Ex Parte Fathollahi et al

Patent Trial and Appeal Board

May 21, 2014

11397307 (P.T.A.B. May. 21, 2014)

UNITED STATES PATENT AND TRADEMARK OFFICE
__________

BEFORE THE PATENT TRIAL AND APPEAL BOARD
__________

Ex parte BAHRAM FATHOLLAHI, JAVIER A. FARINAS,
ANDREA W. CHOW, and STEPHANE MOURADIAN
__________

Appeal 2012-003437
Application 11/397,307
Technology Center 1600
__________

Before LORA M. GREEN, JEFFREY N. FREDMAN, and
CHRISTOPHER G. PAULRAJ, Administrative Patent Judges.

PAULRAJ, Administrative Patent Judge.

DECISION ON APPEAL
This is an appeal1 under 35 U.S.C. § 134 involving claims to a method
for processing a protein sample in a microfluidic device. The Examiner
rejected the claims on the grounds of obviousness. We have jurisdiction
under 35 U.S.C. § 6(b). We affirm.

1 Appellants identify the Real Party in Interest as Caliper Life
Sciences, Inc. (See App. Br. 3.)
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STATEMENT OF THE CASE
Background
The invention generally relates to methods and apparatuses for the
analysis of proteins. “More particularly, embodiments of the present
invention are directed toward the enzymatic digestion of proteins within a
microfluidic device” (Spec. ¶ 2).
The Claims
Claims 1-16 are on appeal. Independent claim 1, the only
independent claim, is representative and reads as follows:
1. A method of processing a protein sample in a
microfluidic device, the method comprising the steps of:
providing a microfluidic device comprising a fluid network
that includes a sample reservoir, a waste reservoir, a plurality of
parallel channels connecting the sample reservoir to the waste
reservoir, and a plurality of reagent reservoirs that introduce
reagents at various points along each of the plurality of parallel
channels, wherein each of the reagent reservoirs is in fluid
communication with at least two of the plurality of parallel
channels;
placing immobilized trypsin in a portion of each of the
plurality of channels;
placing a sample into the sample reservoir;
placing the reagents required to carry out reduction,
alkylation, and quenching into the reagent reservoirs;
applying a single negative pressure driving force to the
waste reservoir that generates a continuous flow of the sample
from the sample reservoir, through the plurality of channels, to the
waste reservoir, wherein the flow sequentially exposes the sample
to the reduction, alkylation, and quenching reagents before the
sample flows through the immobilized trypsin; and
collecting the sample in the waste reservoir.

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The Issues

The Examiner has rejected the claims as follows:
I. Claims 1, 12, 15, and 16 were rejected under 35 U.S.C.
§ 103(a) as unpatentable over the combination of Kopf-Sill
’185,2 Anderson,3 and Timperman.4
II. Claims 2-5 were rejected under 35 U.S.C. § 103(a) as
unpatentable over the combination of Kopf-Sill ’185, Anderson,
and Timperman as further combined with Gao.5
III. Claims 6-11 were rejected under 35 U.S.C. § 103(a) as
unpatentable over the combination of Kopf-Sill ’185, Anderson,
and Timperman as further combined with Kopf-Sill ’358.6
IV. Claim 13 was rejected under 35 U.S.C. § 103(a) as unpatentable
over the combination of Kopf-Sill ’185, Anderson, and
Timperman as further combined with Wang.7

