or more channels of a microfluidic device” (section [0099])
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was made to combine the teachings of Link et al. with the teachings of Takahashi et al. and Quake et al., thereby arriving at the invention as claimed for the following reasons.
As discussed above, Link et al. already disclose a method of generating a plurality of aqueous droplets present in an immiscible carrier fluid, wherein the nozzle is formed within the immiscible fluid with explicit teachings that droplets can contain nucleic acid molecules, such as DNA, and that two or more droplets can be coalesced to mix the contents therein.
While Link et al. do not explicitly disclose that their method can be applied for providing a DNA and PCR reactants in droplets, wherein the two droplets are coalesced to form a PCR-ready droplet, in view of Takahashi et al. who teach that emulsion droplets comprising DNA and PCR reagents can be generated within an immiscible carrier fluid and coalesced for PCR reaction therein, said one of ordinary skill in the art would have been motivated to apply the teachings of Link et al. so as to perform PCR reaction.  And while Link et al. did not explicitly teach that the carrier fluid, oil is provided via a pump that generates a positive pressure, said one of ordinary skill in the art would have recognized that other prior art known pressure driven system utilized for generating droplets in a channel would have worked equally well.  Such a capability was demonstrated by Quake et al. who also teach generation of emulsion droplets comprising PCR reaction mixture in a flowing oil, wherein the artisans explicitly teach the use of pumps which provide a positive pressure flow to oil and the respective reagents from inlets in order to generate emulsion droplets in the main channel (see above).
Therefore, one of ordinary skill in the art would have been motivated to combine the teachings of Link et al., with the teachings of Takahashi et al. and Quake et al., thereby arriving at the claimed method based on the nozzle configuration of Link et al.
In KSR International Co v. Teleflex Inc, the supreme court stated that, “A person of ordinary skill in the art is also a person of ordinary creativity, not an automation” (82 USPQ2d at 1397) and that “in many cases a person of ordinary skill will be able to fit the teachings of multiple patents together like pieces of a puzzle” and take into account, “the inference and creative steps that a person of ordinary skill in the art would employ” (82 USPQ2d at 1396).
Lastly, it is respectfully submitted that such “fitting” of the teachings would not have been “beyond [the] ... skill [level]” of one of ordinary skill in the art
In KSR, the Supreme Court stated:
“When a work is available in one field of endeavor, design incentives and other market forces can prompt variations of it, either in the same field or a different one.  If a person of ordinary skill can implement a predictable variation, 103 likely bars its patentability.  For the same reason, if a technique has been used to improve one device, and a person of ordinary skill in the art would recognize that it would improve similar devices in the same way, using the technique is obvious unless its actual application is beyond his or her skill.” (page 13, emphasis added).
Therefore, the invention as claimed is deemed prima facie obvious over the cited references.
Claims 32-34 are rejected under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Link et al. (WO 2004/091763 A2, published October 2004) in view of Takahashi et al. (US 2006/0257893 A1, published November 16, 2006) and Quake et al. (US 2002/0058332 A1, published May 16, 2002), as applied to claims 1-4, 12, 13, 21-23, 31, and 35 above, and further in view of Dorfman et al. (Analytical Chemistry, 2005, vol. 77, pages 3700-3704).
The teachings of Link et al., Takahashi et al. and Quake et al. have already been discussed above.
Link et al, in view of Takahashi et al., while teaching performing PCR in aqueous droplets in surrounded by an immiscible oil medium, do not teach every possible such medium or variations known in the art.  Consequently, the artisans do not explicitly teach that the oil is fluorinated (claim 32), or that the oil comprises fluorosurfactant (claims 33 and 34).
Dorfman et al. disclose a method of performing a continuous PCR reaction in an aqueous droplets surrounded by an immiscible oil medium (“[p]erformign continuous flow PCR for high-throughput clinical and quantitative applications requires encapsulating the sample and preventing contamination between them … this can be accomplished in two ways: either the sample can be a droplet entrained in an immiscible solvent …”, page 3700, 2nd column).
