Office Action 10: Final Rejection (2022-10-31)
Rejection types: 112 - Written Description/Enablement

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 from a first reservoir into a first inlet channel on a microfluidic substrate, 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 immiscible carrier fluid and into a main channel within the nozzle, wherein the nozzle and the main channel are connected to the microfluidic substrate via a self-aligning fluid interconnect molded into the microfluidic substrate, wherein the self-aligning fluid interconnect positions the nozzle within the immiscible carrier fluid and forms a radial seal between the microfluidic substrate and the main channel; 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 wherein the nozzle design inhibits the aqueous sample fluid from wetting surfaces of the nozzle or the main channel.

2\. (Previously Presented) The method of claim 1, wherein the nozzle comprises a ferrule immersed within the immiscible carrier fluid.

3\. (Previously Presented) The method of claim 2, wherein surfaces of the nozzle and ferrule are wetted entirely by the carrier fluid.

4\. (Previously Presented) The method of claim I, wherein the nozzle has complete isolation from any continuous aqueous phase.

5-11. (Cancelled) 610

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

13\. (Previously Presented) 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.

14-20. (Cancelled) 1519

21\. (Previously Presented) The method of claim 1, wherein the oil supply line provides a constant flow of the carrier fluid into the nozzle while the plurality of droplets are formed.

22\. (Cancelled)

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 I, wherein the immiscible carrier fluid comprises an oil.

32\. (Previously Presented) The method of claim 3 I, 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.