2 Kopf-Sill et al., US 6,858,185 B1, published Feb. 22, 2005.
3 Anderson, US 2004/0072251A1, published Apr. 15, 2004.
4 Timperman, US 2003/0089605 A1, published May 15, 2003.
5 Gao et al., Integrated Microfluidic System Enabling Protein
Digestion, Peptide Separation, and Protein Identification, 73
ANAL. CHEM. 2648-2655 (2001).
6 Kopf-Sill et al., US 2001/0045358 A1, published Nov. 29, 2001.
7 Wang et al., Integration of immobilized trypsin bead beds for
protein digestion within a microfluidic chip incorporating
capillary electrophoresis separations and an electrospray mass
spectrometry interface, 14 RAPID COMMCNS MASS SPEC. 1377-
1383 (2000).
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V. Claim 14 was rejected under 35 U.S.C. § 103(a) as unpatentable
over the combination of Kopf-Sill ’185, Anderson, and
Timperman as further combined with Wolk.8
The only issue presented on appeal is whether the Examiner presented
a prima facie case of obviousness of independent claim 1 based on the
combination of Kopf-Sill ’185, Anderson, and Timperman. Appellants have
not presented any specific arguments for the dependent claims, but rather
only argue that “any claim depending from a nonobvious claim is also
nonobvious” (App. Br. 15-17). Accordingly, the dependent claims stand or
fall with independent claim 1 because they have not been separately argued.
37 C.F.R. § 41.37(c)(1)(vii) (2011).
FINDINGS OF FACT
FF1. Kopf-Sill ’185 discloses a microfluidic device with a
single vacuum source (Kopf-Sill ’185, Title).
FF2. Kopf-Sill ’185 teaches that “[r]eservoirs or wells are
locations at which samples, components, reagents and the like are
added into the device for assays to take place” and that “[t]he
reservoirs are typically placed so that the sample or reagent is
added into the system upstream from the location at which it is
used” (Kopf-Sill ’185, 10:13-19).
FF3. Kopf-Sill ’185 discloses embodiments of a
microfluidic device that include sample wells (602, 702), a main
channel (604, 704, 804) and multiple flow reduction channels in a
parallel configuration (e.g., 608, 610, 708, 710, 808, 810), reagent

8 Wolk et al., US 2001/0049148 A1, published Dec. 6, 2001.
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reservoirs (e.g., 612, 712, 812) that are each fluidly coupled to two
different channels (e.g., 622, 722, 822)), and a waste well (806)
(Kopf-Sill ’185, Figs. 6-8).
FF4. Kopf-Sill ’185 discloses that “an assay of interest is
optionally performed at all of the various concentrations of sample
material in the flow reduction channel,” and that “reagent usage is
decreased by the decrease in flow rate produced by the flow
reduction channels” (Kopf-Sill ’185, 13:1-6).
FF5. Kopf-Sill ’185 teaches that “movement of fluidic
materials through the microfluidic devices of the invention is
driven by a pressure source” and that “[t]he pressure source is
typically a vacuum source applied at the downstream terminus of
the main channel” (Kopf-Sill ’185, 17:11-15).
FF6. Kopf-Sill ’185 teaches that “[w]aste well 806, 806B
includes a vacuum source at one end of the channel system to draw
the suspension through the channel” (Kopf-Sill ’185, 17:56-58).
FF7. Kopf-Sill ’185 teaches that “[b]y directing fluid
through the flow reduction channels . . ., the flow rate in a
microfluidic device is modulated in response to the addition of
material to the main channel,” and that “[a]n added advantage of
this flow rate modulation is that, in addition to maintaining a
continuous or constant flow rate, subsequent reagent addition need
not be in larger amounts to meet an increased flow rate demand”
(Kopf-Sill ’185, 19:55-59).
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FF8. Kopf-Sill ’185 teaches that “[a]lternatively, reagent
materials are added to the flow reduction channels” (Kopf-Sill
’185, 4:7-8).
FF9. Anderson teaches the digestion of a protein sample to
peptides using the following protocol:
The digestion is carried out by first denaturing the protein
sample (e.g., with urea or guanidine HCl), reducing the
disulfide bonds in the proteins (e.g., with dithiothreitol or
mercaptoethanol), alkylating the cysteines (e.g., by addition
of iodoacetamide), quenching excess iodoacetamide by
addition of more dithiothreitol or mercaptoethanol, and
finally (after removal or dilution of the denaturant) addition
of the selected proteolytic enzyme (e.g. trypsin), followed by
incubation to allow digestion.

(Anderson, ¶ 68.)
FF10. Anderson teaches an embodiment where:
[T]hree subsamples of the plasma peptide digest are
prepared: one undiluted (to which is add[ed] the required
amount of the thyroglobulin labeled monitor peptide, and
those of other proteins in Group 3 abundance class: FIG. 9),
a second diluted about 1,000 fold (Group 2, to which we add
about the same amount of the labeled complement C2
monitor peptide), and a third diluted about 1,000,000 fold
(Group 1, to which is add[ed] the albumin monitor peptide).