Dorfrman et al. disclose the use of fluorinated oil as the immiscible medium when performing the continuous emulsion PCR (“an improvement system for performing continuous flow PCR using droplets in an immiscible fluorinated solvent …”, page 3701, 1st column, 2nd paragraph; “primed with either the bulk fluorinated oil FC-40”, page 3701, 1st column, 3rd paragraph) or oil comprising a fluorosurfactant (“or FC-40 containing various amounts of fluoro alcohol surfactant”, page 3701, 1st column, 3rd paragraph).
It would have been prima facie obvious to one of ordinary skill in the art at the time the invention was made to combine the teachings of Link et al., Takahashi et al. and Quake et al. with the teachings of Dorfman et al., thereby arriving at the invention as claimed for the following reasons.
Dorman et al. discuss potential problems associated with performing emulsion droplet PCR in an immiscible medium:
“wall treatment [of the channel] are necessary to prevent adsorption of DNA to the walls and subsequent transfer between samples … wall contamination problem is a significant drawback … bubbles in the stream may introduce irregularities in the thermal history of the droplets, making the technique less attractive … flowing droplet system does not suffer from either of these drawbacks … However, the initial work on this system suffered from contamination between droplets” (page 3701, 1st column, 1st paragraph).
In solving the drawback so identified, Dorman et al. disclose that immiscible solvent, “prevent[ed] the droplet breakage … thereby eliminating the contamination between droplets” (page 3701, 1st column, 2nd paragraph), producing “adequately reproducible amplification between different drops without any detectable contamination between them” (page 3701, 1st column, 2nd paragraph).
Based on such findings, one of ordinary skill in the art would have been motivated to utilize a fluorinate immiscible oil or oil comprising fluorosurfactants into the teachings of Link et al., Takahashi et al. and Quake et al., for the same recognized benefit of producing reproducible amplification reaction between emulsion droplets as well as preventing contamination therebetween.
Given that Link et al., in combination with Takahashi et al., Quake et al. and Dorfman et al. would have performed emulsion amplification reactions in a flowing oil medium, said one of ordinary skill in the art would have also expected that addition of fluorosurfactants or using fluorinated oil in the method produced form the combination of Link et al. and Takahashi et al. would have yielded the same predictable benefit as observed in Dorman et al.
In KSR, the Supreme Court particularly emphasized “the need for caution in granting a patent based on the combination of elements found in the prior art,” Id. at 415, 82 USPQ2d at 1395, and discussed circumstances in which a patent might be determined to be obvious. Importantly, the Supreme Court reaffirmed principles based on its precedent that “[t]he combination of familiar elements according to known methods is likely to be obvious when it does no more than yield predictable results.” Id. at 415-16, 82 USPQ2d at 1395. The Supreme Court stated that there are “[t]hree cases decided after Graham [that] illustrate this doctrine.” Id.  at 416, 82 USPQ2d at 1395. (1) “In United States v. Adams, . . . [t]he Court recognized that when a patent claims a structure already known in the prior art that is altered by the mere substitution of one element for another known in the field, the combination must do more than yield a predictable result.”
Therefore, the invention as claimed is deemed prima facie obvious over the cited references.
Conclusion
No claims are allowed.
Examiner notes that the embodiment depicted on Figure 3, wherein the single oil supply line is designed around the ferrule of the nozzle so as to circulate the ferrule may advance prosecution over the present art of record.  However, any amendment thereto would necessitate a new search and examination and would require a filing of an RCE.
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 extension fee 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 date of this final action.
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CITED REFERENCE SUMMARIES: Ref US20020058332A1: Claims:
A microfluidic device comprising a main channel, and at least one inlet region in communication with the main channel at a droplet extrusion region.
a main channel, and
at least one inlet region in communication with the main channel at a droplet extrusion region.