35-38. (Cancelled) 3637
Page 2 of 7 Page 3 of 7 Page 4 of 7

OFFICE ACTION:
DETAILED ACTION
The present Office Action is responsive to the Amendment received on September 30, 2022.
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, 22, 24-30, and 35-38 are canceled.
Information Disclosure Statement
The IDS received on September 21, 2022 is improper because Applicants have not submitted the requisite fee under 37 CFR 1.17(p).
The IDS form says that the fee under 37 CFR 1.17(p) has been submitted with said IDS.  However, Applicants’ own electronic submission says that the fee is not submitted with said IDS and there is not authorization statement to charge any deficient fees.
Therefore, said IDS has not been considered for failing to comply with 37 CFR 1.97(c)(1) or (c)(2).
Claim Rejections - 35 USC § 112
The rejection of claim 35 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, made in the Office Action mailed on December 29, 2021 is withdrawn in view of the Amendment received on March 22, 2022, canceling the rejected claim.
Rejection – New Grounds, Necessitated by Amendment
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.
Claims 1-4, 12, 13, 21, 23, and 31-34 are 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 1 recites the limitation, “the nozzle design”. The limitation lacks an antecedent basis and it is unclear just exactly what part of the nozzle’s configuration is being referenced by “the” nozzle design.
Claim 23 is indefinite for the following reasons.
Claim 23 recites that carrier fluid into the nozzle provides complete isolation of the nozzle from the aqueous phase of the first aqueous fluid.
Claim 23 depends from claim 21, which further depends from claim 1.
Claim 1 has been amended to recite that the first aqueous fluid and a second aqueous fluid is “coalesced” to form an aqueous sample fluid and that this fluid is what is being formed as a droplet within the immiscible fluid.  Therefore, claim 23 appears to be duplicative to claim 21.
Claims 2-4, 12, 13, 21, 23, and 31-34 are indefinite by way of their dependency on claim 1.
The following is a quotation of the first paragraph of 35 U.S.C. 112(a):
(a) IN GENERAL.—The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor or joint inventor of carrying out the invention.
The following is a quotation of the first paragraph of pre-AIA  35 U.S.C. 112:
The specification shall contain a written description of the invention, and of the manner and process of making and using it, in such full, clear, concise, and exact terms as to enable any person skilled in the art to which it pertains, or with which it is most nearly connected, to make and use the same, and shall set forth the best mode contemplated by the inventor of carrying out his invention.
Claims 1-4, 12, 13, 21, 23, and 31-34 are rejected under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement. The claim(s) contains subject matter which was not described in the specification in such a way as to reasonably convey to one skilled in the relevant art that the inventor or a joint inventor, or for applications subject to pre-AIA  35 U.S.C. 112, the inventor(s), at the time the application was filed, had possession of the claimed invention. This is a New Matter Rejection.
Claim 1 has been amended to recite, inter alia, the limitation that the “nozzle and the main channel are connected to the microfluidic substrate via a self-aligning fluid interconnect molded into the microfluidic substrate”.
There is insufficient support for such a configuration for the following reasons.
Applicants’ reference made to lines 28-30 of page 4 does not support for this limitation because that section has to do with a general discussion on self-aligning fluidic interconnect to connect the inlet channel for introducing a sample fluid to the channel.  The Office notes that Applicants had introduced this embodiment into the claims in the Amendment dated March 22, 2022, which were rejected.
The presently amended claims canceled that limitation and now require that the main channel through which the monodisperse droplets which are formed and travel through; and nozzle are connected via self-aligning fluid interconnect.  The main channel and the nozzle located downstream from where sample fluid (containing sample and reaction reagents) are provided and therefore, cannot support the present configuration.
Applicants also refer to lines 1-6 of page 43, which discusses a face seal between the microfluidic device and the interconnect.  However, this section is explicit in stating that the seal is for making a “leak-free seal between the external fluid lines and the microfluidic device.”
The main channel and the nozzle are not part of the external fluid lines but are found within the microfluidic device.  The external fluid lines are those which are fed into the microfluidic device, such as reagents and sample fluid lines as discussed above.
PNG
media_image1.png
369
837
media_image1.png
Greyscale
Lastly, Applicants reference Figure 12 for support.  The figure is reproduced below:
In order to understand this figure, page 43 is referenced.  On page 43, the specification states Figure 12 shows that the entrance to the molded hole is large enough to guarantee capture but tapers down to the sealing surfaces.  The section does employ the term, “retaining ferrule”.  However, this ferrule appears to be for referencing the length of the tube which connects to the fluid interconnect:
“The external interconnect is made from a hard but flexible material such as 1/32” PEEK tubing.  The features in the microfluidic device can be molded/machined directly onto the tubing ends or molded as individual pieces and mechanically fastened to the tubing.  The retaining ferrule shown in Figure 12 would be attached during manufacturing and provide good absolute referencing of the tube length.  The ferrule could be an off-the-shelf component …” (page 43, lines 17-23)
Therefore, all of the disclosure points to the fluid interconnect providing a connection of the outside fluid source, such as sample and reaction reagents onto the microfluidic device, prior to their entry onto the microfluidic device and not for connecting the ferrule and main channel within the device.
Therefore, a general disclosure for a specific embodiment of outside fluid connection cannot justify support for the presently claimed configuration.
Applicants are requested to provide where in the specification a support is found where the main channel and the ferrule is connected to the microfluidic substrate via a self-aligning fluid interconnect.
Claim Rejections - 35 USC § 103
The rejection of claims 1-4, 12, 13, 21-23, 31, 36, and 38 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), Quake et al. (US 2002/0058332 A1, published May 16, 2002) and Tai et al. (U.S. Patent No. 6,428,053, issued August 6, 2002), made in the Office Action mailed on July 1, 2022 is withdrawn in view of the Amendment received on September 30, 2022.  There is lack of legal motivation to arrive at the structural configuration of employed microfluidic substrate of the claimed method.
The rejection of claims 32-34 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), Quake et al. (US 2002/0058332 A1, published May 16, 2002) and Tai et al. (U.S. Patent No. 6,428,053, issued August 6, 2002), as applied to claims 1-4, 12, 13, 21-23, 31, 36, and 38 above, and further in view of Dorfman et al. (Analytical Chemistry, 2005, vol. 77, pages 3700-3704), made in the Office Action mailed on July 1, 2022 is withdrawn in view of the Amendment received on September 30, 2022.
The rejection of claim 37 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), Quake et al. (US 2002/0058332 A1, published May 16, 2002) and Tai et al. (U.S. Patent No. 6,428,053, issued August 6, 2002), as applied to claims 1-4, 12, 13, 21-23, 31, 36, and 38 above, and further in view of Inamura et al. (US 2003/0038865 A1, published February 2003), made in the Office Action mailed on July 1, 2022 is withdrawn in view of the Amendment received on September 30, 2022, canceling the rejected claim.
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 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.
Inquiries