(Anderson, ¶104.)
FF11. Anderson “teaches supports with binding agents
which can collect from a sample target peptides and proteins of
varying concentrations” (Anderson, ¶29).
FF12. Timperman teaches a microfluidic system for
proteome analysis in which “the sample processing reagent is a
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protease (such as trypsin) immobilized on a first solid phase
disposed in a portion of the channel” (Timperman, ¶9).
FF13. Timperman teaches that “different channels can
comprise different types or amounts of protease . . . to perform a
plurality of reactions of substantially identical samples (e.g.,
obtained from a single sample plug) in parallel,” and that “the
extent of digestion may be controlled by precisely controlling the
amount of time a sample is exposed to protease to produce larger
peptides or peptides comprising overlapping sequences”
(Timperman, ¶¶ 114-115).
FF14. Timperman teaches that “[t]he immobilized protease
allows the use of high concentrations of enzyme with negligible
production of autolysis products” (Timperman, ¶113).
FF15. Timperman teaches “[i]n still another aspect,
however, flow through one or more selected channels of the device
is hydrodynamic and mediated mechanically through valves placed
at appropriate channel junctions as is known in the art,” and
“[t]herefore, in one aspect, flow of reagents in each of the channels
8 of the device 5 is independently controlled” (Timperman, ¶130).
PRINCIPLES OF LAW
“[I]f 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.” KSR Int'l Co. v. Teleflex Inc., 550
U.S. 398, 417 (2007). The question to ask is “whether the improvement is
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more than the predictable use of prior art elements according to their
established functions.” Id. However, “[o]bviousness does not require
absolute predictability of success ... all that is required is a reasonable
expectation of success.” In re Kubin, 561 F.3d 1351, 1360 (Fed. Cir. 2009)
(citing In re O'Farrell, 853 F.2d 894, 903–04 (Fed. Cir. 1988)).
“A reference may be said to teach away when a person of ordinary
skill, upon reading the reference, would be discouraged from following the
path set out in the reference, or would be led in a direction divergent from
the path that was taken by the applicant.” In re Gurley, 27 F.3d 551, 553
(Fed. Cir. 1994). However, the “mere disclosure of alternative designs does
not teach away.” In re Mouttet, 686 F.3d 1322, 1334 (Fed. Cir. 2012) (citing
In re Fulton, 391 F.3d 1195, 1201 (Fed. Cir. 2004)).
ANALYSIS
As recognized by the Examiner, Kopf-Sill ’185 discloses a method of
processing a sample within a microfluidic device comprising a fluid network
with a plurality of parallel channels connecting a sample reservoir to a waste
reservoir, and a plurality of reagent reservoirs that sequentially introduce
reagents at various points along each of the plurality of parallel channels,
wherein each of the reagent reservoirs is in fluid communication with at
least two of the plurality of parallel channels, and wherein (Ans. 5; FF1-7).
The Examiner acknowledges that Kopf-Sill ’185 fails to specifically teach
that the microfluidic device can be used in a method “for processing proteins
involving exposing the sample to reduction, alkylation, and quenching
reagents sequentially, and that immobilized trypsin is placed in a portion of
each of the channels” (id.).
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The Examiner, however, finds that Anderson teaches the digestion of
proteins by exposing the sample proteins to reduction, alkylation, and
quenching reagents sequentially, “wherein the proteins may be subdivided
into undiluted, diluted 1000 fold, and diluted 10000000 fold subsamples”
(Ans. 6; FF9-10). The Examiner further finds that Anderson teaches that
“tryptic peptides created by trypsin digestion of the protein can be collected”
(Ans. 6; FF11).
Additionally, the Examiner finds that
Timperman further teaches that the microfluidic
device configured for protein digestion may comprise a
plurality of channels, each containing different types or
amounts of a protease such as trypsin immobilized on a solid
phase disclosed in a portion of the channels, to maintain the
trypsin within the channel . . ., so that a desired amount of
digested polypeptide products may be obtained