The device of claim 1 wherein a stream of an extrusion fluid passes through the main channel.
The device of claim 2 wherein a sample fluid, which is incompatible with the extrusion fluid, passes through the inlet region so that droplets of the second fluid are sheared into the main channel.
The device of claim 2 wherein the extrusion fluid is a non-polar solvent.
The device of claim 4 wherein the extrusion fluid is decane.
The device of claim 5 wherein the extrusion fluid is selected from the group consisting of tetradecane and hexadecane.
The device of claim 3 wherein the sample fluid comprises an aqueous solution.
The device of claim 7 wherein the aqueous solution comprises ultra pure water, TE buffer, phosphate buffer saline or acetate buffer.
The device of claim 3 wherein the sample fluid contains biological material.
The device of claim 9 wherein each of the droplets contains, on average, no more than one particle of the biological material.
The device of claim 10 wherein the biological material comprises viral particles and each of the droplets contains, on average, no more than one viral particle.
The device of claim 9 wherein the biological material comprises molecules of one or more enzymes.
The device of claim 9 wherein the biological material comprises molecules of one or more substrates.
The device of claim 9 wherein the biological material comprises one or more cells.
The device of claim 9 wherein the biological material comprises one or more viral particles.
The device of claim 1 wherein the droplet extrusion region comprises a T-shaped junction between the inlet region and the main channel.
The device of claim 1 further comprising: a detection region within or coincident with at least a portion of the main channel at or downstream from the droplet extrusion region, and a detector associated with the detection region.
a detection region within or coincident with at least a portion of the main channel at or downstream from the droplet extrusion region, and
a detector associated with the detection region.
The device of claim 17 wherein the detector is an optical detector.
The device of claim 17 further comprising: a discrimination region downstream from the detection region, and a flow control system responsive to the detector and adapted to direct droplets through the discrimination region into a branch channel.
a discrimination region downstream from the detection region, and
a flow control system responsive to the detector and adapted to direct droplets through the discrimination region into a branch channel.
The device of claim 1 wherein the main channel resides in a layer of elastomeric material.
The device of claim 20 wherein the elastomeric layer is adjacent to a substrate layer.
The device of claim 1 , further comprising: a first inlet region in communication with the main channel at a first droplet extrusion region; and a second inlet region in communication with the main channel at a second droplet extrusion region.
a first inlet region in communication with the main channel at a first droplet extrusion region; and
a second inlet region in communication with the main channel at a second droplet extrusion region.
The device of claim 22 wherein: a first fluid containing a first biological material passes through the first inlet region, so that droplets of the fluid containing the first biological material are sheared into the main channel; and a second fluid containing a second biological material passes through the second inlet region, so that droplets of the fluid containing the second biological material are sheared into the main channel.
a first fluid containing a first biological material passes through the first inlet region, so that droplets of the fluid containing the first biological material are sheared into the main channel; and
a second fluid containing a second biological material passes through the second inlet region, so that droplets of the fluid containing the second biological material are sheared into the main channel.
The device of claim 23 wherein droplets containing the first biological material mix with droplets of the second biological material.
The device of claim 24 wherein: the first biological material comprises an enzyme, and the second biological material comprises a substrate to the enzyme.
the first biological material comprises an enzyme, and
the second biological material comprises a substrate to the enzyme.
The device of claim 24 wherein the first biological material interacts with the second biological material upon said mixing.
The device of claim 26 wherein the interaction produces a detectable signal.
A device for sorting biological material comprising: (a) a microfabricated substrate having at least one main channel, an inlet region which meets the main channel at a droplet extrusion region, and at least two branch channels meeting at a junction downstream from the droplet extrusion region; (b) a detection region within or coincident with at least a portion of the main channel and associated with a detector; and (c) a flow control system responding to the detector and adapted to direct biological material into a branch channel.