APPLICANT REBUTTAL: Remarks
With entry of this amendment, claims 1-4, 12, 13, 21, 23, and 31-34 are pending, with claim 1 being the only independent claim. Claim 1 is amended. Support for the amendments may be found the as-filed application, for example, at least at lines 28-30 on page 4; lines 1-6 on page 43 and in FIG. 12.
No new matter is added.
Rejections under 35 U.
S.
C. 103
The Office Action on page 4 states that claims 1-4, 12, 13, 21-23, 31, 36, and 38 are rejected under §103 over WO 2004/091763 to Link in view of U.S. Pub. 2006/0257893 to Takahashi, U.S. Pub. 2002/0058332 to Quake, and U.S. Pat. 6,428,053 to Tai. On pages 13-14, claims 32-34 are so rejected over Link, Takahashi, and Quake and further in view of Dorfman, 2005, Anal Chem 77:3700-3704. On page 17, claim 37 is so rejected over Link, Takahashi, Quake, Tai, and U.S. Pub. 2003/0038865 to Inamura.
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. 1991). 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 981 (CCPA 1974).
Claim 1 recites coalescing first and second aqueous fluids together on a microfluidic substrate. Pressure is used to flow an immiscible carrier fluid into a nozzle disposed within the immiscible carrier fluid and into a main channel within the nozzle. The nozzle (and the main channel therein) are connected to the substrate by a self-aligning fluid interconnect. That interconnect positions the nozzle within the immiscible carrier fluid and forms a radial seal
between the microfluidic substrate and the main channel. Noting that the oil flows
into
the nozzle & main channel, aqueous droplets are formed within the main channel, which is never wetted by aqueous phase liquid.
What is missing from the prior art is a self-aligning fluid interconnect that positions a nozzle within oil, such that aqueous liquid can flow into the nozzle and from droplets without making contact with the nozzle or the main channel.
Link only reports that continuous or discontinuous fluidic streams may be formed or combined or that such a fluidic stream may be manipulated, e.g., by coalescing droplets. Link, Abstract. Link does not disclose any self-aligning interconnect that positions a nozzle within oil and flows the oil
into
the nozzle to form droplets therein. Additionally, Link is also silent on a design that inhibits aqueous fluid from wetting channel walls.
Takahashi, Quake, Dorfman, and Tai do not teach or suggest any nozzle, immersed in an immiscible carrier fluid, where aqueous droplets are formed by flowing the immiscible carrier fluid into the nozzle.
Takahashi reports a device for preparing and detecting genomic material using 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. Takahashi, [0030], [0040]. Takahashi states that the "sipper" is a capillary that aspirates a sample droplet from e.g., a microtiter plate. Id., [0011]. Quake only reports a device with a inlet regions in communication with a main channel at a droplet extrusion region for introducing droplets of a sample into the main channel. Quake, [0076]. Dorfman reports PCR in droplets flowing in an immiscible solvent. Dorfman, Abstract. Dorfman uses droplets in the immiscible solvent flowing in Teflon capillaries through zones of a compact heating system. Id., 3701. Dorfman requires that a Hamilton PSD/2 pump be used to aspirate droplets from a fluid reservoir into a capillary tube.
Id., 3702. Tai reports a micromachined fluidic coupler. Tai, Abstract. However, Tai is wholly silent on what fluids may be used, does not even distinguish between liquids and gases, and makes no mention whatsoever of aqueous liquids, immiscible carrier fluids, or droplets. Like the other references, Tai is silent on any mechanism to avoid aqueous wetting of the channel, and does not teach or suggest a nozzle protruding into an immiscible carrier fluid wherein that carrier fluid flows
into
the nozzle for droplet formation.
The Office Action on page 4 states that Link in Figure 11B shows a nozzle surrounded by oil, but that is not quite correct. In Link's 11B, there is no nozzle protruding into oil. The "narrow opening" identified in the Office Action is simply surrounded by whatever is outside of Link's device (e.g., the plastic chip or the ambient atmosphere). Moreover, the narrow opening shown in Link forms no part of a self-aligning fluidic interconnect, as recited in amended claim 1. Further, Link is silent as to inhibiting channel aqueous phase wetting by sample fluid.
Because that element of amended claim 1 is wholly missing from the prior art, 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 rejections 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: September 30, 2022
Respectfully submitted,