(Ans. 6; FF12-13).
We believe the Examiner has made a prima facie showing that a
person of ordinary skill in the art would have found it obvious to use the
microfluidic device of Kopf-Sill ’185 to perform trypsin digestion of a
protein sample at different concentrations as disclosed by Anderson, and to
further use immobilized trypsin within the channels as disclosed by
Timperman, which would result in the claimed method. We have considered
Appellants’ arguments, but are not persuaded otherwise. Appellants have
not presented any evidence of secondary (objective) considerations rebutting
the prima facie case of obviousness.
Appellants first argue that their “claimed method does not involve
performing method steps on different concentrations of the same sample
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concurrently,” and that their “purpose in processing a protein sample in a
microfluidic device having a plurality of parallel channels is to increase the
rate at which a sample is processed” (App. Br. 13-14 (emphasis in original).
Appellants assert that reducing the flow of sample, as taught by Kopf-Sill
’185, would “confer no benefit in Appellants’ method” and be
“counterproductive” (id. at 14).
We do not find this argument persuasive because the claimed method
does not preclude processing the protein sample at different concentrations
in the manner taught by Kopf-Sill ’185, even assuming arguendo that doing
so would require a reduced flow. Moreover, the claims do not require a
minimum flow rate for performing the trypsin digestion, nor is there any
suggestion that the device of Kopf-Sill ’185 could not achieve a flow rate
required for trypsin digestion in the manner claimed. Therefore, one of
ordinary skill in the art would have found reason to perform the trypsin
digestion protocol of Anderson in the microfluidic device of Kopf-Sill ’185
in order maintain a continuous flow rate while avoiding the need to add
reagents in larger amounts to meet an increased flow demand (FF7). See
KSR, 550 U.S. at 419-20 (“In determining whether the subject matter of a
patent claim is obvious, neither the particular motivation nor the avowed
purpose of the patentee controls. . . . Under the correct [obviousness]
analysis, any need or problem known in the field of endeavor at the time of
invention and addressed by the patent can provide a reason for combining
the elements in the manner claimed.”).
Appellants argue against the Anderson reference on the grounds that
the reference discloses that “the samples are detected rather than being
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collected in a reservoir” (App. Br. 14 (emphasis in original)). However,
Anderson teaches collecting peptides and proteins of varying concentrations
from the sample target (FF11). One of ordinary skill in the art would
therefore have found it obvious to use the waste reservoirs disclosed by
Kopf-Sill ’185 to collect the digested peptides. Appellants further argue that
Anderson provides no teaching regarding the use of a microfluidic device to
carry out reduction, alkylation, and quenching in the manner claimed (App.
Br. 14). However, Kopf-Sill ’185 discloses a device that is capable of doing
so. One cannot show nonobviousness by attacking references individually
where the rejections are based on combinations of references. In re Merck
& Co., Inc., 800 F.2d 1091, 1097 (Fed. Cir. 1986).
With respect to Timperman, Appellants argue that the reference is
similarly silent with regard to continuous flow and collecting a digested
sample in a reservoir (App. Br. 14-15). Again, those capabilities are taught
by Kopf-Sill ’185 as discussed above. Appellants further argue that
Timperman only “teaches a device that is electrophoretically driven,” and
teaches away from the claimed method by stating that “flow of reagents in
each of the channels 8 of the device 5 is independently controlled” when
discussing hydrodynamic flow (App. Br. 15; FF15). But Timperman does
not teach that hydrodynamic flow in each of the channels must necessarily
be independently controlled when using immobilized trypsin in a
microfluidic device, and we therefore do not view that disclosure in
Timperman as teaching away from the claimed method of using a single
negative pressure driving force to generate continuous flow. Moreover, the
fact that Timperman discloses the use of electrophoretic flow does not teach
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away from the use of a vacuum source. To the contrary, Timperman’s
alternative disclosure of hydrodynamic driven flow would provide reason to
try the single vacuum source device disclosed by Kopf-Sill ’185. See
Mouttet, 686 F.3d at 1334 (The “mere disclosure of alternative designs does
not teach away.”).
Kopf-Sill ’185 discloses that reagent materials can be added to the
flow reduction channels instead of being placed in reservoirs (FF8).
Because Timperman discloses that “immobilized protease allows the use of
high concentrations of enzyme with negligible production of autolysis
products,” one of ordinary skill in the art would have found motivation to
use immobilized trypsin in the flow reduction channels of the Kopf-Sill ’185
device so that so that a desired amount of digested polypeptide products may
be obtained (FF14).
Accordingly, we find that the combined teachings of Kopf-Sill ’185,
Anderson, and Timperman establish a prima face case of obviousness that
Appellants have not rebutted.
SUMMARY
We affirm the rejection of claim 1 under 35 U.S.C. § 103(a) as being
rendered obvious by Kopf-Sill ’185, Anderson, and Timperman. Claims 2-
16 fall with claim 1.

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TIME PERIOD FOR RESPONSE
No time period for taking any subsequent action in connection with
this appeal may be extended under 37 C.F.R. § 1.136(a).

AFFIRMED

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