(a) a microfabricated substrate having at least one main channel, an inlet region which meets the main channel at a droplet extrusion region, and at least two branch channels meeting at a junction downstream from the droplet extrusion region;
(b) a detection region within or coincident with at least a portion of the main channel and associated with a detector; and
(c) a flow control system responding to the detector and adapted to direct biological material into a branch channel.
The device of claim 28 wherein: an extrusion fluid passes through the main channel, and a sample fluid, which is incompatible with the first fluid and contains the biological material, passes through the inlet region, so that droplets of the second fluid and containing the biological material are sheared into the main channel.
an extrusion fluid passes through the main channel, and
a sample fluid, which is incompatible with the first fluid and contains the biological material, passes through the inlet region, so that droplets of the second fluid and containing the biological material are sheared into the main channel.
The device of claim 29 wherein the flow control is adapted to direct the droplets into a branch channel.
The device of claim 29 wherein: the extrusion fluid is a non-polar solvent, and the sample fluid is an aqueous solution.
the extrusion fluid is a non-polar solvent, and
the sample fluid is an aqueous solution.
The device of claim 31 wherein the extrusion fluid is decane.
The device of claim 32 wherein the extrusion fluid is selected from the group consisting of tetradecane and hexadecane.
The device of claim 31 wherein the aqueous solution comprises ultra pure water, TE buffer, phosphate buffer saline, or acetate buffer.
The device of claim 29 wherein each of the droplets contains, on average, no more than one particle of the biological material.
The device of claim 35 wherein the biological material comprises viral particles and each of the droplets contains, on average, no more than one viral particle.
The device of claim 29 wherein the biological material comprises one or more viral particles.
A method for sorting biological material, said method comprising: (a) furnishing an extrusion fluid to a main channel of a microfabricated substrate; (b) providing droplets of a sample fluid containing the biological material to the main channel, wherein the sample fluid is incompatible with the extrusion fluid; (c) interrogating the biological material in each droplet for a predetermined characteristic as it passes through a detection region associated with the main channel; and (d) directing the flow of each droplet into a selected branch channel according to the results of the interrogation.
(a) furnishing an extrusion fluid to a main channel of a microfabricated substrate;
(b) providing droplets of a sample fluid containing the biological material to the main channel, wherein the sample fluid is incompatible with the extrusion fluid;
(c) interrogating the biological material in each droplet for a predetermined characteristic as it passes through a detection region associated with the main channel; and
(d) directing the flow of each droplet into a selected branch channel according to the results of the interrogation.
The method of claim 38 wherein the biological material comprises one or more viral particles.
The method of claim 38 wherein: the sample fluid is an aqueous solution; and the extrusion fluid is a non-polar solvent.
the sample fluid is an aqueous solution; and
the extrusion fluid is a non-polar solvent.
The method of claim 40 wherein the aqueous solution comprises ultra pure water, TE buffer, phosphate buffer saline, or acetate buffer.
The method of claim 40 wherein the extrusion fluid is decane.
The method of claim 42 wherein the extrusion fluid is selected from the group consisting of tetradecane and hexadecane.
The method of claim 38 wherein each of the droplets contains, on average, no more than one particle of the biological material.
The method of claim 44 wherein the biological material comprises viral particles, and each of the droplets contains, on average, no more than one viral particle.
The device of claim 27 , wherein the detectable signal is at least one of fluorescence and color change.
The device of claim 23 , wherein the passage of each fluid through an inlet region is independently controlled.
The device of claim 23 , wherein droplets of the first fluid are sheared into the main channel ahead of droplets of the second fluid.
The device of claim 23 , wherein the passage of each fluid is controlled by at least one valve.
The device of claim 47 , wherein the passage of each fluid is controlled by at least one valve.
The device of claim 19 , wherein the flow control comprises the application of radiation pressure.
The device of claim 19 , wherein the flow control comprises an optical beam.