NEXT 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 January 31, 2023 has been entered.
Preliminary Remark
Claims 2, 5-11, 14-20, 22-30, and 35-38 are canceled.
Claims 39 and 40 are new.
Information Disclosure Statement
The IDS received on October 28, 2022 is proper and is being considered by the Examiner.
Claim Rejections - 35 USC § 112
The rejection of claims 1-4, 12, 13, 21, 23, and 31-34 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, made in the Office Action mailed on October 31, 2022 is withdrawn in view of the Amendment received on January 31, 2023.
The new matter rejection of claims 1-4, 12, 13, 21, 23, and 31-34 under 35 U.S.C. 112(a) or 35 U.S.C. 112 (pre-AIA ), first paragraph, as failing to comply with the written description requirement., made in the Office Action mailed on October 31, 2022 is withdrawn in view of the Amendment received on January 31, 2023.
Rejection – New Grounds, Necessitated by Amendment
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.
Claims 3 and 21 are 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 3 is indefinite because the claim 3 does not appear to further limit the parent claim for the following reasons.
The parent claim, claim 1 has been amended to require that the method employs a nozzle that comprises a ferrule “immersed, by design, within the immiscible carrier fluid sample, thereby inhibiting the aqueous sample fluid from wetting the surfaces of the nozzle or the main channel”.
Claim 3, depending from claim 1, recites that the “surfaces of the nozzle and ferrule are wetted entirely by the carrier fluid.”
Claim 3 no longer further limit its parent claim because the nozzle already comprises a ferrule which is “immersed” in the immiscible fluid.  By immersion, the surfaces of the nozzle and ferrule are wetted entirely by the immiscible fluid.  Therefore, there doesn’t appear to be any further limitation in claim 3.
Claim 21 recites the phrase, “the oil supply line”.  It is unclear which of the, “at least one oil supply line” the oil supply line is referring to.
Amending the phrase to recite, “the at least one oil supply line” would overcome this rejection.
Claim Rejections - 35 USC § 103
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 negated by the manner in which the invention was made.
Claims 1, 3, 4, 12, 13, 21, 31, 39, and 40 are rejected under pre-AIA  35 U.S.C. 103(a) as being unpatentable over Cho et al. (EP 1 584 586 A2, published 2005) in view of Kumacheva et al. (WO 2005/103106 A1, published November 2005, priority April 2004) and Bhagat et al. (Journal of Micromechanics and Microengineering, 2007, vol. 17, pages 42-49).
With regard to claim 1, Cho et al. teach a method of performing nucleic acid amplification reaction (“a method and apparatus for amplifying nucleic acids”, Abstract), the method comprising:
providing a first aqueous fluid from a first reservoir into a first inlet channel on a microfluidic substrate, the first aqueous fluid comprising a biological material (see Fig. 6, Target DNA reservoir 442);
providing via a second inlet channel, a second aqueous fluid comprising primer (see Fig. 6, see primer reservoir 444);
coalescing the first aqueous fluid with the second aqueous fluid to form an aqueous sample fluid (see Fig. 6, channel where the aqueous droplets merge therein prior to entry into main channel);
providing an immiscible carrier fluid via at least one oil supply line (see Fig. 6, see oil 152),
applying a pressure to flow the immiscible fluid into a main channel (“oil 131 … inserted into the reaction channel 130 via the respective inlet channels … 151 at a constant flow rate”, section [0053] evidences the presence of a source of pressure driven flow).
With regard to claims 12 and 13 the aqueous fluids further comprise reagents for conducting an amplification (see Fig. 6, element 446, “for example, polymerase, dNTPs, and buffer”, section [0076]).
With regard to claim 21, the oil flow is constant flow (see section [0053]).
With regard to claim 31, the immiscible fluid is oil (“oil introduction”, section [0076]).
With regard to claims 39 and 40, the microfluidic substrate is a chip that includes a heating and cooling lines (see Figure 7, T1, T2, and T3, wherein the PCR reaction is performed as the droplets travel past the different temperature zones of heating and cooling (also, “substrate 110 has two areas having different temperatures, for example, 65oC area and 95oC area … heating unit 120 may also be comprised of three or more heaters having different temperatures”, section [0050]).
Cho et al. do not disclose that the device comprises a nozzle disposed within the immiscible carrier fluid which leads into the main channel where emulsion droplets enter for nucleic acid amplification reaction, wherein the nozzle comprises a ferrule immersed in oil thereby inhibiting the aqueous droplets to wet the surfaces of the nozzle or the main channel (claims 1 and 3), wherein the nozzle has complete isolation from any continuous aqueous phase (claim 4).