The device of claim 3 , wherein the sample fluid and the extrusion fluid have different refractive indexes.
The device of claim 3 , wherein the refractive index of the sample fluid is lower than the refractive index of the extrusion fluid.
The device of claim 28 , wherein the flow control comprises the application of radiation pressure.
The device of claim 28 , wherein the flow control comprises an optical beam.
The device of claim 29 , wherein the sample fluid and the extrusion fluid have different refractive indexes.
The device of claim 29 , wherein the refractive index of the sample fluid is lower than the refractive index of the extrusion fluid.
The method of claim 38 , wherein the refractive index of the extrusion fluid is different from the refractive index of the sample fluid, and wherein directing step comprises the application of radiation pressure.
The method of claim 59 , wherein the application of radiation pressure comprises moving droplets with an optical beam.
Ref US20060257893A1: Claims:
A microfluidic device, comprising: at least one sipper; at least one fluid reservoir connected to at least one microfluidic inline reaction channel, wherein the at least one microfluidic inline reaction channel runs through a reagent assembly area, an amplification area within a first temperature-controlled area, and a detection area within a second temperature-controlled area; and at least one metal trace for heating of and/or fluid movement within the microfluidic inline reaction channel, wherein detection of amplified DNA products occurs at one or more temperatures.
at least one sipper;
at least one fluid reservoir connected to at least one microfluidic inline reaction channel, wherein the at least one microfluidic inline reaction channel runs through a reagent assembly area, an amplification area within a first temperature-controlled area, and a detection area within a second temperature-controlled area; and
at least one metal trace for heating of and/or fluid movement within the microfluidic inline reaction channel,
wherein detection of amplified DNA products occurs at one or more temperatures.
The microfluidic device of claim 1 , wherein the at least one microfluidic inline reaction channel further comprises a valve downstream of the detection area.
A microfluidic device, comprising: at least one sipper; at least one fluid reservoir connected to at least one microfluidic inline reaction channel, wherein the at least one microfluidic inline reaction channel runs through a reagent assembly area, an amplification area, and a detection area, and wherein the microfluidic inline reaction channel further comprises a valve downstream of the detection area; and at least one metal trace for heating of and/or fluid movement within the microfluidic inline reaction channel.
at least one fluid reservoir connected to at least one microfluidic inline reaction channel, wherein the at least one microfluidic inline reaction channel runs through a reagent assembly area, an amplification area, and a detection area, and wherein the microfluidic inline reaction channel further comprises a valve downstream of the detection area; and
at least one metal trace for heating of and/or fluid movement within the microfluidic inline reaction channel.
An instrument that controls fluid movement within, heating and cooling of, and data acquisition from, the microfluidic device of claim 1 , comprising a cartridge that interfaces between the instrument and the microfluidic device.
An instrument that controls fluid movement within, heating and cooling of, and data acquisition from, the microfluidic device of claim 3 , comprising a cartridge that interfaces between the instrument and the microfluidic device.
An apparatus comprising the microfluidic device of claim 3 and an instrument that controls fluid movement within, heating and cooling of, and data acquisition from, the microfluidic device.
The apparatus of claim 6 , further comprising a cartridge that interfaces between the instrument and the microfluidic device.
A method of determining an organism in a sample, the method comprising the steps of: (a) acquiring the sample; (b) isolating at least one copy of the genomic DNA of the organism, if present in the sample; (c) introducing a first mixture comprising SGP primers, nucleotides, DNA polymerase, and intercalators to the genomic DNA of the organism to form a second mixture; (d) heating the second mixture to a first temperature that will cause the genomic DNA, if present, to denature into a first and second genomic DNA template; (e) cooling the second mixture to a second temperature that will cause the primers to anneal to each genomic DNA template; (f) reheating the second mixture to a third temperature that is between the first and second temperatures to allow the primers to remain annealed to the genomic DNA and the DNA polymerase to elongate nucleic acid polymers originating from the annealed primers; (g) maintaining the third temperature for a first length of time; (h) repeating steps (d)-(g) at least once; (i) repeating steps (d)-(f); (j) maintaining the third temperature for a second length of time equal to about 40-60% of the first length of time; (k) recooling the second mixture to a fourth temperature lower than or equal to that of the second temperature to allow formation of higher-order structures containing intercalators; (l) detecting the resulting higher-order structures; (m) performing melting temperature analysis; (n) detecting a waveform profile; and (o) determining a positive waveform profile from the sample if the sample contained the organism.
(a) acquiring the sample;
(b) isolating at least one copy of the genomic DNA of the organism, if present in the sample;
(c) introducing a first mixture comprising SGP primers, nucleotides, DNA polymerase, and intercalators to the genomic DNA of the organism to form a second mixture;
(d) heating the second mixture to a first temperature that will cause the genomic DNA, if present, to denature into a first and second genomic DNA template;
(e) cooling the second mixture to a second temperature that will cause the primers to anneal to each genomic DNA template;
(f) reheating the second mixture to a third temperature that is between the first and second temperatures to allow the primers to remain annealed to the genomic DNA and the DNA polymerase to elongate nucleic acid polymers originating from the annealed primers;
(g) maintaining the third temperature for a first length of time;
(h) repeating steps (d)-(g) at least once;
(i) repeating steps (d)-(f);
(j) maintaining the third temperature for a second length of time equal to about 40-60% of the first length of time;
(k) recooling the second mixture to a fourth temperature lower than or equal to that of the second temperature to allow formation of higher-order structures containing intercalators;
(l) detecting the resulting higher-order structures;
(m) performing melting temperature analysis;
(n) detecting a waveform profile; and
(o) determining a positive waveform profile from the sample if the sample contained the organism.
The method of claim 8 , wherein the third temperature is maintained for a second length of time about 50% of the first length of time.
The method of claim 8 , wherein the number of times steps (d)-(g) are repeated in step (h) is 20-50 times.
The method of claim 10 , wherein the number of times steps (d)-(g) are repeated in step (h) is 22-24 times.
The method of claim 8 , wherein the method further comprises repeating steps (i)-(j) one or more times prior to step (k).
A method of detecting the absence or presence of an organism in a sample, comprising the steps of: (a) acquiring the sample; (b) isolating at least one copy of the genomic material of the organism, if present in the sample; (c) aspirating at least one sample droplet into a microfluidic reaction channel; (d) forming at least one sample plug by mixing the at least one sample droplet with a primer plug, wherein the primer plug comprises amplification reagents that comprise at least one primer, nucleotides, DNA polymerase, and intercalators; (e) heating the at least one sample plug to a first temperature that will cause each copy of the genomic DNA, if present, to denature into a first and second genomic DNA template; (f) cooling the at least one sample plug to a second temperature that will cause the primers to anneal to each genomic DNA template; (g) reheating the at least one sample plug to a third temperature that is between the first and second temperatures as to allow (1) the primers to remain annealed to the genomic DNA and (2) the DNA polymerase to elongate nucleic acid polymers originating from the annealed primers; (h) maintaining the third temperature for a first length of time; (i) repeating steps (e)-(h) at least once; and (j) detecting the resulting amplified products, wherein at least steps (c)-(j) occur within the apparatus of claim 6 .
(b) isolating at least one copy of the genomic material of the organism, if present in the sample;
(c) aspirating at least one sample droplet into a microfluidic reaction channel;
(d) forming at least one sample plug by mixing the at least one sample droplet with a primer plug, wherein the primer plug comprises amplification reagents that comprise at least one primer, nucleotides, DNA polymerase, and intercalators;
(e) heating the at least one sample plug to a first temperature that will cause each copy of the genomic DNA, if present, to denature into a first and second genomic DNA template;
(f) cooling the at least one sample plug to a se
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