PNG
media_image1.png
212
277
media_image1.png
Greyscale
Kumacheva et al. teach a device for producing droplets within a channel, wherein in an embodiment, aqueous droplets are generated and enters a main channel that is prefilled with immiscible fluid, so as to prevent the aqueous droplets from contacting the main channel and its entry (see below Figure 21b, reproduced below):
“present invention discloses a versatile strategy of synthesis of polymeric particles using a ‘lab on chip’ with pre-designed size shape, morphology and composition” (page 9, lines 18-30)
“Figures 21a and 21b is a schematic illustration of the formulation of droplets by two different mechanism using the microfluidic reactor 120 … Immiscible liquids L1 and L2 (e.g., oil and aqueous phases) are forced into the narrow orifice …” (page 20, lines 23-26)
Bhagat et al. teach a prior art known means of coupling a channel to another channel via a ferrule (see Figure 1).
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 Cho et al. with the teachings of Kumacheva et al. and Benett et al., thereby arriving at the invention as claimed.
As discussed above, Cho et al. already teach a microfluidic device comprising separate reservoirs that provide template nucleic acids and reagents for amplification reaction, which are connected to a main channel flowing with oil.  The resulting configuration produces individual droplets of amplification reaction reagents that are surrounded by flowing oil which are flowed past different sections of the microfluidic chip zones having different temperatures.
While Cho et al. did not explicitly teach that the single channel which coalesces the template nucleic acid and amplification reagents leads into the main channel is completely surrounded by oil, such a configuration has been taught by Kumacheva et al. as shown above that result in the same result of providing an emulsified droplet of precisely controlled size (see page 9, lines 29-35).
As to connecting the main channel to the portion where the aqueous droplets are dispersed (see below left region where droplets are formed and the right region (b) the main channel where emulsion droplets flow through in the figure from
PNG
media_image1.png
212
277
media_image1.png
Greyscale
Kumacheva et al.), use of any such prior known coupling means, such as ferrule would have been an obvious implementation, who use does not provide any perceivable functional advantage over that of Kumacheva 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 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 Cho et al. (EP 1 584 586 A2, published 2005) in view of Kumacheva et al. (WO 2005/103106 A1, published November 2005, priority April 2004) and Bhagat et al. (Journal of Micromechanics and Microengineering, 2007, vol. 17, pages 42-49) as applied to claims 1, 3, 4, 12, 13, 21, 31, 39, and 40 above, and further in view of Dorfman et al. (Analytical Chemistry, 2005, vol. 77, pages 3700-3704).
The teachings of Cho et al., Kumacheva et al. and Bhagat et al. have been discussed above.
Cho et al., Kumacheva et al. and Bhagat et al. do not teach that the immiscible fluid oil should be fluorinated (claim 32) or comprise a fluorosurfactant (claims 33 and 34).
Dorfman et al., much like Cho et al., disclose a method of performing a continuous PCR reaction in aqueous droplets surrounded by immiscible oil (“[p]erforming continuous flow PCR for high-throughput clinical and quantitative applications require 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).
Dorfman 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 Cho et al., Kumacheva et al. and Bhagat et al. with the teachings of Dorfman et al., thereby arriving at the invention as claimed for the following reasons.
Dorfman 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 this drawback, Dorfman 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 fluorinated immiscible oil or oil comprising fluorosurfactants into the teachings of Cho et al., Kumacheva et al. and Bhagat et al., for the same recognized benefit of producing reproducible amplification reaction between emulsion droplets as well as preventing contamination therebetween.
Given that Cho et al., in combination with Kumacheva et al. performed emulsion amplification reaction in a flowing oil medium, said one of ordinary skill in the art would have also expected that the addition of fluorosurfactants or using fluorinated oil in the method of Cho et al./Kumacheva et al. would have yielded the same predictable benefit as observed by Dorfman 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

CITED REFERENCE SUMMARIES: Ref WO2004091763A2: Claims:
A method, comprising: producing a fluidic stream comprising a first fluid surrounded by a second, liquid fluid by expelling the first fluid from an outlet of a channel, wherein the fluidic stream has an average cross-sectional dimension perpendicular to its direction of travel that is smaller than an average cross-sectional dimension ofthe outlet ofthe channel.
The method of claim 1 , wherein the fluidic stream comprises droplets.
The method of claim 2, wherein, in at least some ofthe droplets, the second fluid completely surrounds the first fluid.
The method of claim 1 , wherein the fluidic stream is continuous.
The method of claim 1, wherein the average cross-sectional dimension ofthe outlet ofthe channel is less than about 5 mm.
The method of claim 1, wherein the average cross-sectional dimension ofthe outlet ofthe channel is less than about 1 mm.
The method of claim 1, wherein the average cross-sectional dimension ofthe outlet ofthe channel is less than about 100 microns.
The method of claim 1, wherein the average cross-sectional dimension ofthe outlet ofthe channel is less than about 10 microns.
The method of claim 1, wherein the first fluid is liquid.
The method of claim 1 , wherein at least a portion of the fluidic stream has an electric charge.
The method of claim 1 , further comprising hardening the fluidic stream to form a solid article.
The method of claim 1 , wherein the second, liquid fluid is expelled from an outlet of a second channel.
The method of claim 12, wherein the second channel contains the first channel.
The method of claim 1, wherein the fluidic stream is directed through a dimensional restriction.
The method of claim 1 , further comprising surrounding at least a portion ofthe fluidic stream with a sheathing fluid.
The method of claim 15, wherein the sheathing fluid decreases the average cross- sectional dimension ofthe fluidic stream, relative to the average cross-sectional dimension ofthe fluidic stream in the absence ofthe sheathing fluid.
The method of claim 1 , further comprising applying an electric field to at least a portion ofthe fluidic stream.
An apparatus, comprising: a channel; a reduction system constructed and arranged to reduce an average cross-sectional dimension of a fluidic stream exiting an outlet ofthe channel; and a second channel constructed and arranged to surround at least a portion ofthe fluidic stream with a second, liquid fluid.
The apparatus of claim 18, wherein the reduction system comprises an electric field generator.
The apparatus of claim 18, wherein the reduction system comprises a dimensional restriction.
The apparatus of claim 18, wherein the reduction system comprises the second, liquid fluid.
The apparatus of claim 18, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 5 mm.
The apparatus of claim 18, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 1 mm.
The apparatus of claim 18, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 100 microns.
The apparatus of claim 18, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 10 microns.
A method, comprising: surrounding a first fluid with a second fluid by action at least in part of an electric field.
The method of claim 26, wherein the surrounding step comprises encapsulating the first fluid in the second fluid using an electric field.
The method of claim 26, wherein the surrounding step comprises encapsulating the first fluid in the second fluid using a sheathing fluid.
The method of claim 26, wherein the surrounding step comprises encapsulating the first fluid in the second fluid to form a droplet.
The method of claim 29, wherein the droplet has an average cross-sectional dimension of less than about 5 mm.
The method of claim 29, wherein the droplet has an average cross-sectional dimension of less than about 1 mm.
The method of claim 29, wherein the droplet has an average cross-sectional dimension of less than about 100 microns.
The method of claim 29, wherein the droplet has an average cross-sectional dimension of less than about 10 microns.
The method of claim 26, wherein the surrounding step comprises encapsulating the first fluid in the second fluid to form a tube.
The method of claim 26, wherein the first fluid is gaseous.
The method of claim 35, wherein the first fluid comprises air.
The method of claim 26, wherein the first fluid is liquid.
The method of claim 37, wherein the first fluid comprises water.
The method of claim 26, wherein the second fluid is liquid.
The method of claim 39, wherein the second fluid comprises water.
The method of claim 39, wherein the second fluid has a viscosity of at least 1.2 cp.
The method of claim 39, wherein the second fluid has a viscosity of at least 2 cp.
The method of claim 39, wherein the second fluid has a viscosity of at least 5 cp.
The method of claim 39, wherein the second fluid has a viscosity of at least 10 cp.
The method of claim 39, wherein the second fluid has a viscosity of at least 50 cp.
The method of claim 39, wherein the second fluid has a viscosity of at least 100 cp.
The method of claim 26, further comprising encapsulating the first fluid and the second fluid in a third fluid.
The method of claim 47, further comprising encapsulating the first fluid, the second fluid, and the third fluid in a fourth fluid.
The method of claim 26, wherein the second fluid comprises substantially aligned molecules.
The method of claim 26, further comprising hardening the first fluid.
The method of claim 26, further comprising hardening the second fluid.
An apparatus, comprising: a first channel having an outlet for expelling a first fluid; a second channel having an outlet for expelling a second fluid, the second channel positioned to allow the second fluid to surround at least a portion ofthe first fluid at a predetermined location; and an electric field generator positioned to generate an electric field proximate the predetermined location.
The apparatus of claim 52, wherein the second channel is positioned to allow the second fluid to completely surround droplets ofthe first fluid.
The apparatus of claim 52, wherein the second channel is position to allow the second fluid to surround a tube ofthe first fluid.
The apparatus of claim 52, wherein the second channel contains the first channel.
The apparatus of claim 52, further comprising a dimensional restriction proximate the predetermined location.
The apparatus of claim 52, wherein an average cross-sectional dimension ofthe outlet of the first channel is less than about 5 mm.
The apparatus of claim 52, wherein an average cross-sectional dimension ofthe outlet of the first channel is less than about 1 mm.
The apparatus of claim 52, wherein an average cross-sectional dimension ofthe outlet of the first channel is less than about 100 microns.
The apparatus of claim 52, wherein an average cross-sectional dimension ofthe outlet of the first channel is less than about 10 microns.
A method, comprising: producing a fluidic stream of droplets, the droplets having an average diameter of less than about 60 microns and a distribution of diameters such that no more than about 5% ofthe droplets have a diameter greater than about 10% ofthe average diameter.
The method of claim 61, wherein no more than about 5% ofthe droplets have a diameter greater than about 5% ofthe average diameter.
The method of claim 61 , wherein no more than about 5% of the droplets have a diameter greater than about 3% ofthe average diameter.
The method of claim 61 , wherein no more than about 5% of the droplets have a diameter greater than about 1% ofthe average diameter.
The method of claim 61, wherein no more than about 5% ofthe droplets have a diameter greater than about 0.1% of the average diameter.
The method of claim 61, wherein no more than about 5% ofthe droplets have a diameter greater than about 0.01% ofthe average diameter.
The method of claim 61, wherein no more than about 3% ofthe droplets have a diameter greater than about 10%> ofthe average diameter.
The method of claim 61, wherein no more than about 1% ofthe droplets have a diameter greater than about 10% ofthe average diameter.
The method of claim 61 , wherein no more than about 0.1 % of the droplets have a diameter greater than about 10% ofthe average diameter.
The method of claim 61, wherein no more than about 0.01% ofthe droplets have a diameter greater than about 10% ofthe average diameter.
-71. The method of claim 61 , wherein the fluidic stream of droplets is produced from a channel.
The method ' of claim 61, wherein the droplets have an average diameter of less than about 50 microns.
The method of claim 61 , wherein the droplets have an average diameter of less than about 40 microns.
The method of claim 61 , wherein the droplets have an average diameter of less than about 25 microns.
The method of claim 61, wherein the droplets have an average diameter of less than about 10 microns.
The method of claim 61 , wherein the droplets have an average diameter of less than about 5 microns.
The method of claim 61, wherein the droplets have an average diameter of less than about 1 microns.
The method of claim 61 , wherein the droplets have an average diameter of less than about 0.3 microns.
The method of claim 61 , wherein the droplets have an average diameter of less than about 0.1 microns.
The method of claim 61 , wherein the droplets have an average diameter of less than about 0.01 microns.
The method of claim 61 , further comprising adding at least a portion of the fluidic stream of droplets to a liquid.
An apparatus, comprising: a channel having an outlet, through which a fluid can be expelled, the channel being positioned to allow the fluid to be directed at a reduction system; wherein the reduction system is constructed and arranged to produce, from the  fluid, a fluidic stream of droplets having an average diameter of less than about 60 microns and a distribution of diameters such that no more than about 5% ofthe droplets have a diameter greater than about 10% ofthe average diameter.
The apparatus of claim 82, wherein the reduction system comprises an electric field generator.
The apparatus of claim 82, wherein the reduction system comprises a dimensional restriction.
The apparatus of claim 82, further comprising a second channel positioned to allow the fluid to be encapsulated by a sheathing fluid.
The apparatus of claim 82, further comprising a second channel positioned to allow at least some ofthe droplets to be encapsulated by a sheathing fluid.
The apparatus of claim 82, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 5 mm.
The apparatus of claim 82, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 1 mm.
The apparatus of claim 82, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 100 microns.
The apparatus of claim 82, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 10 microns.
A method comprising: reducing a cross-sectional dimension of a fluidic stream exiting an outlet of a channel using an electric field.
The method of claim 91 , wherein the fluidic stream is continuous.
The method of claim 91 , wherein the fluidic stream comprises droplets.
The method of claim 91 , wherein the reducing step comprises reducing the cross- sectional dimension by producing a Taylor cone.
The method of claim 91, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 5 mm.
The method of claim 91, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 1 mm.
The method of claim 91, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 100 microns.
The method of claim 91, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 10 microns.
The method of claim 91 , wherein the fluid exiting the channel comprises at least one component having a viscosity of at least about 1.2 cp.
The method of claim 91 , further comprising surrounding at least a portion of the fluidic stream with a sheathing fluid.
The method of claim 91 , further comprising hardening at least a portion of the fluidic stream.
An apparatus, comprising: a channel having an outlet; and  an electric field generator constructed and arranged to reduce a cross-sectional dimension of a fluid exiting the outlet ofthe channel.
The apparatus of claim 102, wherein the electric field generator is constructed and arranged to allow the fluid to produce a Taylor cone.
The apparatus of claim 102, further comprising a second channel positioned to allow the fluid to be encapsulated by a sheathing fluid.
The apparatus of claim 102, wherein an average cross-sectional dimension of the outlet of the channel is less than about 5 mm.
The apparatus of claim 102, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 1 mm.
The apparatus of claim 102, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 100 microns.
The apparatus of claim 102, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 10 microns.
A method, comprising: encapsulating a cell in a fluid droplet.
The method of claim 109, wherein the fluid has a cross-sectional dimension of less than about 1 mm.
The method of claim 109, wherein an average cross-sectional dimension of the outlet of the channel is less than about 100 microns.
The method of claim 109, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 10 microns.
The method of claim 109, further comprising encapsulating the fluid in a second fluid.
The method of claim 109, wherein the encapsulating step comprises encapsulating the cell in a droplet.
The method of claim 109, wherein the encapsulating step comprises encapsulating the cell in the fluid droplet using an electric field.
The method of claim 109, wherein the encapsulating step comprises encapsulating the cell in the fluid droplet using a Taylor cone.
The method of claim 109, wherein the encapsulating step comprises charging the cell with an electric charge and charging the fluid with an opposite electric charge.
A method, comprising: producing a fluid Taylor cone within a liquid.
The method of claim 118, wherein the producing step comprises: inducing a charge in a first fluid exiting an outlet of a channel sufficient to cause the first fluid to form a Taylor cone; and sheathing the Taylor cone within a second, sheathing liquid.
The method of claim 119, further comprising: stabilizing the Taylor cone by passing the first fluid through a dimensional restriction downstream ofthe outlet ofthe channel.
The method of claim 120, comprising passing at least one of the first fluid and the second, sheathing liquid through the dimensional restriction.
The method of claim 121, wherein the dimensional restriction has a cross-sectional dimension smaller than an inner diameter ofthe channel outlet.
The method of claim 121, comprising passing both the first fluid and the second, sheathing liquid through the dimensional restriction.
The method of claim 118, wherein the first fluid comprises a liquid having a viscosity greater than about 1.2 cp.
The method of claim 118, further comprising allowing the Taylor cone to produce a fluidic stream of droplets.
The method of claim 118, furtlier comprising allowing the Taylor cone to produce a tube of fluid.
An apparatus, comprising: a channel having an outlet; an electric field generator positioned to induce an electric field in proximity ofthe outlet ofthe channel; and a dimensional restriction having a cross-sectional dimension smaller than an average cross-sectional dimension ofthe outlet ofthe channel, the dimensional restriction in fluidic communication with the channel.
The apparatus of claim 127, wherein the dimensional restriction is substantially aligned with the channel.
The apparatus of claim 127, wherein the electric field generator is positioned to create a Taylor cone in a fluid exiting the channel outlet.
The apparatus of claim 127, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 5 mm.
The apparatus of claim 127, wherein an average cross-sectional dimension of the outlet of the channel is less than about 1 mm.
The apparatus of claim 127, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 100 microns.
The apparatus of claim 127, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 10 microns.
An apparatus, comprising: a channel having an outlet; an electric field generator positioned to create a Taylor cone in a fluid exiting the outlet ofthe channel; and a stabilization system able to restrict motion ofthe Taylor cone.
The apparatus of claim 134, wherein the stabilization system comprises a dimensional restriction.
The apparatus of claim 134, wherein the stabilization system is substantially aligned with the channel.
The apparatus of claim 134, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 5 mm.
The apparatus of claim 134, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 1 mm.
The apparatus of claim 134, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 100 microns.
The apparatus of claim 134, wherein an average cross-sectional dimension ofthe outlet of the channel is less than about 10 microns.
An apparatus, comprising: a first channel having an outlet; a second channel having an outlet; and an electric field generator positioned to create a first Taylor cone in a first fluid exiting the outlet ofthe first channel and a second Taylor cone in a second fluid exiting the outlet ofthe second channel.
The apparatus of claim 141 , wherein the second Taylor cone contains the first Taylor cone.
The apparatus of claim 141, further comprising a third channel having an outlet, wherein the electric field generator is positioned to create a third Taylor cone in a third fluid exiting the outlet ofthe third channel.
The apparatus of claim 143, wherein the third Taylor cone contains the first Taylor cone and the second Taylor cone.
The apparatus of claim 141, further comprising a dimensional restriction that at least one ofthe first fluid and the second fluid passes through.
The apparatus of claim 145, wherein the dimensional restriction is substantially aligned with at least one ofthe first channel and the second channel.
The apparatus of claim 141, wherein an average cross-sectional dimension ofthe outlet of at least one ofthe first channel and the second channel is less than about 5 mm.
The apparatus of claim 141, wherein an average cross-sectional dimension ofthe outlet of at least one ofthe first channel and the second channel is less than about 1 mm.
The apparatus of claim 141, wherein an average cross-sectional dimension ofthe outlet of at least one ofthe first channel and the second channel is less than about 100 microns.
The apparatus of claim 141, wherein an average cross-sectional dimension ofthe outlet of at least one ofthe first channel and the second channel is less than about 10 microns.
A method, comprising : producing a first fluid Taylor cone and a second fluid Taylor cone within the first fluid Taylor cone.
The method of claim 151, wherein the first fluid Taylor cone is liquid.
The method of claim 152, wherein the first fluid Taylor cone comprises water.
The method of claim 151, wherein the second fluid Taylor cone is gaseous.
The method of claim 154, wherein the second fluid Taylor cone comprises air.
The method of claim 15
[Truncated to input limit]